title: O-GlcNAcylation Modulates Expression and Abundance of N-Glycosylation Machinery in an Inherited Glycosylation Disorder

city: Greenwood

first: Heather

last: Flanagan-Steet

state: SC

address: 101 Gregor Mendel circle

country: USA

institute: Greenwood Genetic Center

summary: Much of the N-glycosylation machinery was identified decades ago, but the regulatory mechanisms influencing normal and disease states are only beginning to be discovered. Recent studies highlight a role for key metabolites and metabolic enzymes in this process, revising the view of the relationship between different metabolic pathways. Using a multi-omic approach in a zebrafish, we discovered a mechanism where O-GlcNAcylation directly influences the expression and abundance of two enzymes essential for the earliest steps of N-linked glycosylation, the nus1-encoded NgBR and Dpagt1. Here we show that phosphomannomutase (pmm2) deficiency is associated with increased levels of UDP-GlcNAc and protein O-GlcNAcylation. Biochemical studies show that O-GlcNAc modification of the NgBR and Dpagt1 increases their abundance in pmm2m/m mutants. Modulating O-GlcNAc levels, NgBR abundance or Dpagt1 activity exacerbated cartilage phenotypes in pmm2 mutants, suggesting that O-GlcNAc mediated increases in the N-glycosylation machinery protects against more severe pathology.  These findings highlight nucleotide-sugar donors as metabolic sensors that coordinate two different glycosylation pathways, demonstrating how their interplay is relevant to disease outcome in the most common congenital disorder of glycosylation.

overall_design: We did bulk total RNA-seq with ribo-depletion of whole zebrafish larvae at 2 ages- 5dpf and 8dpf for two genotypes- wildtype WT and Mutant where the mutant is a loss of function of Pmm2. Each sample consisted of 15 larvae and we  sequenced 5 biological replicates per genotype and age combination.

first: Sneha

last: Mokashi


title: Nuclear translocation of TfR1 facilitating tumor progression via p53-mediated gene regulation [RNA-seq]

status: Public on Dec 27 2024

city: Beijing

first: Qizhi

last: Wang

address: Chaoyang District

country: China

institute: Chinese Academy of Sciences

type: Expression profiling by high throughput sequencing

summary: It is well-known that p53-mediated gene regulation exerts a profound impact on the occurrence, progression, metastasis, and various other facets of tumor. Wherein, activation of p53 downstream DNA damage repair-related genes is able to promote the survival of cancer cells during tumor progression.Notably, it has been substantiated that the transcriptional activity of p53 could be modulated by interactions with other proteins.Therefore, we asked whether the interaction with TfR1 was relevant to the transcriptional regulation of p53 target genes. Our results support the positive regulation of nuclear TfR1 on DNA damage repair-related genes, especially for the NER pathway.

overall_design: In order to investigate the regulation of TfR1 on genes in HCT-116 cells, we detected the transcripts before and after knocking down TfR1, and added cisplatin stimulation on this basis, and collected cells for transcript detection

first: Yaxin

last: Hou


title: Ctbp2 and histone modification ChIP-seq profiles in DNA-damaged stem cells

city: Seoul

first: Han-Teo

last: Lee

address: Daehakro 103, Jongro

department: College of Medicine

country: South Korea

institute: Hong-Duk Youn

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Cells are exposed to both intrinsic and extrinsic DNA damage stresses in various environmental conditions, necessitating appropriate control mechanisms. Stem cells, characterized by their pluripotency and self-renewal capabilities, must regulate these mechanisms more rigorously due to their high hierarchical status. Previous studies have suggested that stem cells induce differentiation to prevent the propagation of inappropriate DNA damage. However, this approach paradoxically results in the loss of stemness when the DNA damage is repaired. In this study, we aim to demonstrate that stem cells possess mechanisms to protect their stemness in response to DNA damage. To this end, we performed chromatin immunoprecipitation sequencing (ChIP-seq) for Ctbp2, a well-known regulator of stemness escape, and various histone markers.

overall_design: Chromatin immunoprecipitation DNA sequencing (ChIP-seq) for H3K27Ac and Ctbp2 in DMSO- or 500 nM adriamycin-treated mouse embryonic stem cells (E14)

first: Hong-Duk

last: Youn


title: Epigenomic landscape of human cumulus cells in premature ovarian insufficiency using single-base resolution methylome and hydroxymethylome. [RNA-Seq]

city: Xi'an

first: Wenhao

last: Shi

state: Shaanxi

address: Houzai Gate 73#

department: The assisted Reproduction Center

institute: Northwest Women’s and Children’s Hospital

summary: Premature ovarian insufficiency (POI) has recently been reported to be linked with epigenetic changes. Previous studies have focused on the regulation of individual genes associated with ovarian function through single-gene epigenetic variations, however, there is a deficiency in the comprehensive comprehension of the epigenetic profile for POI. Therefore, we conducted a multi-omics study integrating methylation, hydroxymethylation, and transcriptome sequencing analyses in cumulus cells from women with POI and their matched controls. Our results showed significantly higher global methylation and hydroxymethylation levels in POI women compared to controls. The number of hyper-methylated/hyper-hydroxymethylated regions exceeded hypo-methylated/hypo-hydroxymethylated regions in differentially methylated and hydroxymethylated regions across chromosomes and genetic elements. Methylation within genebodies remained high and exhibited a slight negative correlation with gene expression. Variation in methylation levels was notable among promoter regions, with gene expression decreasing as methylation levels increased (displaying a pronounced negative correlation). Hydroxymethylation levels in both the genebody and promoter regions remained consistently low and showed no overall association with gene expression. Furthermore, we found that changes in methylation were linked to the epigenetic age clock rather than being specifically associated with POI causative genes or ovarian function genes. Our study sheds light on critical features of POI, characterized by widespread DNA hyper-methylation and hyper-hydroxymethylation across the genome. The observed relationship between hyper/hypo-methylation and gene expression regulation may provide valuable insights for identifying potential epigenetic biomarkers and treatment targets for POI.

overall_design: We conducted WGBS-seq, ACE-seq, and RNA-seq analyses on six samples obtained from cumulus cells of three women with POI and three age- and BMI-matched control women.

first: Dongyang

first: Juanzi

first: Chen

last: Huang


title: Effect of porcine spermatogonia germ cells on de novo morphogenesis of testis trough xenotransplatation.

city: Gwangjin-gu

first: Min-Gi

last: Han

state: Seoul

address: Neungdong-ro 120

department: 1Department of Stem Cells and Regenerative Technology

institute: Konkuk University

summary: This study explores the role of porcine spermatogonia germ cells and testicular fibroblast in testis development and spermatogenesis through xenotransplantation. Previous research demonstrated the importance of Sertoli cells in maintaining testicular structure and function during testis development. However, the contribution of spermatogonia to these processes remains unclear. H&E staining revealed that FO tissues, no testicular structures were formed; instead, heterogeneous tissues with simple tubular formations were observed. Conversely, in GF tissues, structures resembling testicular tissue developed, displaying various stages of testis development including elongated spermatid. Immunofluorescence staining showed seminiferous tubules expressing PGP 9.5, VASA, and ACR2, confirming the presence of spermatogenesis. Interestingly, FO tissues were dominantly expressed ESR1, indicating bipotent gonadal property. RNA-seq analysis further revealed that GF tissue showed spermatogenesis-linked GOBPs with gene expression of mature Sertoli cells, including NR5A1, AR, and DMRT1. However, FO tissues displayed early tissue formation processes with expression of BMP4, WNT5A, ESR1, TGFBR3 and ACVR, generally inhibited in normal testis. These findings highlight the critical role of germ cells in formation of testicular structures through xenotransplantation and suggest the potential for spermatogenesis within these structures. However, further research is needed to verify the integrity and fertilizing capacity of the sperm, as these aspects have not been confirmed in this study.

overall_design: To address this, both germ cells and fibroblasts (GF) and fibroblast only (FO) were transplanted to investigate whether the presence of pSGCs influences testicular tissue remodeling and development. After a 6-month period,  RNA-seq analyses were conducted on reconstitution tissues.

first: Hyuk

last: Song


title: Effect of miR-662 and miR-24-2-5p overexpressions on gene expression in human MDA-MB-231-luc2-NW1 (NW1) and MCF7 cell lines.

city: Genova

first: Davide

last: Ceresa

state: APO/FPO EUROPE / ATLANTIC

address: Largo R. Benzi, 10

department: Cellular Oncology

country: Italy

institute: IRCCS Ospedale Policlinico San Martino

summary: MicroRNAs (miRNAs), small non-coding RNAs, are powerful and extremely versatile regulators of the gene expression in cells, playing an important role in every step of cancer progression. Here, we report a list of transcripts regulated by miR-662 and miR-24-2-5p transient overexpressions using miRNA MIMIC transfection (MIMIC-miR-662 and MIMIC-miR-24-2-5p, respectively) compared to control (MIMIC-negCTRL-FAM+) at 36 hours post-transfection in human MDA-MB-231-luc2-NW1 (NW1) (for miR-662 and miR-24-2-5p) and MCF7 (only for miR-24-2-5p) breast cancer cell lines. We found 38 transcripts significantly deregulated (17 downregulated, 21 upregulated) in miR-662-overexpressing NW1 cells compared to mock-transfected cells; 222 (187 downregulated, 35 upregulated) and 34 (32 downregulated, 2 upregulated) transcripts deregulated in miR-24-2-5p-overexpressing NW1 and MCF7 cells, respectively, compared to mock-transfected cells. Furthermore, a total of 30 transcripts (29 downregulated, 1 upregulated) were shared between miR-24-2-5p-overexpressing NW1 and MCF7 cells compared to mock-transfected cells. For miR-662, which we found to promote breast cancer metastasis by stimulating cancer cell stemness (Puppo et al., 2023), we observed the deregulation of Cell Migration Inducing Hyaluronidase 1 (CEMIP), Interleukin 6 receptor (IL6R), High Mobility Group AT-Hook 2 (HMGA2), Small Ubiquitin Like Modifier 3 (SUMO3), and Polymerase (DNA) Delta Interacting Protein 2 (POLDIP2), which reinforced the evidence of the involvement of miR-662 in the acquisition of a stem-like phenotype by human NW1 breast cancer cells and miR-662 contribution to breast cancer bone metastasis formation. For miR-24-2-5p, we found that the downregulation of two transcripts, WD Repeat and FYVE Domain Containing 1(WDFY1) and SH3 Domain Binding Glutamate Rich Protein Like 2 (SH3BGRL2), might explain a possible role of miR-24-2-5p as a tumour suppressor by reducing breast cancer cell invasive properties.

overall_design: To investigate the functional role of two miRNAs, miR-662 and miR-24-2-5p, in the metastatic progression of breast cancer cells to distant organs, particularly to bone, we transiently overexpressed them in human breast cancer cell lines (MDA-MB-231-luc2-NW1 a.k.a. NW1, MCF7), and we then analysed whole transcriptomic changes at 36 hours post-transfection by RNA sequencing. MiR-662 and miR-24-2-5p transient overexpressions were obtained by miRNA MIMICs (50nM) transfection (MIMIC-miR-662 and MIMIC-miR-24-2-5p, respectively) using Lipofectamine 2000 as transfection agent. Three independent transfections (N=3) were performed for each condition. Gene expression profiling analysis was conducted by using data obtained by the RNA sequencing on two different cell lines (NW1, MCF7), where sample triplicates have been collected at 36 hours post-transfection. Each experimental condition (NW1/MIMIC-miR-662, NW1/MIMIC-miR-24-25p, MCF7/MIMIC-miR-24-2-5p) was compared to the relative control (NW1/MIMIC-negCTRL, MCF7/MIMIC-negCTRL).

first: Margherita

last: Puppo

first: Philippe

last: Clézardin


title: Methylation-modulated PFTK1 regulates Gefitinib resistance via Wnt/β-catenin signalling in EGFR mutant non-small-cell lung cancer cells

city: Guangzhou

first: Xiaoting

last: Jia

address: Hengzhigang Road 78#

institute: Affiliated tumor hospital of GZMU

summary: We implemented mRNA microarray analysis in HCC827 and Gefitinib resistant cells HCC827/GR to identify differential expression of  mRNAs in NSCLC.

overall_design: The global expression profile of mRNA was detected from two kinds of cells (HCC827 and HCC827/GR)  using LncRNA + mRNA Human Gene Expression Microarray V4.0. The RNAs  were labeled with Cy3 dye.

first: Jingjie

last: Tian


title: The neurodevelopmental regulatory role and clinical value of the multi-molecular axis in early-onset schizophrenia [miRNA]

city: Taiyuan

first: Sha

last: Liu

state: Shanxi

address: No. 56 Xinjian South Road, Yingze District, Taiyuan, Shanxi province

department: Department of Psychiatry

institute: First Hospital of Shanxi Medical University

type: Non-coding RNA profiling by high throughput sequencing

summary: Background: Biomarkers identified by previous epigenetic studies of schizophrenia have focused solely on diagnostics or pathology, potentially leading to a disconnection between them. This study focused on early-onset schizophrenia (EOS), aiming to identify the multi-molecular axis centered on circular RNAs (circRNAs), microRNAs (miRNAs), and messenger RNAs (mRNAs) with both neurobiological function and diagnostic value. Methods: Based on the discovery cohort and validation cohort-1, screening of candidate genes on the axis were completed via high-throughput sequencing, expressive and structural validation. Combining dual-luciferase reporter assays with overexpression/knockdown experiments, the axis with binding and regulatory relationships has been established. Subsequently, the functions of genes on the axis were explored through zebrafish embryo manipulation and neural differentiation. The clinical value of the entire axis was assessed in the validation cohort-2. Results: The regulatory effects of hsa-circ-CORO1C on hsa-miR-708-3p, hsa-miR-708-3p on target JARID2 and LNPEP were elucidated. Among them, hsa-miR-708-3p caused aberrant phenotypes including significant craniocerebral malformation and impaired neuron axon growth. JARID2 and LNPEP could facilitate neuronal differentiation and augment synaptic formation. The combined diagnostic efficacy of the whole axis, where hsa-circ-CORO1C could serve as a sponge for hsa-miR-708-3p to alleviate its suppressive effects on JARID2 and LNPEP, surpassed any individual gene we found in EOS. Conclusions: Our study demonstrated a multi-molecular axis, hsa-circ-CORO1C–hsa-miR-708-3p–JARID2+LNPEP, in EOS for the first time. By integrating evidence from genetic, neurophenotypic, and clinical perspectives, we have expanded the comprehension of the pathological mechanism and provided the reference for identifying reliable objective diagnostic biomarkers for EOS.

overall_design: In the cohort of 10 drug-naïve, first-episode patients with early-onset schizophrenia and 10 matched healthy controls, differentially expressed circRNAs and miRNAs were identified via Illumina high-throughput sequencing.

first: Zexuan

last: Li

first: Xinzhe

last: Du

first: Yong

last: Xu


title: Chemotherapeutic agents and leucine deprivation induce codon-biased aberrant protein production in cancer

city: Amsterdam

first: Pierre-Rene

last: Körner

address: Plesmanlaan 121

department: Oncogenomics

country: Netherlands

institute: Netherlands Cancer Institute (NKI)

summary: Messenger RNA (mRNA) translation is a tightly controlled process frequently deregulated in cancer. Key to this deregulation are transfer RNAs (tRNAs), whose expression, processing, and post-transcriptional modifications are often altered in cancer to support cellular transformation. In conditions of limiting levels of amino acids, this deregulated control of protein synthesis leads to aberrant protein production in the form of ribosomal frameshifting or misincorporation of non-cognate amino acids. Here, we studied leucine, an essential amino acid coded for in mRNA by six different codons. Surprisingly, we found that leucine deprivation leads to ribosomal stalling and aberrant protein production in various cancer cell types, predominantly at one codon, UUA. Similar effects were observed after treatment with chemotherapeutic agents, implying a shared mechanism controlling the downstream effects on mRNA translation. In both conditions, a limitation in the availability of tRNALeu(UAA) for protein production was shown to be the cause for this dominant effect on UUA codons. The induced aberrant proteins can be processed and immune-presented as neoepitopes, and can direct T-cell killing. Altogether, we uncovered a novel mode of interplay between DNA damage, regulation of tRNA availability for mRNA translation, and aberrant protein production in cancer that could be exploited for anti-cancer therapy.

overall_design: Ribosome profiling to measure mRNA levels and mRNA translation

first: Adva

last: Kochavi

first: Remco

last: Nagel

first: Pierre-René

first: Onno

last: B Bleijerveld

first: Chun-Pu

last: Lin

first: Zowi

last: Huinen

first: Yuval

last: Malka

first: Natalie

last: Proost

first: Marieke

last: van de Ven

first: Xiaodong

last: Feng

first: Jasmine

last: Montenegro Navarro

first: Abhijeet

last: Pataskar

first: Daniel

last: S Peeper

first: Julien

last: Champagne

first: Reuven

last: Agami


title: Spatial transcriptomics: P. aeruginosa corneal infection

city: Ithaca

first: Hao

last: Zhou

address: 200 Kimball Hall

institute: Cornell University

summary: To holistically unravel the complexity of pathogen-host interactions within infected tissues we leverage a dual spatial transcriptomic approach that, for the first time, simultaneously captures the expression of Pseudomonas aeruginosa genes alongside the entire host transcriptome in a model of ocular infection. This innovative method reveals differential pathogen and host-specific gene expression patterns across specific anatomical regions generating a unified transcriptional map of infection. By integrating these data, we developed a predictive ridge regression model trained on images from infected tissues.

overall_design: Overnight cultures of P. aeruginosa were grown to an OD600 of 0.20 in LB broth, and cultures were then washed and resuspended in 1x sterile PBS (Corning). C57BL/6J mice were anesthetized with ketamine/xylazine (MWI Animal Health) and corneal epithelium was abraded with three, parallel 10mm scratches, performed with a 26 ½ -gauge needle (BD). Pseudomonas was then applied to the abraded cornea topically with 2µl containing 5x105 CFU in PBS in the PA14 infection studies and with 5ml of 1x106 CFU in the PAO1 infection studies. For use in the infection experiments, 100µl of overnight culture was added to 10ml of fresh LB and was subcultured for 2.5-3h to reach OD600=0.20. 1ml of culture was spun down at 3,000 x g (rotor, centrifuge) for 10 min and diluted to a desired inoculum. Experiments were carried out predominantly with male 7-8 week old C57Bl6/J mice. After 24h or 48h, mice were euthanized, and corneas were imaged by brightfield microscopy to observe opacification. Bacterial CFU was quantified at 24h and 48h by serial dilutions of whole eye homogenates onto LB plates and were counted manually.

first: Oscar

last: Negrón

first: Serena

last: Abbondante

first: Nicole

last: Chumbler

first: Eric

last: Pearlman

first: Mihaela

last: Gadjeva


title: Efficient suppression of premature termination codons with alanine by engineered chimeric pyrrolysine tRNAs

city: Indianapolis

first: Jeffery

middle: M

last: Tharp

state: Indiana

address: 635 Barnhill Drive

department: Biochemistry and Molecular Biology

institute: Indiana University School of Medicine

summary: Mutations that introduce premature termination codons (PTCs) within protein-coding genes are associated with incurable and severe genetic diseases. Many PTC-associated disorders are life-threatening and have no approved medical treatment options. Suppressor transfer RNAs (sup-tRNAs) with the capacity to promote translational readthrough of PTCs represent a promising therapeutic strategy to treat these conditions; however, developing novel sup-tRNAs with high efficiency and specificity often requires extensive engineering and screening. Moreover, these efforts are not always successful at producing more efficient sup-tRNAs. Here we show that a pyrrolysine tRNA (tRNAPyl), which naturally translates the UAG stop codon, offers an attractive scaffold for developing effective sup-tRNAs that restore protein synthesis from PTC-containing genes. We created a series of rationally designed Pyrrolysine tRNA Scaffold Suppressor-tRNAs (PASS-tRNAs) that are substrates of bacterial and human alanyl-tRNA synthetase. Using a PTC-containing fluorescent reporter gene, PASS-tRNAs restore protein synthesis to wild-type levels in bacterial cells. In human cells, PASS-tRNAs display robust and consistent PTC suppression in multiple reporter genes, including pathogenic mutations in the tumor suppressor gene BRCA1 associated with breast and ovarian cancer. Moreover, PTC suppression occurred with high codon specificity and no observed cytotoxic effects. Collectively, these results unveil a class of sup-tRNAs with great potential for tRNA-based therapeutics.

overall_design: Ribosome protected footprints and total RNA libraries were prepared from HEK 293 cells transfected with a sup-tRNA-expressing plasmid or an empty vector control to determine the impact of sup-tRNA expression on native stop codon readthrough and gene expression.

last: Vargas-Rodriguez


title: Breast organoid suspension cultures maintain long-term estrogen receptor expression and responsiveness

city: Boston

first: Kung-Chi

last: Chang

state: MA

address: 240 Longwood Avenue, Building C/Rom 501

department: Cell Department

institute: Harvard medical School

summary: Organoid cultures offer a powerful technology to investigate many different aspects of development, physiology, and pathology of diverse tissues. Unlike standard tissue culture of primary breast epithelial cells, breast organoids preserve the epithelial lineages and architecture of the normal tissue. However, existing organoid culture methods are tedious, difficult to scale, and do not robustly retain estrogen receptor (ER) expression and responsiveness in long-term culture. Here, we describe a modified culture method to generate and maintain organoids as suspension cultures. This method improves organoid growth and uniformity over the traditional Matrigel dome embedding method while still maintaining the fidelity of the three major epithelial lineages. Using this adopted method, we are able to isolate and culture hormone sensing (HS) cells alone that retain ER responsiveness upon estrogen stimulation in long-term culture. This culture system presents a valuable opportunity to study the events involved in initiation and evolution of ER-positive breast cancer.

overall_design:
To evaluate if hormone sensing cells still have ER activation upon Estrogen treatment (sample1-12).
To compare Hormone sensing (HS), luminal adaptive secretory precursor fate (LASP), Basal (BA) isolated from human breast organoid cultured in matrigel domes or suspension cultures (sample 13-27).

first: Joan

middle: S

last: Brugge


title: Multimodal Omics Elucidation of Metabolic Transition during the Development of Hepatic Organoids

city: Jinan

first: Shule

last: Zhang

address: 107 Wenhua Xilu

institute: Qilu Hospital of Shandong University

summary: Hepatic organoids (HO） serve as optimal models simulating liver development, where elucidating the changes in transcriptome and proteome during their development is crucial for understanding the in vitro simulation of liver biological development. Given that HO are embedded in a matrix gel during their maturation stage, extracting metabolites while fully preserving the cellular state poses significant challenges. Through transcriptomics, it is possible to infer alterations in metabolic flux, offering an alternative perspective on metabolic transformation. In this study, we conducted transcriptomic and proteomic sequencing across four developmental stages of HO, analyzed differential genes and proteins, the correlation between transcriptome and proteome, the transcription-translation ratio, and changes in metabolic flux.

overall_design: RNA sequencing was performed on the following three stages of the differentiation of HO, including: foregut (FG), hepatoblast (HB), HO1, and HO2. Three replicates were performed for each stage.

first: Dong

last: Li

first: Xianyu

first: Linghong

last: Liu

first: Huixian

last: Ma

first: Qing

last: Shi

first: Xiuli

last: Ju


title: Hepcidin Inhibits Hepatocyte Apoptosis via the PERK Pathway in Acute Liver Injury and Fibrosis

first: Changying

address: Panjiayuan Street 17, Chaoyang District

institute: Chinese Academy of Medical Sciences and Peking Union Medical College

summary: RNA transcription profiling of hepatocytes was compared between C57BL/6 mice and Hamp-/- mice for 24h

overall_design: Four-week-old male WT and Hamp-/- mice were randomly divided into the CCl4 and vehicle groups (n=6). The mice were administered 12.5% CCl4 (CCl4: olive oil= 1:7) or vehicle (olive oil) for 24 h.


title: Epigenomic landscape of human cumulus cells in premature ovarian insufficiency using single-base resolution methylome and hydroxymethylome. [ACE-seq]

first: Wenhao

first: Dongyang

last: Wang

first: Juanzi

first: Chen

last: Huang


title: Interpreting Ovarian Dynamics: Transcriptomic and Metabolomic Insights into Porcine Gonadal Development and Gestation

city: Chengdu

first: Binyun

last: Pan

address: No. 211 Huimin Road, Wenjiang District

institute: Sichuan Agricultural University

summary: The ovary is the central organ of the reproductive system, responsible for producing oocytes, synthesizing and secreting steroid hormones. The healthy development and regular cyclical changes of the ovaries are crucial for regulating the reproductive process. This study conducted high-throughput RNA-seq, miRNA-seq, and non targeted metabolite analysis on the ovarian tissues of Chenghua sows at four stages: day 3 after birth (D3), puberty (Pub), one year after birth (Y1), and day 105 of pregnancy (Pre). By comparing the expression changes of mRNA, miRNA, and metabolites during infancy, adolescence, sexual maturity, and pregnancy, multiple key genes and metabolites related to ovarian development and pregnancy maintenance were identified. Research has found that several genes rich in steroid synthesis pathways, such as CYP11A1, HSD3B1, HSD17B1, and SCARB1, are closely related to ovarian development. Joint analysis was also conducted on differential metabolites in this pathway. In addition, the study also identified key genes that maintain pregnancy, such as LUM, FN1, PLAUR, and SELP. A differentiated miRNA targeted mRNA interaction network has been constructed. Clarifying the molecular regulatory mechanisms of ovarian development and pregnancy in sows may provide new theoretical insights for improving sow fertility.

overall_design: In this study, ovarian tissues were collected at four time points: on postnatal day 3 (D3), at postnatal day 120 just entering puberty (Pub), 1st year (Y1) of birth during sexual maturity, and 105th day of pregnancy (Pre). These sows were obtained from the Chengdu Livestock and Poultry Genetic Resources Protection Center (Sichuan, China). Every Chenghua sows was in good health with no history of reproductive diseases. They were kept under the same environmental conditions with the comparable level of nutrition. Three healthy sows were selected from each age groups, labeled as D3-1, D3-2, D3-3, and so on, and their ovaries were collected for this study. The ovary tissue was first washed with 1X PBS and cutted into 3–5 cm2 fragments and immediately snap-frozen in liquid nitrogen for total RNA extraction.

first: Yanzhi

last: Jiang


title: H3K14ac binding by its Triple Tudor Domain globally directs SETDB1 activity

city: Ulm

first: Ole

last: Ammerpohl

address: Albert-Einstein-Allee 11

country: Germany

institute: UK Ulm

summary: SETDB1 is a major H3K9 methyltransferase. It contains a unique Triple Tudor Domain (3TD) which specifically bind the dual modification of H3K14ac in the presence of H3K9me1/2/3. In this study, we explored the role of the 3TD H3K14ac interaction in the methylation of H3K9 methylation activity of SETDB1. We performed genome wide DNA methylation profiling using Infinium MethylationEPIC BeadChip (EPIC, Illumina Inc., San Diego, CA, USA) in HCT116 and SETDB1 KO cells and anlyse the role of H3K14ac in SETDB1 dependent DNA methylation.

overall_design: Bisulphite converted genomic DNA isolated from HCT116 wild type and HCT116 SETDB1-knockout cells was hybridized to the Illumina MethylationEPIC Beadchip (biological duplicates).

first: Thyagarajan

middle: T

last: Chandrasekaran

first: Alexandra

middle: G

last: Kouroukli

first: Albert

last: Jeltsch


title: Molecular lineage and spatial distribution of subplate neurons in the human fetal cerebral cortex.

city: Xiamen

first: Xueyu

last: Guo

state: Fujian

address: 668 Jimei Avenue, Jimei District

department: Center for Precision Medicine

institute: Huaqiao University

summary: Subplate neurons (SPNs) are among the firstborn neurons in the human fetal cerebral cortex, and play a critical role in establishing intra- and extracortical connections. In this study, we created spatial landscapes and molecular lineages of SPNs by performing spatial transcriptomics, and revealed precise position information of various cortical cell types including subplate neurons and their high diversity in the human fetal cortex. Through the spatial distribution of subplate neurons, we identified the molecular signatures of subplate neurons that are closer to early-born neurons than late-born neurons. We also analyzed human-specific genes and extracellular matrix genes enriched in subplate neurons, highlighting the potential contributions of subplate neurons to cortical neurogenesis and early structural folding.

overall_design: Cerebral cortex tissue sections from fetal (gestational week 12 and 13) were subject to spatial transcriptomics analysis.

first: Tao

last: Sun


title: Diurnal control of H3K27me1 deposition shapes expression of a subset of cell cycle and DNA damage response genes [ChIP-seq]

city: Madrid

first: Crisanto

last: Gutierrez

address: Nicolas Cabrera 1

department: Genome Dynamics and Function

country: Spain

institute: Centro de Biologia Molecular Severo Ochoa

summary: Rhythmic oscillation of biological processes helps organisms to adapt their physiological responses to the most appropriate time of the day. Chromatin remodeling has been described as one of the molecular mechanisms controlling these oscillations. The importance of these changes in transcriptional activation, as well as in the maintenance of heterochromatic regions has been widely demonstrated. However, little is still known on how diurnal changes can impact the global status of chromatin modifications and, hence, control gene expression. In plants, the repressive mark H3K27me1, deposited by ARABIDOPSIS TRITHORAX-RELATED PROTEIN 5 and 6 (ATXR5 and 6) methyltransferases, is largely associated with transposable elements but it also covers lowly expressed genes. Here we show that this histone modification is preferentially deposited during the night. In euchromatic regions it is found along the bodies of DNA damage response genes (DDR), where it is needed for their proper expression. The absence of H3K27me1 translates in an enhanced expression of DDR genes that follows a rhythmic oscillation pattern. This evidences a link between chromatin modifications and their synchronization with the diurnal cycle in order to accurately modulate the activation of biological processes to the most appropriate time of the day.

overall_design: We profiled H3K27me1 and H3 marks by chromatin-immunoprecipitation DNA-sequencing (ChIP-seq) and analyzed the H3K27me1 data normalized to total H3 in wild type (WT) and atxr5/6 double mutant plants at two time points, ZT4 (midday) and ZT16 (midnight). Seedlings were gown under short days (8h light - 16h dark) conditions and harvested 14 days after sowing. Two biological replicates were performed and 1 gr of seedlings was used as starting material.

first: Jorge

last: Fung-Uceda

first: María

last: Gomez

first: Laura

middle: R

last: Casillas

first: Anna

last: Gil


title: Diurnal control of H3K27me1 deposition shapes expression of a subset of cell cycle and DNA damage response genes

overall_design: To study the impact of diurnal oscillation of H3K27me1 deposition in gene expression, we generated the transcriptomes profiles of wild type (WT) and atxr5/6 double mutant plants at two time points, ZT4 (midday) and ZT16 (midnight). Seedlings were gown under short days (8h light - 16h dark) conditions and harvested 14 days after sowing. Three biological replicates were performed.

last: Sol Gomez

last: Rodriguez Casillas

last: Gonzalez Gil


title: Effect of TGFb/MEKi treatment on human melanoma cell lines

city: Zurich

first: Lukas

last: Sommer

state: Zurich

address: Winterthurerstrasse 190

department: Institute of Anatomy

country: Switzerland

institute: University of Zurich

summary: The TGFβ signaling pathway is known for its pleiotropic functions in a plethora of biological processes. In melanoma, TGFβ signaling promotes invasiveness and metastasis formation. However, its involvement in the response to therapy is controversial. While several studies have linked TGFβ signaling to elevated resistance to targeted therapy in melanoma, separate findings have indicated a favorable treatment response through TGFβ-mediated increase of cell death. We now found that the outcome of TGFβ signaling in the context of targeted therapy is dose dependent. Unlike low doses, high levels of TGFβ signal activation induce apoptosis upon simultaneous MAPK pathway inhibition, even in targeted therapy resistant melanoma cell lines. Using transcriptomic analyses, combined with genomic target identification of the critical TGFβ signaling effector SMAD4, we demonstrate that parallel activation of TGFβ signaling and MAPK pathway inhibition causes a complete switch of TGFβ target genes from promoting pro-invasive processes to fueling pro-apoptotic pathways. Investigations of underlying mechanisms identified a novel apoptosis-inducing gene signature. Functional validation of signature members confirmed the deubiquitinating enzyme BAP1, the ubiquitin conjugation factor UBE4B and the pro-apoptotic BCL2 family member BCL2L11 (BIM) to mediate apoptosis in this condition. Using a modified, synthetic version of the TGFβ1 mRNA for intra-tumoral injections, we additionally showcase a potential therapeutic application of this treatment combination.

overall_design: In total 6 human melanoma cell lines were treated with 4 different conditions (control, TGFb only, MEKi only or TGFb + MEKi) for 20h before RNA was isolated and transcriptome sequencing was performed. For two cell lines M130830 and M170117 three replicates were sequenced, for the remaining 4 cell lines only one replicate per condition was sequenced.

first: Benjamin

last: Loos

first: Adrian

last: Salas-Bastos


title: Change in the ability of granulosa cells to elongate transzonal projections during follicle development

city: Kobe

first: Mihoko

last: Fushii

address: Nada-ku

country: Japan

institute: Kobe University

summary: Cumulus cells (CCs) and mural granulosa cells (MGCs) have different phenotypes and different roles in the follicle. However, our previous reports have suggested that CCs and MGCs from bovine early antral follicles become MGC-like cells and CC-like cells, respectively. In this study, we examined whether granulosa cells (GCs) from secondary follicles and MGCs from late antral follicles were able to reconstruct complexes with transzonal projection (TZP)-free denuded oocytes (DOs) and regenerate TZPs similar to MGCs from early antral follicles. Furthermore, to confirm that the regenerated TZPs were functional, the development of reconstructed complexes and oocyte growth in the complexes were determined. After coculture, GCs were able to reconstruct complexes with DOs and regenerate TZPs similar to MGCs from early antral follicles. In addition, the oocytes in integrally reconstructed complexes grew fully and acquired meiotic competence, suggesting that the regenerated TZPs are functional. In contrast, MGCs from late antral follicles had lost the ability to elongate TZPs. Since the ability to regenerate TZPs differed among the cells, the transcriptome was analyzed among GCs, CCs, and MGCs collected from follicles of different sizes. The characteristic of TZP generation coincided with transcriptome changes in two directions from GCs to CCs and MGCs. In conclusion, until the early antral follicle stage, bovine GCs, CCs, and MGCs have common characteristics to elongate TZPs and form antrum-like structures to support oocyte growth in vitro. Furthermore, as the follicle develops, MGCs lose the ability to elongate TZPs.

overall_design: We investigated whether GCs from secondary follicles and MGCs from late antral follicles were able to reconstruct complexes with TZP-free denuded oocytes and regenerate TZPs similar to MGCs from early antral follicles using in vitro growth culture. To confirm that the characteristic of TZP generation was coincident with transcriptome changes, we performed gene expression analysis using data obtained from RNA-seq of 5 different cells at different size follicles.

first: Hirohisa

last: Kyogoku


title: Inhibition of the mineralocorticoid Receptor reduces Cirrhosis associated changes of hepatocyte Glucose and Lipid Metabolism in hepatocytes

city: Halle

first: Alexander

last: Nolze

address: Magdeburger Strasse 6

institute: MLU Halle-Wittenberg

summary: Background & aims: Recent studies have suggested that mineralocorticoid receptor (MR) activation may contribute to the progression of liver cirrhosis. Hypoxia, a common feature in liver cirrhosis, has been found to induce MR activation hepatocytes. However, the impact of non-physiological MR activation remains poorly understood. In this study, we investigate the effect of cirrhosis-induced MR activation in hepatocytes. Methods: RNA sequencing followed by gene ontology term enrichment analysis was performed on liver samples from rats treated for 12 weeks with or without CCl4 and for the last four weeks with or without eplerenone (MR antagonist). We investigated if these changes can be mimicked by hypoxia and if those changes lead to alterations of glucose and lipid metabolism in primary rat hepatocytes (pRH) and human HepG2 cells.

overall_design: All animal experiments were conducted in accordance with the german law and the directive 2010/63/EU. They were ethically and legally approved by the governmental, local animal committees (42502-2-1123 MLU, Landesverwaltungsamt Sachsen-Anhalt or TWZ 16-2021, Jena University hospital). All animal studies were conducted in compliance with the ARRIVE guidelines and the National Institutes of Health (NIH) Guide for the Care and Use of Laboratory Animals. The rats were kept in a specific pathogen-free environment, with ad libitum access to water and standard rodent diet at constant temperature of 22±2°C with a 12/12 h light-dark cycle. Cirrhosis was induced by inhalation of carbon tetrachloride (CCl4) and ingestion of phenobarbital via the drinking water (0.35 g/L) three times a week for 12 weeks in age-matched, male Wistar rats (purchased from Charles river) with an initial weight of 75-100 g. Eplerenone treatment (Inspra, 100 mg kg 1 BW) per os (hazelnut spread, 5g/day) was initiated after eight weeks of CCl4 inhalation, and continued until 12 weeks of CCl4 inhalation. Livers were isolated 7-10 days after the last dose of CCl4. At the end of the experiment, the animals were anesthetized with ketamine/xylazine (100/5 mg/kg i.p.; Ketaset 100 mg/ml, Zoetis, 2% Rompun, Bayer Healthcare) and euthanized by exsanguination while in deep narcosis. RNA sequencing from liver samples was performed. The quality of the RNA samples was assessed using a 2100 Bioanalyzer (Agilent Technologies). All samples had an RNA Integrity Number (RIN) above 7 (with 10 as the maximal possible value). Novogene Co., Ltd. (Cambridge, UK) carried out the sequencing, libraries preparation (poly(A) enrichment), and the paired-end sequencing (2 × 150 bp) runs on a NovaSeq6000 Illumina system. Adaptor clipping and data quality control were provided by the service company as well. Read mapping to the rat genome rn6 was performed with HISAT2 (v. 2.1.0), and featureCounts (2.0.0, –M –t exon) was used to count the mapped reads. Gene annotation was performed using BiomaRt (v.2.44.4) to access the Ensembl archive v101.  Three treatment groups  were defined: a) control: animals received no treatment, b)CCl4: liver cirrhosis was induced by 12 week of treatment with CCl4 & phenobarbital, c) CCl4 + eplerenone: liver cirrhosis was induced by 12 weeks of treatment with CCl4 & phenobarbital, after 8 weeks rats were supplemented with eplerenone per os in hazelnut spread.

first: Mohammad

last: Mohib

first: Sindy

last: Rabe

first: Michael

last: Rooney

first: Quratul

last: Ain

last: Zipprich

last: Gekle

first: Barbara

last: Schreier


title: Mitochondrial DNA mutations are associated with response to anti-VEGF therapy in ovarian cancer PDX models

city: Padova

first: Daniele

last: Boso

address: Via Gattamelata, 64

department: Basic and Translational Oncology Unit

institute: Veneto Institute of Oncology IOV - IRCCS

summary: Mitochondrial DNA (mtDNA) mutations have been reported in several solid tumors including ovarian cancer (OC), the most lethal gynecologic malignancy, and raised interest as they potentially induce mitochondrial dysfunction and rewiring of cellular metabolism. In this study, we characterized mtDNA mutations in ovarian cancer patient derived xenografts (PDX) and investigated their impact on cancer cells at multiple levels. To correlate the presence of mtDNA mutations and metabolic pathways that were modulated at the transcriptional level, RNA-seq analysis was performed in ascites-derived cells from 19 OC PDX (here named PDOVCA). Results indicated that mtDNA mutated (>50% VAF) PDX were endowed with upregulated glycolysis and other pathways connected with cancer metabolism. Functional analysis demonstrated that mtDNA mutations modulated mitochondria activity, assembly capacity and morphology of mitochondrial complexes in PDX cells. Moreover, PDX cells bearing homoplasmic mtDNA mutations behave as glucose addicted and could barely survive glucose starvation in vitro. These findings led us to investigate whether mtDNA mutations correlated with response to anti-VEGF therapy, which was shown to reduce glucose availability in tumors. Strikingly, PDX bearing homoplasmic pathogenic mtDNA mutations had improved survival upon anti-VEGF treatment in mouse models, compared with PDX lacking mtDNA mutations or bearing them at low frequency. These results hint at mtDNA mutations as new biomarkers of response to antiangiogenic drugs.

overall_design:
Ovarian cancer patient-derived xenografts were developed as previosuly described (Indraccolo S. et al., Eur J Cancer, 2006) and propagated by injecting tumor cells intra-peritoneally into 8-week-old female NOD/SCID mice. The ascitic component developed by 19 PDX was harvested and 1 million cells was collected for RNAseq analysis.
We performed gene expression profiling analysis using data obtained from RNA-seq of 19 different PDOVCA ascites-derived cells
Comparative gene expression profiling analysis of RNA-seq data between mtDNA mutated (VAF>50%) and wild type PDOVCA (VAF<50%)

first: Ilaria

last: Piga

first: Sonia

last: Minuzzo

first: Chiara

last: Trento

last: Billato

last: Romualdi

first: Stefano

last: Indraccolo


title: PCBP1/2 and TDP43 Function As NAT10 Adaptors In ac4C Writer Complex Towards mRNAs in Mammalian Cells [293T_acRIPseq]

city: Hangzhou

first: Heng-yu

last: Fan

state: Zhejiang

address: Yuhangtang Road

institute: Life Sciences  Institute

summary: Massive numbers of modified bases in mRNAs sculpt the epitranscriptome and play vital roles in RNA metabolism. The only known acetylated RNA modification, N-4-acetylcytidine (ac4C), is highly conserved across cell types and among species. Although the GCN5-related acetyltransferase 10 (NAT10) functions as an ac4C writer, the mechanism underlying the acetylation process is largely unknown. In this study, we identified the NAT10/PCBP/TDP43 complex as an mRNA ac4C writer in mammalian cells. We identified RNA-binding proteins (RBPs) affiliated with two different families, PCBP1/2 (poly(rC)-binding protein 1/2) and TDP43 (TAR DNA binding protein 43), as NAT10 adaptors for mRNA tethering and substrate selection. Knockdown of the adaptors resulted in decreased mRNA acetylation abundance in HEK293T cells, with globally reduced density in 5`-untranslated region (UTR) and coding sequence (CDS) and ablated cytidine-rich ac4C motifs. The adaptors also affect the ac4C sites by recruiting NAT10 to their binding sequences. The presence of the NAT10/PCBP/TDP43 complex in mouse testes highlights its potential physiological functions in vivo. These findings reveal the composition of the mRNA ac4C writer complex in mammalian cells and expand our knowledge of mRNA acetylation and ac4C site preferences.

overall_design: Polyadenylated RNAs were isolated from total RNAs by Oligo(dT)25 Dynabeads. Libraries were constructed by following the Smart-seq2 procedure.

first: Zhiyan

last: Jiang

first: Yuke

last: Wu

first: Hengyu


title: PCBP1/2 and TDP43 Function As NAT10 Adaptors In ac4C Writer Complex Towards mRNAs in Mammalian Cells

summary: This SuperSeries is composed of the SubSeries listed below.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: PCBP1/2 and TDP43 Function As NAT10 Adaptors In ac4C Writer Complex Towards mRNAs in Mammalian Cells [acRIPseq]


title: Single-cell analysis reveals the loss of progenitor subpopulation of valvular endothelial cells leads to functional mitral regurgitation

city: Beijing

first: mengxia

last: fu

address: xicheng road

institute: fuwai hospital

overall_design:
We conducted single-cell RNA sequencing analysis of mitral valve leaflets extracted from six patients, comprising three individuals with moderate-to-severe functional mitral regurgitation and three nondiseased mitral valve controls. Bioinformatics was used to identify the cell types, describe the cell functions, and investigate cellular developmental trajectories and interactions.
------------------------
Authors state:
Raw sequence data were provided at China Genomic Sequence Archive (GSA). The accession number will be included once the paper is accepted

first: Xiaohu

last: Wang

first: Mengxia

last: Fu

first: Jiangping

last: Song


title: Machine learning-optimized targeted detection of alternative splicing

city: Philadelphia

first: Peter

last: Choi

state: PA

address: 3501 Civic Center Blvd, CTRB 4400

department: Pathology and Laboratory Medicine

institute: The Children's Hospital of Philadelphia

summary: RNA-sequencing (RNA-seq) is widely used for analysis of alternative splicing, but in practice, has inherent biases which hinder its ability to detect and quantify splicing events. To address this, we present a targeted RNA-seq method that specifically enriches for splicing-informative junction-spanning reads. Local Splicing Variation sequencing (LSV-seq) utilizes multiplexed reverse transcription from highly scalable pools of primers anchored near splice junctions of interest. Primers are designed using Optimal Prime, a novel dedicated machine learning algorithm trained on the performance of thousands of primer sequences. LSV-seq achieves high on-target capture rates and concordance with RNA-seq, while requiring several-fold lower sequencing depth. We use LSV-seq to target events with low coverage in GTEx RNA-seq data and discover hundreds of previously hidden tissue-specific splicing events. Our results demonstrate the ability of LSV-seq to capture alternative splicing with exceptional sensitivity and highlight its potential to improve the detection of other RNA features of interest.

overall_design: RNA-seq and LSV-seq of Jurkat cells (JSL1 clone) either left unstimulated or stimulated with PMA. Additionally, LSV-seq of human brain cerebellum, heart atrial appendage and liver tissues. Three replicates for each condition.

first: Kevin

last: Yang

first: Yoseph

last: Barash


title: Expression data from the murine salivary gland (SG; submandibular gland) 8 weeks after the ovariectomy (OVX) or sham surgery (Cont)

city: Yangsan-si

first: Yoojin

last: seo

state: Gyeongsangnam-do

address: 49, Pusandaehak-ro, Mulgeum-eup

institute: School of Dentistry, Pusan National University

summary:
In general, women are more susceptible to xerostomia than men. Clinical studies have indicated a noteworthy increase in the incidence of xerostomia following menopause, along with a negative correlation between saliva estrogen levels and the severity of xerostomia symptoms. Similarly, ovariectomy (OVX) in rodents not only decreases saliva secretion but also triggers pathological alterations in the SG.
We used microarrays to detail the global programme of gene expression underlying OVX-mediated SG dysfuction.

overall_design: six-week-old female BALB/c mice (were anesthetized and either a bilateral OVX (designated as OVX group) or sham surgery (designated as Cont group) was performed as established protocol. Microarray-based transcriptomic profiling of SGs (n=3 for each group) was conducted 8 wks after the surgery.

first: Hyung-Sik

last: Kim

last: Seo


title: Microglial APOE3 Christchurch protects neurons through suppression of ferroptosis in a human iPSC-derived Alzheimer’s disease model

city: Monrovia

first: Xiwei

state: CA

address: 655 HUNTINGTON DR.

institute: City of Hope

summary: Alzheimer's Disease (AD) is the most prevalent neurodegenerative disorder characterized by extracellular amyloid plaque and neuronal tangle formation. A 2019 study discovered an association between the APOE3 Christchurch (APOE3Ch) variant and delayed AD progression in a PSEN1 mutation carrier. However, whether the APOE3Ch variant is causal to the delayed onset of AD and what is the underlying mechanism remains to be explored. In this study, we established neuron-microglia cocultures and neuroimmune organoids using CRISPR/Cas9-edited isogenic APOE3 and APOE3Ch microglia derived from human induced pluripotent stem cells (hiPSCs) along with PSEN1 mutant neurons or brain organoids. We show that APOE3Ch microglia exhibit enhanced phagocytic capacity and increased Tau clearance in neuron-microglia co-cultures. Moreover, APOE3Ch microglia were able to reduce phosphorylated Tau levels in PSEN1 mutant brain organoids. We also demonstrated that APOE3Ch microglia could preserve neural network activity in co-cultured PSEN1 neurons when challenged with synaptosomes prepared from AD patient brains. RNA-seq analysis identified phagocytosis and ferroptosis among pathways of differentially expressed genes from APOE3 and APOE3Ch microglia. Subsequent validation analyses established a causal link between reduced ferroptosis and enhanced phagocytosis including Tau clearance by APOE3Ch microglia, providing mechanistic insights into the neuroprotective role of APOE3Ch microglia. These findings together demonstrate that the APOE3Ch variant plays a causal role in microglial neuroprotection which can be exploited in therapeutic development for AD.

overall_design: Microglia cocultured with with either WT or PSEN1 mutant neurons  were purified with immunoprecipitation before RNA sequencing. Briefly, 2 ×10^6 microglia were cocultured with DRNs for 10 days, the microglia were collected with a gentle blow with 1mL tips to the surface, the suspend media were collected, the attached cells were singlized with TrypLE. Combined microglia from supernatant fraction and singlized fraction and allow to bind with CD11b MicroBeads (Miltenyi Biotec, Cat # 130-049-601) following manufacture's protocol. Total RNA was isolated from the purified cells with TRIzol. RNA quality control was performed by the Integrative Genomics Core at City of Hope.

first: Guoqiang

last: Sun

first: Cheng

first: Yanhong

last: Shi


title: Gene expression in 10 d Wild-type and Amh-cre Hdac3 KO Sertoli cells

city: Nanjing

first: Qing

last: He

state: Jiangsu

address: 101 Longmian Avenue

department: State Key Laboratory of Reproductive Medicine

institute: Nanjing Medical University

summary: This study aims to explore the transcriptional changes in Sertoli cells depleted with HDAC3.  Sertoli cells were isolated from wild-type and Amh-cre Hdac3 KO mice at postnatal day 10.

overall_design: For each group, we have  biological triplicates. Total RNA was extracted using TRIzol reagent. Strand-specific libraries were prepared using the TruSeq Stranded Total RNA Sample Preparation kit (Illumina) according to the manufacturer's instructions.

first: Mengrou

last: Liu

first: Lan

last: Ye


title: RNA-seq of HeLa cells and RPE1 cells after USP39 KD using RNAi

city: Frankfurt am Main

first: Michaela

last: Müller-McNicoll

address: Max-von-Laue-Str. 13

institute: Goethe University Frankfurt

summary: We used HeLa cells and RPE1 cells after knockdown (KD) of USP39 using siRNAs. We compared differential expression and alternative splicing between Control and KD cells.

overall_design: We used 3 biological replicates for ctrl siRNAs and 3 biological replicates for the KD siRNAs. PolyA-selected RNA was sequenced with 150-bp paired end reads with 50 Million reads per replicate.

first: Ivan

last: Dikic


title: RNA sequencing after heroin or immune challenge in male rats

first: Stephanie

last: Daws

address: 3500 N Broad St

institute: Temple University

summary: We report the miRNAs differentially expressed in the nucleus accumbens of adult male rats after either heroin self-administration, experimenter administered heroin, lipopolysaccharide (LPS) or LPS and experimenter administered heroin

overall_design: RNA profiles in nucleus accumbens of adult male Sprague dawley rats after heroin or LPS

last: Sillivan


title: Endosomal trafficking bypassed by RAB5B-CD109 interplay promotes axonogenesis in KRAS-mutant pancreatic cancer

city: Guangzhou

first: Dingwen

last: Zhang

state: Guangdong

address: 106 Zhongshan 2nd Road, Guangzhou, Guangdong Province

department: Department of Pancreatic Surgery

institute: Guangdong Provincial People's Hospital

summary: Perineural invasion (PNI) represents a unique biological feature causing the poor prognosis of pancreatic ductal adenocarcinoma (PDAC), especially with KRAS mutation. Extracellular vesicle (EVs)-packaged circRNAs function as essential mediator for tumor microenviroment communication to trigger PDAC cells invading into the surrounding regions and lead to distant metastasis. However, the regulatory mechanism of exosomal circRNA in PNI of KRAS-mutated PDAC has not been elucidated. Herein, we identified an KRASG12D-mutation responsive exosomal circRNA-circPNIT, which was positively correlated with PNI of PDAC. Functionally, KRASG12D PDAC-derived EV-packaged circPNIT promotes axonogenesis and PNI both in vitro and in vivo. Mechanistically, circPNIT-mediated Rab5B-CD109 interplay bypassed traditional endosomal trafficking to anchor Rab5B in the lipid rafts of multivescular body and fostered the secretion of circPNIT as a cargo of CD109+EVs. Subsequently, CD109+EVs achieved the targeted delivery of circPNIT to neurons by recognizing their surface receptor TRPV1 and facilitated DSCAML1 transcription-induced axonogenesis, which in turn enhanced the PNI by activating GFRα1/RET pathway. Importantly, we established an engineered si-cirPNIT-loaded CD109+EVs to dramatically inhibit PNI of PDAC in a KRASG12D/+ Trp53R172H/+ Pdx-1-Cre mice model. Collectively, Our findings highlight the mechanism underlying EV-packaed circRNA mediating PNI of KRAS mutated-PDAC dependent of a Rab5B endosomal bypass, supporting circPINT as an effective target for the treatment of neuro-metastatic PDAC

overall_design: To explored the mechanism by which KRASG12D PDAC EV-packaged circPNIT regulated axonogenesis after the internalization by dorsal root ganglia cells. Next generation sequencing analysis was performed on 3 pairs of independent dorsal root ganglia cells induced by PANC-1-EVcircPNIT and PANC-1-EVvector, respectively.

first: Yuming

last: Luo

first: Yan

last: Lin

first: Min

last: Yu


title: Comparison of transcriptional profiles of CD62L+ and CD62L-  subsets of in vivo primed CD44hiCD4+ T cells  by exposure to ovalbumin plus papain

city: Jinan

first: Hua

last: Tang

state: Shandong

address: Qingdao Road 6699

institute: Shandong First Medical University &Shandong Academy of Medical Sciences

summary: Memory type 2 CD4+ T helper (Th2) cells play critical roles in allergic asthma, but their generation and maintenance remain poorly understood. Specially, roles of conventional dendritic cells (cDCs), including CD11b+CD103- cDC2s and CD11b-CD103+ cDC1s, in this process are still undefined. Here, we report that fate decision of memory Th2 cells was dictated by cDC subsets at priming stage. To be specific, temporary depletion of CD103+ cDC1s during priming promoted the generation of allergen-specific circulating memory Th2 cells in exposure to ovalbumin plus papain, suggesting that precursors of memory Th2 cells emerged at priming stage. Subsequent data revealed that activated CD44hiCD62L+CD4+ T cells, which were elevated by cDC1 depletion during priming, exhibited memory-precursor-like transcriptional and functional properties. Taken together, CD103+ cDC1s reduced number of CD62L+ memory precursors, and thus restrained generation of memory Th2 cells.

overall_design: Naïve CD45.1+CD4+OT-II T cells (specifically recognize ovalbumin), sorted from un-treated CD45.1×OT-II mice, were injected into C57BL/6 (WT) and Batf3-/- mice (CD45.2+), and recipients were immunized with ovalbumin plus papain 24 h after injection. CD62L+ and CD62L- subsets of activated CD44hiCD45.1+T cells were sorted from mediastinal lymph nodes of WT and Batf3-/- mice 5 days after immunization. Naïve CD45.1+CD4+OT-II T cells were used as control. Three independent replicates were prepared for each cell subset.

first: Jianguo

first: Linqing

last: Du

first: Shuying

last: Yi

first: Yanan

last: Bi

first: Lina

last: Li

first: Guangming

last: Zhu

first: Xiuli

last: Jing

first: Wengang

last: Song


title: Control of protein accumulation by LON2 peroxisomal protease and autophagy

city: Houston

first: Bonnie

last: Bartel

state: Texas

department: BioSciences

institute: Rice University

summary: Peroxisomes are membrane-bound eukaryotic organelles that house essential metabolic pathways. Many of these pathways generate damaging reactive oxygen species (ROS) that also function as signaling molecules. In Arabidopsis thaliana, damaged or obsolete peroxisomal proteins can be degraded by the LON2 peroxisomal protease. In addition, peroxisomes can be entirely degraded through a specialized form of autophagy (pexophagy), a process that is amplified when LON2 is dysfunctional. As a first step towards surveying the transcriptional landscape of dysfunctional LON2 , we used RNA-seq analysis to compare transcript that are induced and reduced when LON2 is dysfunctional in the presence and absence of autophagy.

overall_design: RNA-seq of 6 day old continuous light grown whole seeding extract of Arabidopsis wild type Col-0 (3 biological replicates) as a control, atg2-4 mututant (3 biological replicates), lon2-8 mutant (3 biological replicates), lon2-8 atg2-4 double mutant (3 biological replicates).

first: DurreShahwar

last: Muhammad

first: Natalie

last: Clark


title: Charaterization of transcriptomic profile of subcutaneous adipose tissue from Constitutive Thin women

city: Pierre Bénite

first: emmanuelle

last: meugnier

address: Cens Eli Bat 2 D- Hopital Lyon Sud

country: France

summary: NOT PROVIDED; REQUESTED

overall_design: PeriOmbilical Subcutaneous Adipose tissue biopsies from Constitutive Thin (CT) women and Lean women were compared using Affymetrix Chips

first: Emmanuelle

last: Meugnier

first: Hubert

last: Vidal


title: Expression data from SAMP8 and SAMR1 mice brains in TFQA treatments

status: Public on Dec 26 2024

city: Tsukuba City

first: Hongyu

state: Ibaraki

address: Tennodai 1-1-1

department: Life and Earth Sciences

institute: University of Tsukuba

summary:
TFQA treatment in SAMP8 mice showed the significant performance on improving learning and memory compared with no treatment SAMP8 mice brains.  TFQA treatments in SAMP8 mice also had significant activation of neurotrophin signaling pathway, by increasing BDNF.
We used microarrays to detail the global programme of gene expression underlying treatment and identified distinct classes of regulated genes during this process.

overall_design: SAMP8 and SAMR1 mice brains were selected  for RNA extraction on Affymetrix microarrays. We selected 6 months old adult male SAMP8 and SAMR1 mice. After 37 days administration of TFQA,  mice were dissected and brains were isolated.


title: Multiplexed mosaic tumor models reveal natural phenotypic variations in drug response within and between populations [8 cancer cell lines]

city: San Francisco

first: Hani

last: Goodarzi

department: Department of Biochemistry and Biophysics

institute: UCSF

summary: Heterogeneity between patients poses a significant challenge in cancer drug development. Although success of a drug candidate in the clinic depends on efficacy across large numbers of diverse patients, only a few in vivo preclinical models are used for assaying a preclinical drug candidate due to the lack of scalability of traditional xenograft models. To address this limitation, we developed GENEVA, a scalable platform that enables the measurement of molecular and phenotypic responses to drug perturbations at single-cell resolution, both in 3D and in vivo. GENEVA models the genetic diversity of cancer by combining multiple patient-derived cell lines and cancer cell lines into pooled 3D cultures and xenograft models, allowing us to study drug responses across a wide range of genetic backgrounds within a single experiment. This platform integrates high-resolution transcriptomics with multiplexed single-cell profiling, providing a unique ability to uncover the interplay between cancer genotypes and drug responses. In this study, we apply GENEVA to investigate KRAS G12C inhibitors and demonstrate that mitochondrial activation is a key driver of cell death following KRAS inhibition. Additionally, we identify epithelial to mesenchymal transition (EMT) as a prominent resistance mechanism to KRAS G12C inhibition using in vivo GENEVA mice. These findings highlight the utility of GENEVA in identifying novel therapeutic targets and optimizing combination therapies, while underscoring its potential to bridge the gap between preclinical cancer models and patient outcomes by modeling the complexity of tumor heterogeneity in a scalable and robust assay.

overall_design: A total of 8 cancer cell lines with varying genetic backgrounds, including BRAF(V600E) and KRAS G12C mutations, were pooled together into a single experimental setup. These pooled cells were cultured in 3D conditions and then split into 4 treatment groups:   Vehicle control (DMSO),   ARS-1620 (0.4 µM, 1.6 µM, 25 µM)  After six days of treatment, cells were harvested and subjected to scRNA-seq

first: Johnny

middle: X

first: Jung

last: Suh

first: Katerina

middle: D

last: Popova

first: Kristle

last: Garcia

first: Tanvi

last: Joshi

first: Bruce

last: Culbertson

first: Jessica

middle: B

last: Spinelli

first: Vishvak

last: Subramanyam

first: Kevin

last: Lou

first: Kevan

last: Shokat

first: Jonathan

last: Weissman


title: Exacerbated Salmonellosis in poly(ADP-ribose) polymerase 14 deficient mice

city: Turku

first: Madhukar

last: Vedantham

address: Kiinamyllynkatu 10-D

country: Finland

institute: University of Turku

summary: Salmonella enterica serovar Typhimurium infection triggers a robust immune response in the host, but the molecular mechanisms regulating this response are not fully understood. Parp14 has been shown to enhance type I interferon responses and influence bacterial resistance, particularly in macrophages. To explore the role of Parp14 in immune regulation, we investigated Salmonella infection in Parp14 knockout mice over 5 days. On day 1 post-infection (p.i.), Parp14-deficient mice displayed elevated bacterial loads in the liver, but no differences in other tissues. By day 5, bacterial counts in the proximal colon were slightly lower in Parp14 knockout mice compared to wild-type controls, suggesting Parp14 regulates immune responses across multiple cell types, including epithelial cells. Histopathological analyses revealed increased immune cell infiltration, goblet cell loss, and tissue erosion in the proximal colon of Parp14-deficient mice on day 5 p.i. RNA-seq data from proximal colon tissue identified impaired expression of immune response genes, including Ccl2, Il1b, and Il6, which are critical for early-stage infection defense. Further analysis revealed downregulation of genes involved in cellular adhesion and cytoskeleton maintenance. These findings indicate that Parp14 is essential for an effective early innate immune response and for maintaining epithelial integrity during Salmonella infection, highlighting its potential as a therapeutic target for bacterial infections.

overall_design: To investigate the transcriptome changes in colon of Parp14-deficient mice when compared to wt mice in resting state and post Salmonella infection, we performed bulk tissue RNA-Seq analysis of the mouse colon proximal sections 1-day post-infection.

first: Arto

last: Pulliainen


title: Multiplexed mosaic tumor models reveal natural phenotypic variations in drug response within and between populations [11 cancer cell lines]

overall_design:
A total of 11 cancer cell lines with varying genetic backgrounds, including BRAF(V600E) and KRAS G12C mutations, were pooled together into a single experimental setup. These pooled cells were cultured in 3D conditions and then split into three treatment groups:
Vehicle control (DMSO),
Vemurafenib (0.8 µM) targeting BRAF(V600E) mutants,
ARS-1620 (1 µM) targeting KRAS G12C mutants.
After six days of treatment, cells were harvested and subjected to scRNA-seq

last: Yu


title: Type I interferon signaling and peroxisomal dysfunction contribute to enhanced inflammatory cytokine production in Irgm1-deficient macrophages

city: Durham

first: Gregory

middle: Alan

last: Taylor

state: NC

address: DUMC Box 3003

department: Medicine

institute: Duke University

summary: The human IRGM gene has been linked to inflammatory diseases including sepsis and Crohn’s disease. Decreased expression of human IRGM, or of the mouse orthologues Irgm1 and Irgm2, leads to increased production of a number of inflammatory chemokines and cytokines in vivo and/or in cultured macrophages. Prior work has indicated that increased cytokine production is instigated by metabolic alterations and by changes in mitochondrial homeostasis; however, a comprehensive mechanism has not been elucidated. In the studies presented here, RNA deep sequencing and quantitative PCR were used to show that increases in cytokine production, as well as most changes in the transcriptional profile of Irgm1-/- bone marrow-derived macrophages (BMM), are dependent on increased type I IFN production seen in those cells. Metabolic alterations that drive increased cytokines in Irgm1-/- BMM - specifically increases in glycolysis and increased accumulation of acyl-carnitines - were unaffected by quenching type I IFN signaling. Dysregulation of peroxisomal homeostasis was identified as a novel upstream pathway that governs type I IFN production and inflammatory cytokine production. Collectively, these results enhance our understanding of the complex biochemical changes that are triggered by lack of Irgm1 and contribute to inflammatory disease seen with Irgm1-deficiency.

overall_design: To gain more insight into the transcriptional changes that are initiated by Irgm1 deficiency and their dependence on type I IFN, we performed deep RNA sequencing and Gene Expression Enrichment Analysis (GSEA) comparing BMM cultured from WT, Irgm1-/-, and IFNAR/Irgm1-/- mice

middle: A

first: Joseph

last: Prinz

first: Devi

last: Swain Lenz


title: Epididymis-specific RNase A family genes regulate fertility and small RNA processing

city: Santa Cruz

first: Upasna

last: Sharma

address: 1156 High Street

department: Molecular, Cell & Developmental Biology

institute: UC Santa Cruz

summary: Sperm small RNAs are implicated in intergenerational transmission of paternal environmental effects. Small RNAs generated by cleavage of tRNAs, known as tRNA fragments (tRFs) or tRNA-derived RNAs (tDRs or tsRNAs), are an abundant class of RNAs in mature sperm, and can be modulated by environmental conditions. The biogenesis of tRFs in the male reproductive tract remains poorly understood. Angiogenin, a member of the Ribonuclease A superfamily (RNase A), cleaves tRNAs to generate tRFs in response to cellular stress. Four paralogs of Angiogenin, namely Rnase9, Rnase10, Rnase11, and Rnase12, are specifically expressed in the epididymis —a long, convoluted tubule where sperm mature and acquire fertility and motility. The biological functions of these genes remain largely unknown. Here, by generating mice deleted for all four genes (Rnase9-12-/-, termed “KO” for Knock Out), we report that these genes regulate fertility and small RNA levels. KO mice showed complete male sterility. KO sperm fertilized oocytes in vitro but failed to efficiently fertilize oocytes in vivo due to an inability of sperm to pass through the utero-tubular junction. Intriguingly, there were decreased levels of fragments of tRNAs (tRFs) and rRNAs (rRNA-derived small RNAs or rsRNAs) in the KO epididymis and epididymal luminal fluid. RNases 9-12 did not show ribonucleolytic activity in-vitro, suggesting that the changes in tRF and rsRNA levels are independent of enzymatic activity of these proteins.  Importantly, KO sperm showed a dramatic decrease in the levels of tRFs, demonstrating a role of epididymis-specific Rnase9-12 genes in regulating sperm small RNA composition. Together, our results reveal an unexpected role of four epididymis-specific non-canonical RNase A family genes in regulating fertility and small RNA processing.

overall_design: Small RNA sequencing of various male reproductive tract tissues including sperm,  epididymis, and epididymal fluid isolated from wild type mice or mice deleted for four epididymis-specific genes Rnase9, Rnase10, Rnase11 and Rnase12 (Rnase9-12 KO)

first: Joshua

middle: F

last: Shaffer

first: Alka

last: Gupta

first: Geetika

last: Kharkwal

first: Edgardo

middle: E

last: Linares

first: Andrew

last: Holmes

first: Julian

last: Swartz

first: Sol

last: Katzman


title: The effect of CXCL10 on macrophages

city: Shanghai

first: Zhe

last: Wang

state: Shanghai

address: Siping Road, Shanghai

institute: Tongji university

summary: To explore the effect of CXCL10 on the immune microenvironment, we induced macrophages from thp1 cell line and treated macrophages with the addition of CXCL10 and their controls.

overall_design: The human leukemia monocytic cell line THP-1 were treated with PMA (100ng/ml) for 48h to differentiate THP-1 monocyte to macrophages. We add the recombinant human CXCL10 (100ng/mL) or the control solutions in the induced macrophages for 24h


title: Transcriptomic signatures of antibody mediated rejection in early biopsies with negative histology in HLA incompatible kidney transplantation

city: Praha 4

first: Petra

last: Hruba

address: Videnska 1958

department: Transplant Laboratory

country: Czech Republic

institute: Institute for Clinical and Experimental Medicine

summary: Presensitized patients with circulating donor-specific antibodies (DSA) prior to transplantation are at risk for antibody-mediated rejection (ABMR). Peritransplant desensitization mitigates but does not eliminate the alloimmune response. We examined the possibility that subthreshold ABMR activity undetected by histology could be operating in some early biopsies.

overall_design: Whole-genome mRNA profiling was performed with Affymetrix microarrays in 30 kidney transplant biopsies, retrospectively selected from all biopsies performed within the first month with available MMDx reports.  Kidney allograft biopsy transcriptome in presensitized patients (DSA+) who had received desensitization and did not develop active or probable ABMR by histology (R-/DSA+, n=10) was compared with biopsies showing active/probable ABMR (R+/DSA+, n=5).  As negative controls, biopsies without rejection by histology performed within the identical time frame in patients without donor-specific antibodies at transplantation (R-/DSA-, n=15) were used .

first: Jiri

last: Klema

last: Mrazova

first: Eva

last: Girmanova

first: Matej

last: Roder

first: Ludek

last: Voska

first: Martin

last: Kment

first: Martina

last: Mackova

first: Klara

last: Osickova

first: Vladimir

last: Hanzal

first: Philip

last: Halloran

first: Ondrej

last: Viklicky


title: Heterogeneous Immune Landscapes and Macrophage Dynamics in Primary and Lung Metastatic Adenoid Cystic Carcinoma of the Head and Neck

city: Beijing

first: Xiaohong

last: Chen

address: 1,DongJiaoMinXiang Street, DongCheng District,Beijing, P.R.China.100730

department: Department of Otolaryngology Head and Neck Surgery

institute: Beijing Tongren Hospital, Capital Medical University

summary: Recurrent or metastatic adenoid cystic carcinoma (ACC) of the head and neck is rare and highly aggressive. Due to the ineffectiveness of immune checkpoint therapies, this study aims to investigate the tumor immune microenvironment of primary tumor tissues and lung metastatic tissues and to comprehend the challenges of immunotherapy. We analyzed RNA sequencing data and constructed immune landscapes from 25 primary tumors and 34 lung metastases.  Compared to adjacent normal tissues, both primary and lung metastatic ACC showed low immune infiltration. Lung metastases had higher immune infiltration levels and antigen presentation scores but also higher T cell exclusion and dysfunction scores. Higher immune cell infiltration in lung metastases is countered by the presence of suppressive tumor-associated macrophages, which may limit effective anti-tumor responses.

overall_design: To investigate the heterogeneous tumor immune microenvironment between primary ACC and lung metastatic ACC, we performed RNA sequencing data on 25 primary ACC tumors and 34 lung metastatic tumor samples.

first: Xuelian

first: Tingyao

last: Ma

first: Shujing

last: Zhang

first: Guoliang

last: Yang

first: Yue

last: Zhao


title: Aberrantly Expressed RECQL4 Helicase Supports Proliferation and Drug Resistance of Human Glioma Cells and Glioma Stem Cells

city: Warsaw

first: Bartosz

last: Wojtas

address: Pasteura 3

country: Poland

institute: Nencki Institute of Experimental Biology

summary: RECQL4 depletion in human glioma cells with genetic tools slightly impaired cell viability and DNA replication, induced changes in gene expression and increased sensitivity of glioma cells and glioma stem cells to chemotherapeutics

overall_design: RecQL4 knock-out and wild type LN18 and LN229 cell lines were used to the experiments.

first: Sylwia

middle: K

last: Krol

first: Agnieszka

last: Kaczmarczyk

first: Kamil

last: Wojnicki

first: Bartoisz

first: Bartlomiej

last: Gielniewski

first: Wieslawa

last: Grajkowska

first: Katarzyna

last: Kotulska

first: Cezary

last: Szczylik

first: Ryszard

last: Czepko

first: Mariusz

last: Banach

first: Wojciech

last: Kaspera

last: Szopa

first: Andrzej

last: Marchel

first: Tomasz

last: Czernicki

first: Bozena

last: Kaminska


title: Template switching enables chemical probing of native RNA structures

city: Ann Arbor

first: Chase

middle: Andrew

last: Weidmann

state: MI

address: 1150 W. Medical Center Drive

department: Biological Chemistry

institute: University of Michigan

summary: RNAs are often studied in non-native sequence contexts to facilitate structural studies. However, seemingly innocuous changes to an RNA sequence may perturb the native structure and generate inaccurate or ambiguous structural models. To facilitate the investigation of native RNA secondary structure by selective 2′ hydroxyl acylation analyzed by primer extension (SHAPE), we engineered an approach that couples minimal enzymatic steps to RNA chemical probing and mutational profiling (MaP) reverse transcription (RT) methods - a process we call template switching and mutational profiling (Switch-MaP). In Switch-MaP, RT templates and additional library sequences are added post-probing through ligation and template switching, capturing reactivities for every nucleotide. For a candidate SAM-I riboswitch, we compared RNA structure models generated by the Switch-MaP approach to those of traditional primer-based MaP, including RNAs with or without appended structure cassettes. Primer-based MaP masked reactivity data in the 5′ and 3′ ends of the RNA, producing ambiguous ensembles inconsistent with the conserved SAM-I riboswitch secondary structure. Structure cassettes enabled unambiguous modeling of an aptamer construct but introduced non-native interactions in the full-length riboswitch. In contrast, Switch-MaP provided reactivity data for each nucleotide in each RNA and enabled unambiguous modeling of secondary structure, consistent with the conserved SAM-I fold. Switch-MaP is an alternative approach to primer-based and cassette-based chemical probing methods that precludes primer masking and the formation of alternative secondary structures due to non-native sequence elements.

overall_design: We developed a template-switching approach to mutational profiling of SHAPE chemical probes to obtain conformational flexibility measurements for all nucleotides of a candidate riboswitch. We compared structural models generated from this new approach to two alternative procedures.

first: Ian

last: Hall

first: Martin

last: O'Steen

first: Sophie

last: Gold

first: Sarah

last: Keane


title: USP4 depletion-driven RAB7A ubiquitylation impairs autophagosome-lysosome fusion and aggravates periodontitis

city: Hangzhou, Zhejiang Province

first: Sen

last: Kang

address: 166 North Qiutao Road

institute: Stomatology Hospital, School of Stomatology, Zhejiang University School of Medicine

summary: In exploring the pathogenesis of periodontitis, we collected clinical gingival tissues from healthy donors and patients with severe periodontitis for RNA-Seq analysis, and calculated the significant statistical correlation values (R) between autophagy-related genes and inflammation-related genes in human gingival tissues, which  could inform a potential therapeutic target for the treatment of periodontitis.

overall_design: To explore the probable relationship between autophagy-related genes and inflammation-related genes, Clinical human gingiva tissues were collected from severe periodontitis patients (Stage IIIC) and healthy individuals.  We then performed gene expression profiling analysis using data obtained from RNA-Seq of these samples.

first: Peihui

last: Ding


title: Effect of Electrospun Poly (L-lactide-co-caprolactone) and Formulated Porcine Fibrinogen for Diabetic Foot Ulcers

city: shanghai

first: guili

last: wang

state: 上海

address: shanghai longhang road jinshan district

institute: fudan

summary:
OBJECTIVE To compare the effect of topical application PLCL/Fg dressing with alginate dressing (ConvaTec Ltd) when treating DFUs.
DESIGN, SETTING, AND PARTICIPANTS A single-center, prospective, randomized, patient-blinded clinical trial was conducted from February 1, 2023, to November 16, 2023. The eligible patients with DFUs of 1 to 20 cm2 present for at least 1 month and with Wagner grade 1 or 2. They were randomized 1:1 to receive PLCL/Fg or control alginate dressing. In PLCL/Fg group, 10 wounds samples were got from five patients at the time of the first dressing application and 50% wound healing time point. The wound specimens were subjected to messenger RNA sequencing.
INTERVENTIONS Participants received PLCL/Fg dressing 1 to 3 times per week or Alginate dressing 3 times per week for 12 weeks.
MAIN OUTCOMES AND MEASURES Primary end point was the incidence of complete healing. Complete healing was defined as complete re-epithelialization of the ulcer bed and absence of secretion. Secondary end points were time to relevant 50% wound reduction, infection, treatment-associated adverse events, and the microenvironment changing during wound healing.

overall_design: A single-center, prospective, randomized, patient-blinded clinical trial was conducted from February 1, 2023, to November 16, 2023. The eligible patients with DFUs of 1 to 20 cm2 present for at least 1 month and with Wagner grade 1 or 2. They were randomized 1:1 to receive PLCL/Fg or control alginate dressing. In PLCL/Fg group, 10 wounds samples were obtained from five patients at the time of the first dressing application and 50% wound healing time point. The wound specimens were subjected to messenger RNA sequencing.

first: Guili

last: Wang

first: Zhihui

last: Dong


title: Macropinocytosis promotes foamy monocyte formation in hypercholesterolemic mice

city: Augusta

first: Gabor

last: Csanyi

state: GA

address: 1460 Laney Walker Ave

department: Vascular Biology Center

institute: Augusta University

summary: Atherosclerosis-associated vascular disease is the leading cause of death worldwide. Clinical and experimental data demonstrated that circulating monocytes internalize plasma lipoproteins and become lipid-laden foamy cells in hypercholesterolemic subjects. This study was designed to identify the endocytic mechanisms responsible for foamy monocyte formation, perform functional and transcriptomic analysis of foamy and non-foamy monocytes, and characterize specific monocyte subsets in the circulation from normocholesterolemic controls and hypercholesterolemic patients. The presence of foamy monocytes was confirmed in hypercholesterolemic mice and humans via flow cytometry analysis. High resolution scanning electron microscopy (SEM) and quantification of FITC/TRITC-dextran internalization demonstrated macropinocytosis stimulation in human (THP-1) and wild type murine monocytes in vitro. Stimulation of macropinocytosis induced foamy monocyte formation in the presence of unmodified, native LDL (nLDL) and oxidized LDL (ox-LDL) in vitro. Genetic blockade of macropinocytosis (LysMCre+ Nhe1f/f) inhibited foamy monocyte formation in hypercholesterolemic mice in vivo and attenuated monocyte adhesion to atherosclerotic aortas ex vivo. qRT-PCR quantified mRNA levels of major scavenger receptors (SR) in foamy and non-foamy monocytes and identified CD36 as a major SR increasing in response to lipid loading. Deletion of CD36 (Cd36-/-) inhibited foamy monocyte formation in hypercholesterolemic mice. Mechanistic studies identified NADPH oxidase 2 (Nox2)-derived superoxide (O2⋅−) as an important downstream signaling molecule stimulating macropinocytosis in monocytes. Bulk RNA-sequencing characterized transcriptional differences between non-foamy and foamy monocytes and macrophages. Flow cytometry analysis of CD14 and CD16 expression demonstrated a significant increase in intermediate monocytes in hypercholesterolemic patients compared to normocholesterolemic controls. These results provide novel insights into the mechanisms of foamy monocyte formation and potentially identify new therapeutic targets in the treatment of atherosclerosis.

overall_design: To investigate differential gene expression in foamy and non-foamy monocytes and macrophages, primary splenic monocytes were isolated from wild type C57BL/6 mice and differentiated into macrophages. Cells were treated with vehicle or ox-LDL. Bulk RNA sequencing was performed to assess transcriptional changes.

first: WonMo

last: Ahn

first: Faith

middle: N

last: Burnett

first: Kamila

last: Wojnar-Lason

first: Jaser

last: Doja

first: William

last: Bryant

first: Amritha

last: Sreekumar

first: Pushpankur

last: Ghoshal

first: Bhupesh

last: Singla

first: Graydon

last: Gonsalvez

first: Ryan

last: Harris

first: Xiaoling

first: Joseph

last: Miano


title: Inhibiting intracellular CD28 in cancer cells enhances antitumor immunity and overcomes anti-PD-1 resistance via targeting PD-L1

city: Tianjin

first: Zhen

last: Yang

address: Weijin Street

institute: Nankai University

summary: To determine how deficiency of Cd28 influences 4T1 cells, we performed a transcriptional analysis

overall_design: RNA form WT and Cd28 knockout 4T1 cells were isolated by Trizol

first: Xinpeng

last: Liu

first: Xuetao

last: Cao


title: ChIP sequencing of wild-type, TRαLKO, TRβKO intestine at stage 54 with and without T3 treratment.

city: Bethesda

last: Dale

state: MD

address: Rm 10D39, 10 Center Drive

department: National Institute of Child Health and Human Development

institute: National Institutes of Health

summary: Conclusions: Our study represents the first detailed comparative analysis of TR binding between wild-type,TRαLKO, TRβKO intestine, with biologic replicates, generated by ChIP-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles.

overall_design: Examination of 3 biological replicate from the different Xenopus tropicalis tadpoles. Intestines from WT, TRαLKO, TRβKO stage 54 tadpoles were treated with T3 for 18 hours when indicated.

first: Yuta

last: Tanizaki

first: Hongen

last: Zhang

first: Yun-Bo

last: Shi


title: Tumor suppressive role of CELF2 in skin tumorignesis

city: Austin

first: Liu

last: Liang

address: 801 16th Ave NE

institute: The HorMel Institute

summary: To investigate the function of CELF2 in non-melanoma skin cancer, we performed immunofluorescence staining studies and found lower CELF2 expression in human squamous cell carcinoma (SCC) tumors than in adjacent normal skin. CELF2 expression was also downregulated during both ultraviolet light- and chemical-induced skin tumorigenesis in mice, suggesting that CELF2 loss might promote skin cancer development. By using shRNA-mediated knockdown (KD) of CELF2 expression, we showed that CELF2 deficiency significantly increased SCC cell proliferation and colony growth in vitro and increased SCC tumor growth in a xenograft mouse model. Although control SCC cells were sensitive to anticancer drugs such as doxorubicin, CELF2-KD SCC cells were resistant to drug-induced tumor growth retardation. Through RNA-seq analysis, we identified that CELF2 loss led to activation of KRT80 and GDF15, which could confer growth and survival advantages to CELF2-deficient SCC cells.

overall_design: We used cultured human SCC13 cells, mouse models, and clinical samples to analyze the function of CELF2 in skin tumorigenesis.

first: Bindeshwar

last: Sah

first: Jasvinder

last: Singh

first: Yao

last: Shen

first: Liang


title: Analysis of an HCA-7 derived cell line (CC-CR) and cell lines generated with knockout of miR-100, miR-125b, or both miRNAs.

city: Nashville

first: Hua-Chang

last: Chen

state: TN

address: 2525 West End Avenue, Suite 1100

department: Department of Biostatistics

institute: Vanderbilt University Medical Center

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Tim-3 on natural killer cells regulates immune response via antibody dependent cellular cytotoxicity in HER2-positive gastric cancer

city: Fukuoka

first: Chikanori

last: Tsutsumi

address: 3-1-1 Maidashi, Higashi-ku

department: Department of Surgery and Oncology, Graduate School of Medical Sciences

institute: Kyushu University

summary: The present study suggests that combined therapy with anti-Tim-3 antibodies and trastuzumab can be a new treatment strategy for HER2-positive GC.

overall_design:
We performed scRNA-seq analysis from 24 gastric cancers.
Please note that the following samples have been included and re-analyzed in the current study:
GSE201347/PRJNA830909:
GEO sample title SRA Experiment BioSample
GSM6061522 gastric cancer [G41T] SRX14976673 SAMN27744099
GSM6061523 gastric cancer [G42T] SRX14976675 SAMN27744098
GSM6061524 gastric cancer [G43T] SRX14976674 SAMN27744097
GSM6061525 gastric cancer [G44T] SRX14976676 SAMN27744096
GSM6061526 gastric cancer [G45T] SRX14976677 SAMN27744095
GSM6061527 gastric cancer [G47T] SRX14976678 SAMN27744094
GSM6061528 gastric cancer [G49T] SRX14976679 SAMN27744093
GSM6061529 gastric cancer [G50T] SRX14976680 SAMN27744092
GSM6061530 gastric cancer [G51T] SRX14976681 SAMN27744091
GSM6061531 gastric cancer [G52T] SRX14976682 SAMN27744090
GSM6061532 gastric cancer [G55T] SRX14976683 SAMN27744089
GSE231539/PRJNA966751:
GSM7290598 gastric cancer-Chemotherapy [G54T] SRX20201151 SAMN34550038
GSM7290599 gastric cancer-Chemotherapy [G57T] SRX20201152 SAMN34550037

first: Kenoki

last: Ohuchida


title: Comparative gene expression profiling of parental A375 melanoma cells and a panel of spontaneously vemurafenib-resistant  derivative cell lines

city: Iowa City

first: Jesse

last: Riordan

state: IA

address: 375 Newton Road

institute: University of Iowa

summary: The use of selective BRAF inhibitors (BRAFi) has produced remarkable outcomes for patients with advanced cutaneous melanoma harboring a BRAFV600E mutation. Unfortunately, the majority of patients eventually develop drug-resistant disease through a variety of mechanisms. We previously described the creation of a panel of spontaneously BRAFi/MEKi-resistant A375 melanoma cells that displayed multiple different resistance mechanisms arising within the same parental cell population. These resistance mechanisms include (i) rearrangements in the BRAF gene resulting in truncation of the N-terminal regulatory domain or tandem duplication of the BRAF kinase domain, (ii) loss of Ras-GAP NF1, and (iii) a slower arising form of drug resistance associated with a more spread cell morphology and increased phospho-MEK S298, a characteristic of cells with increased Rac1 signaling. We provide here comprehensive gene expression profiling for this panel of cell lines. Rac1 mutation, particularly the P29S variant, is common in melanoma patients, where it is associated with BRAFi/MEKi resistance and worse clinical outcomes. Here we delineate mechanisms of Rac1-driven MAPKi-resistance and identify strategies to inhibit the growth of this class of cutaneous melanomas. We find that Rac1-driven melanomas manifest pleiotropic resistance mechanisms including (i) reduced dependence on BRAF/MEK, (ii) activation of alternative MAPK pathways utilizing Jun kinase and p38 MAP kinase, and (iii) a partial reliance on YAP/TAZ signaling. Importantly, although Rac1-driven melanoma cells display reduced dependence on BRAF/MEK, they are not completely ERK-independent. Moreover, the presence of activated Rac1 appears to create a dependency on focal adhesion kinase (FAK) signaling in undifferentiated melanoma cells. As a result, we find that, despite the pleiotropic mechanisms of Rac1-driven MAPKi resistance, combined inhibition of MEK and FAK with avutometinib plus defactinib is a promising approach for suppressing the growth of Rac1-driven cells. Thus, the avutometinib plus defactinib combination, which is currently being investigated for brain metastatic cutaneous melanoma may also have utility against Rac1-driven MAPKi-resistance in heavily pre-treated, advanced disease.

overall_design: RNAseq profiling of parental A375 cells, A735 cells treated with vemurafenib for 24 or 96 hours, and a panel of 8 spontaneously vemurafenib-resistant A375 derivative cell lines

first: Adam

middle: J

last: Dupuy

first: Christopher

last: Stipp


title: ZBTB48 is a pioneer factor regulating B-cell-specific CIITA expression [RNA-seq]

city: Singapore

first: Dennis

last: Kappei

address: 14 Medical Drive, #11-01 (Bench 11), Centre for Translational Medicine

department: Cancer Science Institute of Singapore

country: Singapore

institute: National University of Singapore

summary: CIITA is the master regulator of MHC II gene expression and hence the adaptive immune response. CIITA expression itself is tightly regulated by three cell type-specific promoters, pI, pIII, and by pIV. Here, we demonstrate that the telomere-binding protein and transcriptional activator ZBTB48 directly binds to both the critical activating elements within CIITA pIII and is essential for its gene expression. ZBTB48 establishes open chromatin at CIITA pIII upstream of activating H3K4me3 modifications both priming CIITA transcription for IFNg-induction and ensuring constitutive expression in primary murine B cells. Hence, ZBTB48 acts as a molecular on-off-switch for B-cell-specific CIITA expression.

overall_design:
To comprehensively interrogate the effect of ZBTB48 on transcription upon IFNγ stimulation in U2OS cells using five WT and ZBTB48 KO clones each.
Each of the five U2OS WT and ZBTB48 KO clones were treated with 250U/ml IFNγ for 24 hours.
We globally analysed expression differences in our RNA-seq data comparing the five WT and ZBTB48 KO clones as biological replicates.

first: Grishma

last: Rane

first: Vartika

last: Khanchnadani


title: ZBTB48 is a pioneer factor regulating B-cell-specific CIITA expression [FAIRE-seq]

overall_design:
To investigate the pioneering function of ZBTB48 at promoters in U2OS cells using two WT and ZBTB48 KO clones each.
Formaldehyde assisted isolation of regulatory elements DNA-sequencing (FAIRE-seq) was performed in two U2OS WT and ZBTB48 KO clones each were treated with 250U/ml IFNγ for 24 hours.
Open chromatin was interogated in the two WT and ZBTB48 KO clones, specifically at promoters bound by ZBTB48.


title: Analysis of an HCA-7 derived cell line (CC-CR) and  cell lines generated with knockout of miR-100, miR-125b, or both miRNAs. (smallRNAseq_human_miR_KO.8556)

summary: We report the results of RNA-Seq from an HCA-7 derived cell line with high levels of miR-100 and miR-125b (CC-CR) and knockout cell lines generated from the high expressing cells. CC-CR cells had miR-100, miR-125b or both miRNAs knocked out using CRISPR Cas9.  Since miRNAs function to negatively regulate their target, the knockout cell lines should have the potential mRNA targets upregulated in comparison to CC-CR cells. Results were used to identify potential mRNAs that were upregulated in the knockout cell lines compared to CC-CR cells

overall_design: miRNAs of interest were knocked out using CRISPR-Cas9. RNAseq was conducted to determine potential targets of the miRNAs

first: James

last: Patton

first: Hannah

last: Nelson


title: Analysis of an HCA-7 derived cell line (CC-CR) and  cell lines generated with knockout of miR-100, miR-125b, or both miRNAs (RNAseq_human_miR_KO.7223)


title: Analysis of  a pair of HCA-7 derived cells with low expressing (CC) or high expressing (CC-CR) levels of miR-100 and miR-125b and RNA collected from Ago2 RNA immunoprecipitation (smallRNAseq_human_AGO2_IP.8614)

summary: We report the results of RNA-Seq from a pair of HCA-7 derived cells with low, CC, and high, CC-CR, cells as well as RNA-seq from AGO2 RNA immunoprecipitation from the two cell lines in order to identify potential targets of miR-100 and miR-125b. Results were used to identify potential mRNAs that were upregulated in CC cells (low miR-100 and  miR-125b), compared to CC-CR cells. In addition, we searched for potential mRNA targets that were enriched in the AGO2 immunoprecipitation of CC-CR cells.

overall_design: Samples were collected from CC and CC-CR whole cell lysate and from AGO2 RNA immunoprecipitation.


title: Analysis of  a pair of HCA-7 derived cells with low expressing (CC) or high expressing (CC-CR) levels of miR-100 and miR-125b and RNA collected from Ago2 RNA immunoprecipitation (RNAseq_human_AGO2_IP.8635)


title: Analysis of a pair of HCA-7 derived cells with low expressing (CC) or high expressing (CC-CR) levels of miR-100 and miR-125b and RNA collected from Ago2 RNA immunoprecipitation


title: Analysis of m6A levels in K562 treted with the FTO inhibitor FB23-3 by m6A-mRNA&lncRNA Epitranscriptomic Microarray

city: Rome

first: Alessandro

last: Fatica

address: P.ale Aldo Moro 5

department: Biology and Biotechnologies

institute: Sapienza Un iversity of Rome

summary: FTO, an N6-methyladenosine demethylase, has emerged as a promising target for the treatment of specific acute myeloid leukemia (AML) subtypes. Here, we investigate the antiproliferative effects of the FTO inhibitor FB23-2 in leukemia. We demonstrate that FB23-2 potently inhibits proliferation across both AML and CML cell lines, irrespective of their responsiveness to FTO depletion. Interestingly, FB23-2 induces cell cycle arrest without a concurrent increase in m6A levels, suggesting an alternative mechanism of action.

overall_design: Two-condition experiment, FB23-2 vs. DMSO treated cells. Biological replicates: 3 control DMSO, 3 FB23-2, independently grown and harvested. One replicate per array.

first: Francesco

last: Fazi


title: ZBTB48 is a pioneer factor regulating B-cell-specific CIITA expression.


title: ZBTB48 is a pioneer factor regulating B-cell-specific CIITA expression [ChIP-seq]

overall_design:
To investigate the effect of loss of ZBTB48 on H3K4me3 activation mark at promoters regulated by ZBTB48 upon IFNγ stimulation in U2OS cells using three WT and ZBTB48 KO clones each.
Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for histone modification H3K4me3 in each of the three U2OS WT and ZBTB48 KO clones treated with 250U/ml IFNγ for 24 hours.
We compared enrichment of H3K4me3 in the three WT and ZBTB48 KO clones as biological replicates, specifically at promoters bound by ZBTB48.