title: Land plant-specific H3K27 methyltransferases ATXR5 and ATXR6 control plant development and stress responses [H3K27me3 ChIP-seq]

city: Beijing

first: Xiaoyi

last: Li

address: west beichen

department: Chinese Academy of Sciences

institute: Institute of Genetics and Developmental Biology

summary: The colonization of land by plants relies on numerous evolutionary innovations crucial for terrestrial adaptation. These include advances in sexual reproduction and the ability to properly respond to various environmental stresses, which involve precise control of their regulatory genes. A notable genetic innovation in land plants is the emergence of histone lysine methyltransferases ATXR5/6, which specifically catalyse the repressive histone H3 lysine 27 monomethylation (H3K27me1). However, the evolutionary reason for land plants to develop these enzymes remains unclear. In this study, we investigate the importance of ATXR5/6 by generating strong atxr5;axtr6 double mutants in Arabidopsis. Our results show that ATXR5/6 are essential for plant reproductive development and play a critical role in supporting normal plant growth by repressing the transcription of stress responsive genes. In addition, ATXR5/6 are necessary for maintaining H3K27 trimethylation (H3K27me3), likely by providing H3K27me1 as a substrate for further methylation. We also demonstrate that the function of ATXR5/6 in regulating reproductive development and responsive genes is conserved in the monocot rice. We propose that land plants may have evolved ATXR5/6 to assist in the transcriptional regulation necessary for terrestrial adaptation

overall_design: The deposition of H3K27me1 by ATXR5/6 is considered replication coupled. Similarly, the maintenance of another repressive histone mark, H3K27me3, is also closely linked with DNA replication. It has been proposed that H3K27me1 deposition might facilitate the restoration of H3K27me3 after DNA replication. To assess the requirement of ATXR5/6 in H3K27me3 maintenance, we performed ChIP-seq to examine H3K27me3 profiles in WT and atxr5;atxr5;atxr6hyp;atxr6c-1

first: Danhua

last: Jiang

first: Jie

last: Pan


title: Land plant-specific H3K27 methyltransferases ATXR5 and ATXR6 control plant development and stress responses [Arabidopsis RNA-seq]

status: Public on Nov 07 2024

country: China

type: Expression profiling by high throughput sequencing

overall_design: To explore potential reasons for the additional developmental abnormalities in atxr5;atxr5;atxr6hyp;atxr6c-1 compared to atxr5;atxr6hyp


title: Jade1 and the HBO1 complex are spatial-selective cofactors of Oct4

city: Salt Lake City

first: Yifan

last: Wu

state: UT

address: 2000 Cir of Hope Dr

department: Pathology

country: USA

institute: University of Utah

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Oct4 is a master regulator of pluripotency. Potential Oct4 interactors have been cataloged extensively but the manner and significance of these interactions are incompletely defined. Like other POU domain proteins, Oct4 is capable of binding to DNA in multiple configurations, however the relationship between these configurations and cofactor recruitment (and hence transcription output) are unknown. Here, we show that Oct4 interacts with common and unique proteins when bound to DNA in different configurations. One of these proteins is Jade1, a component of the HBO histone acetyltransferase complex. Jade1 preferentially associates with Oct4 when bound to More palindromic Octamer-Related Element (MORE) DNA sequences that bind Oct4 dimers and are associated with strong gene expression. We show that the Oct4 N-terminus is critical for this interaction. ChIP-seq using HBO1, the enzymatic component of the complex, identifies a preference for binding adjacent to Oct4 at MORE sites. Using purified recombinant proteins and nucleosome complexes, we show that the HBO1 complex acetylates histone H3K9 within nucleosomes more efficiently when Oct4 is co-bound to a MORE site. Histone acetylation is further increased when Oct4 is mutated to favor dimeric MORE binding. Cryo-electron microscopy reveals that Oct4 bound to a MORE near the nucleosome entry/exit site partially unwinds DNA from nucleosome core particles, and identifies additional mass associated with the HBO1 complex. These results identify a novel mechanism of transcriptional regulation by Oct4.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for HBO1 in mouse ES cells.

first: Asit

last: Manna

first: Hiroshi

last: Handa

first: Mahesh

middle: B

last: Chandrasekharan

first: Dean

last: Tantin


title: TRPV4 activation induces TMJ degeneration via p38 MAPK/c-Fos/CXCLs axis

first: Xiaotong

last: Yu

address: 22# Zhongguancun South Avenue, Haidian District

institute: Peking university

summary: Degenerative joint disease mainly manifests abnormal bone and cartilage remodeling, articular disc deformation and synovial inflammation, and it is closely related to intraarticular overload. Activation of the mechanosensitive channel transient receptor potential vanilloid 4 (TRPV4) can lead to degeneration of the temporomandibular joint (TMJ) disc. However, the potential mechanism by which TRPV4 leads to TMJ degeneration are still unclear. The results showed that TRPV4 activation promoted upregulation of chemokines including CXCL6 and CXCL13 in disc cells, and such chemokine release facilitated the proliferation and migration of FLSs and aggravated the TMJ degeneration in rat. Mechanistically, TRPV4-induced p38 MAPK signaling pathway activation promoted chemokine expression via the nuclear translocation of p38 and c-Fos, thereby promoting the proliferation and migration by CXCL-CXCR interaction.

overall_design: The primary rat TMJ disc cells were divided into two groups: the control group and the GSK101 group. The disc cells in the GSK101 group were treated with the TRPV4 agonist GSK101 for 24 hours, while those in the control group were treated with DMSO at the same. concentration for 24 hours.

first: XiaoTong

first: ShengJie

last: Cui

first: FuJia

last: Yang

first: Yan

last: Gu


title: Keystone genes for tooth development [Vole RNA-Seq]

city: Helsinki

first: Jukka

last: Jernvall

address: P.O. Box 56 (Viikinkaari 9)

country: Finland

institute: University of Helsinki

summary: The expression levels of developing vole molar and jaw transcriptome were measured. each of first molars at embryonic days 13-16 (E13, E14, E15, E16), second molars at E16, and jaw tissues at E14.

overall_design: Wild type tooth and jaw germs were dissected from embryonic stages corresponding to E13, E14, E15, E16 development.

first: Outi

last: Hallikas

first: Rishi

last: Das Roy

first: Mona

last: Christensen


title: Distinct pathways for removal of defective RNA polymerase II transcription complexes at a promoter-proximal pause checkpointm [RN21236 dTAG TTchem-seq]

city: London

first: Richard

middle: James

last: Mitter

state: LONDON

address: 1 Midland Road

department: Bioinformatics & Biostatistics

country: United Kingdom

institute: The Francis Crick Institute

summary: The purpose and function of Integrator and RNA polymerase II (RNAPII) promoter-proximal pausing remain uncertain. Here, we show that when Integrator function is compromised by loss of INTS6, RNAPII interacts increasingly with proteins from alternative pathways for its DNA dissociation, including CUL3-ARMC5 (CRL3ARMC5), which ubiquitylates Ser5-phosphorylated Rpb1 and targets it for degradation. ARMC5-dependent RNAPII ubiquitylation is activated by defects in factors required for correctly regulated promoter-proximal pausing, including Integrator, DSIF and mRNA capping enzyme. This ARMC5 checkpoint curtails an appreciable fraction of RNAPII transcription, with ARMC5 knockout cells producing uncapped, nascent RNA transcripts that fail to mature into stable mRNA. Concomitant loss of INTS6 and ARMC5 greatly stabilizes RNAPII at the pause and has severe consequences for cell growth and viability. Our data support a model in which CRL3ARMC5 functions alongside Integrator in a checkpoint mechanism that removes faulty RNAPII complexes at promoter-proximal pause sites to safeguard transcription integrity.

overall_design: TTchem-seq (Gregersen et al., 2020) looking at the effects of rapid depletion of INTS4 in HEK293 FlpIn TREx cells, where endogenous INTS4 harbours an FKBP12F36V degron tag at its N-terminus.

first: Daniel

last: Blears

first: Jiangman

last: Lou

first: Nova

last: Fong

first: Ryan

last: Sheridan

first: David

last: Bentley

first: Jesper

last: Svejstrup


title: Ldha positively regulates the lactylation of Hdac1 to hinder the 2-cell state of embryonic stem cells [RNA-seq]

city: New York

first: XIANLE

last: SHI

state: NY

address: 168 street

institute: CUMC

summary: Protein lactylation is a process that is fueled by lactate, generated by the enzyme lactate dehydrogenase (Ldh) from pyruvate. Despite prior research, the precise role of protein lactylation in controlling the identity of mouse embryonic stem cells (ESCs) is still not fully understood. We observed that inhibiting or eliminating Ldha causes a reduction in global protein lactylation in ESCs, and RNA-seq analysis suggests that Ldha inhibition induces a 2-cell-like cell (2CLC) signature in ESCs. To probe the underlying mechanisms, we performed quantitative lactylation proteomics analysis, we discovered that Hdac1, a gene with significant regulatory roles during the 2-cell stage (2C), undergoes lactylation modification. Additionally, we observed that treatment with an Ldh (lactate dehydrogenase) inhibitor can decrease the lactylation levels of Hdac1. Mechanistically, we discovered that Ldha positively regulates the lactylation of Hdac1, promoting its direct binding to zygotic genome activation (ZGA) gene promoters and has stronger deacetylase activity. This leads to the removal of acetyl groups from H3K27 on these loci, effectively suppressing the expression of 2C genes. Our study presents novel evidence supporting protein lactylation's potential as a means of inhibiting the generation of 2CLCs and modulating acetylation activity.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for mESCs and its KO (Ldha KO, and Ldhb KO) or inhibitor treatment (SO treatment) derivatives.

first: Qiman

last: Dong

first: Lingyi

last: Chen


title: The responses of WT root and sgn3 myb36 root  inoculated with CHA0 for 0h, 3h, 6h and 10h

status: Public on Nov 06 2024

city: Shanghai

first: Yuanjie

last: Tang

state: Shanghai

address: 300 Fenglin Road, Xuhui District

department: CAS Center for Excellence in Molecular Plant Sciences

institute: CHINESE ACADEMY OF SCIENCES

summary: The casparian strip serves as a crucial diffusion barrier in the endodermis, playing a vital role in controlling the flow of substances between the root and its external environment. Here, we observed that when the casparian strip is absent, bacteria can accumulate abundantly on the roots. To investigate the impact of this bacterial accumulation on the root indued by casparian strip deficiency , we inoculated Col-0 and sgn3 myb36 roots with CHA0 in a hydroponic system. Ultimately, we found that the bacterial accumulation on the roots, resulting from the absence of the Casparian strip, strongly induces the immune response of the roots. This indicates that the casparian strip plays an important role in regulating the interaction between the root and microorganisms.

overall_design: To investigate the impact of this bacterial accumulation on the root indued by casparian strip deficiency, we inoculated Col-0 and sgn3 myb36 root with CHA0 for 0h, 3h, 6h and 10h in a hydroponic system. We then performed gene expression analysis using rna seq data from Col-0 and sgn3 myb36 root treated with CHA0 at 4 time points. For DEG analysis, data from another three time points was compared with 0h.

first: Feng

last: Zhou


first: Sasha

middle: Leilani

last: Fulton

address: 3227 Broadway

institute: Columbia University

summary: Major Depressive Disorder (MDD) arises from a complex interaction between genetics and environmental influences such as stress, which leads to persistent changes in frontolimbic gene expression and cellular function. Although relatively understudied in depression, the orbitofrontal cortex (OFC) exhibits significant changes in neuronal activity in MDD, as well as widespread gene expression changes, including consistent upregulation of inflammatory stress pathways. Inflammation signaling in non-neuronal cell-types can disrupt normal neurotransmission, though the molecular mechanisms that regulate this interaction are not well understood. Our laboratory has implemented nuclei-sorting coupled with ATAC-seq in human postmortem OFC to characterize over 200 MDD-specific open chromatin regions (OCRs), which were detected exclusively in the non-neuronal cell population. From this dataset, we identified ZBTB7A, a chromatin remodeling protein with binding motifs significantly overrepresented in these MDD-specific OCRs. ZBTB7A has been shown to orchestrate chromatin accessibility for NF-kB target genes, however, its functional role in psychiatric disease has not yet been explored. In a series of studies utilizing astrocyte-specific viral manipulations, we found that overexpressing this chromatin remodeler in rodent OFC astrocytes was sufficient to induce aberrant expression of inflammatory genes, behavioral deficits, and neuronal hyperactivity in response to a mild stressor, compared to GFP-expressing mice. Together, these findings demonstrate a novel chromatin-based mechanism for increased inflammatory signaling in astrocytes that ultimately impairs neuroadaptive responses to stress with direct implications for OFC dysfunction in MDD pathology.

overall_design: To explore the functional consequences of ZBTB7A Overexpression in rodent OFC astrocytes in the context of a mild stressor, we used an AAV6-GFAP-ZBTB7A vector that overexpresses ZBTB7A vs. GFP. This AAV6 serotype AAV vector has previously been shown to preferentially express transgenes in adult rodent astrocytes. We delivered this AAV6-GFAP-ZBTB7A vector into rodent OFC vs. GFP empty vector controls in order to evaluate the impact of overexpressing ZBTB7A in this region during a mild stressor. Mice were infected intra-OFC with one of the two viruses (on day 0), and give 3 weeks for the transgene to reach maximal expression, followed by either a 1d Subthreshold Social defeat stressor paradigm or control paradigm. Twenty-four hours after the stressor, mice were subjected to three behavioral tests (social interaction, sucrose preference, and forced swim) over a period of 7d, and then mice were sacrificed, and virally infected tissues were microdissected and processed for RNA-seq.

first: Ian

last: Maze


title: The responses of CHA0 treated with WT(Col-0) root exudates, Gln and sgn3 myb36 root exudates

summary: In our previous work, we found that the root exudates of sgn3 myb36 promoted the colonization of CHA0 on roots. Through LC-MS, we identified a large amount of glutamine (Gln) in the root exudates of sgn3 myb36. Therefore, we aim to use RNA-seq to uncover whether the root exudates of sgn3 myb36 and Gln have the same regulatory effects on CHA0. By conducting differential analysis with the CK (CHA0 treated with wild-type root exudates), we hope to identify the specific regulatory mechanisms of sgn3 myb36 and Gln on CHA0.


title: Undernutrition disrupts jejunal and ileal microbiota and epithelial tissue homeostasis in a pregnant sheep model

city: Hefei

first: weibin

last: wu

address: Changjiang Western Road 130

institute: Anhui Agricultural University

summary: In the current study, five late gestation multiparous ewes were restricted to a 30% feeding level to create a model of malnutrition, while five other ewes were fed normally as controls. All ewes were sacrificed and jejunal and ileal samples were collected for transcriptome sequencing to study metabolic changes.

overall_design: Compare the differences in transcriptome profiles between the jejunal and ileal sample from CON and TR ovis aries.

first: Weibin

last: Wu

first: Huizhen

last: Lu

first: Shengyong

last: Mao

first: Yanfeng

last: Xue


title: RNA-Seq Analysis to investigate the impact of  hypoxia on the host responses to Nontypeable Haemophilus influenzae infection in the human explanted lung

city: Oxford

first: Lee

last: Page

state: Oxfordshire

address: Centre for Clinical Vaccinology and Tropical Medicine (CCVTM), Churchill Hospital, Old Road, Headington, Oxford

department: Department of Paediatrics

institute: University of Oxford

summary: Hypoxia is a common microenvironmental condition in the mucus-obstructed, infected, and inflamed airways of lung disease patients and is known to influence immune responses in the lungs. However, the effects of hypoxia on lung host defence against respiratory bacterial infections relevant to chronic pulmonary disease are less understood. Nontypeable Haemophilus influenzae (NTHi), an opportunistic respiratory pathogen, frequently colonizes the lower airways of patients with chronic lung diseases and is associated with poor clinical outcomes. Human ex vivo lung explants were infected with a clinical strain of NTHi under normoxic and hypoxic conditions for 24 hours. The host response to infection and the effect of hypoxia were investigated using RNA sequencing.

overall_design: RNA quality was assessed using an Agilent Bioanalyzer 2100 system before sequencing of enriched poly-(A)-tailed mRNA, which was conducted by Novogene (Cambridge). Libraries were generated using the NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina® (NEB, USA), and their quality was evaluated on the Agilent Bioanalyzer 2100 system. Quantification of the libraries was performed using a Qubit 2.0 fluorometer (Life Technologies). Sequencing was executed on the NovaSeq 6000 Illumina® platform, generating 150-base pair (bp) paired-end reads at a sequencing depth of 40 million reads per sample. To ensure high-quality data reads containing adapter sequences, poly-N stretches, and low-quality reads were removed, resulting in clean reads. The Spliced Transcripts Alignment to a Reference (STAR) software (version 2.5) was utilized for mapping the reads to the human genome (GRCh38), with reads quantified using HTSeq (Anders et al., 2015).

first: Jodie

last: Ackland

first: Claire

last: Simms


title: FOXP2 expression was decreased in placental tissue sample of patients with preeclampsia compared with healthy pregnant women

city: Shenyang

first: Hong

last: Cui

address: 36# Sanhao Street

institute: Shengjing Hospital of China Medical University

summary: Preeclampsia (PE) is a common pregnancy disorder, and it complicates 5~7% of all pregnancies. At present, termination of pregnancy is the only curative strategy for PE. To explore a potential target for PE treatment, we collected placental tissue samples from patients with preeclampsia (PE) and normal pregnant women (N), and performed high-throughput RNA sequencing. We found that FOXP2 was significantly downregulated in placental samples of patients with PE compared with the controls.

overall_design: In the present study, placental tissue samples were collected from patients with PE (n = 6) and normal pregnant women (n = 6) and used for high-throughput RNA-sequencing to analyze gene expression profiles.


title: Gene expression profile (targeted mouse immune response panel) and V(D)J sequences at single cell level of day 10 post-IAV infected splenic T cells in WT mice and LCKFREE mice

city: Clayton

first: Justin

middle: Boxuan

last: Zhang

state: VIC

address: Willington road

country: Australia

institute: Monash University

summary: The LckC20.23A (LCKFREE) mutation in mice dissociates LCK from the CD4 and CD8 coreceptors. We compared the transcriptional profile of conventional splenic CD8 and CD4 T cell subsets at day 10 post-IAV infection of LCKFREE mice to WT mice to assess the functional impact of dissociating LCK from coreceptors..

overall_design: 3xC57Bl/6 and 3x LckC20.23A aged 8 weeks old were infected intranasally with 10E4 pfu HKx31 IAV infection and their spleens harvested at day 10 post-infection. Mouse splenocytes were stained and sorted using BD® FACSAria™ Fusion (BD Biosciences)  into CD8 TN, TVM, and effector/memory T (TE/EM) cells and CD4 TN and TE/EM cells based on expression of CD44, CD49d, and CD62L. Sorted populations from the same individual were pooled and sample-tagged using the BD® Single-Cell Multiplexing Kit. Single-cell capture and cDNA synthesis were performed using the BD Rhapsody™ Single-Cell Analysis System. Targeted scRNA-seq analysis was performed using the BD Rhapsody™ Immune Response Targeted Panel (mouse) containing primers of 397 immune-related genes. A paired TCRαβ sequence library was prepared using the BD Rhapsody™ Mouse TCR/BCR Amplification Kit.

first: Pirooz

last: Zareie

first: Nicole

middle: L

last: La Gruta


title: Bone marrow endosteum houses Hedgehog-susceptible Dlx5-expressing osteoblast precursor cells [Fgfr3CE_Cxcl12GFP]

city: Houston

first: Shion

last: Orikasa

state: TX

address: 7500 Cambridge St

institute: University of Texas Health Science Center at Houston School of Dentistry

summary: A single cell transcriptional profile of osteoblasts and bone marrow stromal cells of the murine femur at postnatal day 49 (P49), lineage-marked by Fgfr3-creER at P21

overall_design: GFP+ and tdTomato+ cells were FACS-isolated from Cxcl12-GFP; Fgfr3-creER; R26R-tdTomato femur bone marrow at P49 (tamoxifen at P21). 71,956 cells were loaded onto 10X Genomics Chromium Single Cell Expression Solution version 2 platform. cDNA was sequenced by Illumina HiSeq4000

first: Noriaki

last: Ono

first: Yuki

last: Matsushita


title: Opposing activity of SOX9 and Wnt in the developing otocyst ensures SOX10 control of endolymphatic development

city: Hong Kong

first: Ka Chi

last: Chu

address: 21 Sassoon Rd, Pok Fu Lam

country: Hong Kong

institute: HKU

summary: Sensorineural deafness can occur in campomelic dysplasia (CD). In a mouse model of the SOX9Y440X/+ CD mutation, SOX10 downregulation was previously shown to be associated with impaired development of the endolymphatic system at mid-gestation. The underlying molecular causes are unknown. Here we found at E10.5 when the otocyst lacks overt inner ear structures, Sox10 was downregulated in the dorsal and medial aspects of the Sox9Y440X/+ otic epithelium, regions associated with specification of the endolymphatic sac and duct.  Single-cell transcriptomic profiling revealed differential elevation of Wnt signaling pathway genes associated with the domain-specific Sox10 reduction. Otocyst-specific forced elevation of Wnt signaling by expression of stabilized β-catenin suppressed Sox10 expression. Similarly, WNT exerted an inhibitory effect on SOX10 expression in human inner ear organoids derived from pluripotent stem cells. We propose a conserved regulatory model of SOX9-WNT antagonism governing SOX10 action in the otocyst underlies the specification of endolymphatic cell fate, with important implications for understanding human deafness syndromes.

overall_design: E10.5 wild type and Sox9Y440X/+ (CD) otic vesicles (OVs) were isolated and dissociated for single-cell profiling. One additional replicate was included for the wild type.

first: Ka

middle: C

first: Kathryn

middle: E

last: Cheah


title: Land plant-specific H3K27 methyltransferases ATXR5 and ATXR6 control plant development and stress responses [Rice RNA-seq]

first: Xiaoyi

last: Li

overall_design: To test the function of ATXR5/6 in other plants, we generated the null osatxr5;osatxr6 mutant in rice and found that OsATXR5/6 are also essential for plant reproduction

first: Danhua

last: Jiang

first: Jie

last: Pan


title: HOXD1 inhibits lung adenocarcinoma progression and is regulated by DNA methylation [ChIP-seq]

city: Ji'nan

first: Lianlian

address: No. 6699 Qingdao Road

institute: Shandong First Medical University(Shandong Academy Of Medical Sciences)

summary: The homeobox (HOX) family encodes highly conserved transcription factors and plays a crucial role in embryonic development and tumorigenesis. Homeobox D1 (HOXD1) is a member of the HOX family, whose biological functions in lung cancer are unclear. The UALCAN database analysis of HOXD1 expression patterns revealed that HOXD1 is downregulated in lung adenocarcinoma patient samples relative to adjacent normal tissue. Western blotting validated low HOXD1 expression in lung adenocarcinoma (LUAD) cell lines. The Kaplan-Meier plotter database demonstrated that HOXD1 expression reduction in LUAD was found to correlate with worse overall survival. Meanwhile, in vitro study showed that HOXD1 overexpression suppressed LUAD cells proliferation, migration, and invasion. In the mouse tumor model, upregulated HOXD1 retarded tumor growth. In addition, targeted bisulfite sequencing and ChIP assay found that DNA hypermethylation occurred in the promoter region of the HOXD1 gene and was associated with DNA methyltransferases. In additionMoreover, upregulated HOXD1 as a transcriptional factor increased the transcriptional expression of BMP2 and BMP6. Taken together, the dysregulation of HOXD1 mediated by DNA methylation inhibited the initiation and progression of lung adenocarcinoma by regulating the expression of BMP/BMP6.

overall_design: To better identify the downstream target gene potentially regulated by HOXD1 as a transcription factor in LUAD, ChIP-seq was performed on HOXD1-overexpression A549 cells to analyze the downstream regulatory network of HOXD1.

first: Xin

last: Hu


title: Immune Exhaustion in ME/CFS and long COVID

city: Southport

first: Natalie

last: Eaton-Fitch

state: Queensland

address: Parklands Dr

institute: Griffith University

summary:
Gene expression analysis of RNA was performed using the commercially available NanoString® nCounter Immune Exhaustion gene expression panel (NanoString Technologies, Seattle, WA, USA). This panel contains 785 genes to elucidate mechanisms behind T cell, B cell and NK cell exhaustion in disease.
Ribonucleic acid (RNA) was extracted from peripheral blood mononuclear cells (PBMCs) isolated from ME/CFS (n=14), long COVID (n=15), and healthy control (HC; n=18) participants. ME/CFS participants were included according to Canadian Consensus Criteria for ME. Long COVID participants were eligible according to the working case definition for Post COVID-19 Condition published by the World Health Organization.

overall_design: Raw gene expression data was normalised against positive and negative controls to account for background noise and platform-associated variation. Normalisation and analysis were performed using Rosalind Bio (San Diego, CA, USA) using geometric means of housekeeping genes (ABCF1, ALAS1, EEF1G, G6PD, GAPDH, GUSB, HPRT1, OAZ1, POLR1B, POLR2A, PPIA, RPL19, SDHA, TBP, TUBB) (Supplementary Material 1). Differential expression is reported between ME/CFS and long COVID with HC.

first: Sonya

last: Marshall-Gradisnik


title: HOXD1 inhibits lung adenocarcinoma progression and is regulated by DNA methylation [TBS]

type: Methylation profiling by high throughput sequencing

overall_design: To investigate that the HOXD1 promoter undergoes DNA methylation, targeted bisulfite sequencing (TBS) was performed to detect methylation levels of HOXD1 in BEAS-2B, A549, and H1299 cells.


title: Ldha positively regulates the lactylation of Hdac1 to hinder the 2-cell state of embryonic stem cells [ChIP-seq]

first: XIANLE

last: SHI

overall_design: Chromatin immunoprecipitation followed by sequencing (ChIP–seq) for H3K27ac in embryonic stem cells

first: Qiman

last: Dong

first: Lingyi

last: Chen


title: Transcriptomic insights into Multiple System Atrophy from a PLP-α-Synuclein transgenic mouse model

city: Phoenix

first: Ignazio

middle: Stefano

last: Piras

state: Arizona

address: 445 N. Fifth Street, Phoenix

institute: Translational Genomic Research Institute

summary: Multiple system atrophy (MSA) is a rare, neurodegenerative disorder with rapid motor and non-motor symptom progression. MSA is characterized by protein aggregations of α-synuclein found in the cytoplasm of oligodendrocytes. Despite this pathological hallmark, there is still little known about the cause of this disease, resulting in poor treatment options and quality of life post-diagnosis. In this study, we investigated differentially expressed genes via RNA-sequencing of brain samples from a validated PLP-α-synuclein transgenic mouse model, identifying a total of 40 DEGs in the MSA group compared to control, with top detected genes being Gm15446, Mcm6, Aldh7a1 and Gm3435. We observed a significant enrichment of immune pathways and endothelial cell genes among the upregulated genes, whereas downregulated genes were significantly enriched for oligodendrocyte and neuronal genes. We then calculated possible overlap of these DEGs with previously profiled human MSA RNA, resulting in the identification of significant downregulation of the Tsr2 gene. Identifying key gene expression profiles specific to MSA patients is crucial to further understanding the cause, and possible prevention, of this rapidly progressive neurodegenerative disorder.

overall_design: Homozygous male and female transgenic PLP-α-synuclein mice (MGI:3604008) overexpressing human α-synuclein under the PLP promoter and age- and sex-matched non-transgenic C57Bl/6 background mice were used in this study. All animals were bred and housed in a temperature-controlled room under a 12/12 hour dark/light cycle, with free access to food and water and under special pathogen free conditions in the animal facility of the Medical University of Innsbruck. The experiments were performed in accordance with the Austrian law under permission BMWFW-66.011/0125-WF/V/3b/2015 and BMWF-66.011/0120-II/3b/2013. At the age of 12 months, mice were sacrificed under overdose of thiopental, brains were extracted, each hemisphere was snap frozen in liquid nitrogen, and kept at -80°C until further processing. RNA was isolated from hemisphere samples using the RNeasy Kit (Qiagen). Sequencing libraries were prepared with 250 ng of total RNA using Illumina Stranded Total RNA with Ribo-Zero Plus library preparation (Illumina Inc). PCR-enriched fragments were validated on a 2200 TapeStation (Agilent Technologies) and quantitated via qPCR. The final library was sequenced by 100 bp paired-end sequencing on a HiSeq 2500 (Illumina) at the Collaborative Sequencing Center (Translational Genomics Research Institute, Phoenix, AZ).

first: Leigh

last: L Nicholson

middle: S

first: Matthew

middle: D

last: DeBoth

first: Ashley

last: Siniard

first: Antonio

last: Heras-Garvin

first: Nadia

last: Stefanova

middle: J

last: Huentelman


title: Conditional knock-out of Transferrin receptor 1 in Alb-cre H11-myc mice

first: Jialei

last: Sun

address: 180 Fenglin Road

institute: Zhongshan Hospital, Fudan University

summary: Disrupted iron metabolism is commonly observed in various types of cancer. However, the role of iron signaling in controlling the tumor microenvironment and its clinical relevance remain unclear. We found that intra-tumoral iron signals stabilized PD-L1 protein, dampening CD8+ T cell responses and promoting tumor evasion. Blocking iron uptake decreased PD-L1 expression and enhanced CD8+ T cell infiltration, leading to attenuated tumor growth. Mechanistically, iron preserved PD-L1 protein integrity by inhibiting its ubiquitination. Iron-induced lipid peroxidation in tumors promoted the generation of 4-Hydroxynonenal (4-HNE), which subsequently adducted to the PD-L1 cytoplasmic domain for its carbonylation. 4-HNE-mediated carbonylation outcompeted PD-L1 ubiquitination, resulting in PD-L1 protein stabilization. Finally, we introduced a designed peptide in tumor cells to diminish PD-L1 carbonylation by competing for 4-HNE availability. This led to decreased PD-L1 expression and enhanced CD8+ T cell immunity, hindering tumor growth. Importantly, such peptide therapy showed effective outcomes in anti-PD-L1 non-responding tumors. Thus, our findings reveal alternative strategies for overcoming PD-L1-mediated immune evasion in cancer.

overall_design:
To investigate how Tfrc controls tumor microenviroment, we established liver conditional Tfrc knock-out mice and established c-Myc driven spontaneous HCC model.
We then perform mRNA-seq from bulk tumor tissue samples of 2 Tfrc knock-out mice and 2 wild type mice.
Comparative gene expresssion profiling analysis of RNA-seq data for Tfrc knock-out and wild type HCC tissues.

first: Jimin

last: Zhu

first: Xizhong

last: Shen

first: Chuan

last: Wu


title: Splenic FO B and MZ B cells from Rack1(f/f) and Rack1(f/f;CD19-Cre) mice were sorted and subjected to bulk RNA-seq

city: shihezi

first: XT

last: Z

address: shihezi

institute: Shihezi university

summary: Transcription factor Pax5 activates genes essential for B-cell development and function. However, the regulation of Pax5 expression remains elusive. Adaptor Rack1 can interact with multiple transcription factors and modulate their activation and/or stability. Despite that, its role in the transcriptional control of B-cell fates is largely unknown. Here we show that CD19-driven Rack1 deficiency leads to pro-B accumulation and simultaneous reduction of B cells at later developmental stages. The generation of bone marrow chimeras indicates a cell-intrinsic role of Rack1 in B-cell homeostasis. Moreover, Rack1 augments BCR and TLR signaling in mature B cells. Based on diminished CD19 expression upon Rack1 deficiency, further exploration reveals that Rack1 maintains Pax5 protein levels through direct interaction and consequent prevention of Pax5 ubiquitination. Accordingly, Mb1-driven Rack1 deficiency almost completely blocks B-cell development at the pro-B cell stage. Thus, Rack1 regulates B-cell development and function through, at least partially, binding to and stabilizing Pax5.

overall_design: To explore the gene clusters affected by the specific knockdown of Rack1 in B cells and identify the possible target genes.

first: Jiyan

last: Zhang

first: Xueting

first: Chenke

last: Ma

first: Yucheng

last: Lu


title: Antiallergic Effects of Luteolin in IgE-Induced bone marrow-derived mast cells

city: Gangneung

first: Myungsuk

last: Kim

address: 679, Saimdang-ro, Gangneung-si, Gangwon-do, Republic of Korea

country: South Korea

institute: KIST

summary: This study investigated the antiallergic effects of Chamaecrista nomame extract (CN) and its isolated compound luteolin in a mouse model of ovalbumin (OVA)-induced asthma.  We aimed to identify active substances within CN, elucidate their molecular signaling mechanisms, and assess their therapeutic potential for treating allergic asthma.

overall_design: We screened 302 plant extracts for anti-degranulation activity, identifying CN as a promising candidate. In vitro assessments on RBL-2H3 cells and a mouse model of OVA-induced asthma were used to evaluate CN's effects.  RNA sequencing and gene expression analysis in bone marrow-derived mast cells (BMMCs) were conducted to explore the molecular mechanisms.  Luteolin was identified as a key bioactive component of CN.

first: Wook-Bin

last: Lee


title: A novel method for characterizing cell-cell interactions at single-cell resolution reveals unique immune signatures in blood T cell-monocyte complexes during infection (Dengue scRNA-Seq)

city: La Jolla

first: Ningxin

last: Kang

state: California

address: 9420 Athena Cir

institute: La Jolla Institute for Immunology

summary: We designed a novel method to study the transcriptome of single cells forming complexes in a high-throughput fashion, without the need for bioinformatic deconvolution. We applied this method to the study of T cells and monocytes forming complexes in blood isolated from individuals with dengue fever, with blood collected at the acute and convalescent phase of infection. We found that the transcriptomic signatures of T cells and monocytes forming complexes significantly overlapped with those identified in T cell-monocyte complexes in active tuberculosis, with upregulation of genes associated with T cell activation, cytotoxicity, cell adhesion and MHC-II. Thus, using our novel method, we identified that T cells and monocyte forming complexes in blood hold unique immune signatures distinct from singlets, and that they may be a previously overlooked valuable biomarker to monitor immune synaptic interactions during infection.

overall_design:
PBMC were isolated, cryopreserved and thawed on the day of the experiment. T cell-monocyte complexes were identified by flow cytometry as live CD3+CD14+CD19- events and sorted in bulk. Cell sorting effectively disrupts the physical connection between cells forming complexes, with the vast majority of resulting cells being singlet CD3+ or singlet CD14+ cells remaining in suspension. The single-cell suspension was then used for droplet single-cell sequencing using the 10X genomics platform.
-----------------------------------
Authors state the following concerning the raw data "privacy concerns".

first: Julie

middle: G

last: Burel

first: Gregory

last: Seumois

first: Ashu

last: Chawla

first: Bjoern

last: Peters


title: Next Generation Sequencing Facilitates Quantitative Analysis of Bone Marrow-Derived Macrophages Transcriptomes for Acly-E14 ASO effects

first: Xingguang

last: Liu

address: Xiangyin road 800

department: National Key Laboratory of Immunity & Inflammation

institute: Naval medical university

summary: Purpose: The purpose of this study is to detect activated or silenced genes during  bone marrow derived macrophages (BMDMs) transfected with control siRNA or Acly-E14 siRNA. Gene expression differences between two samples could be found using transcriptome profiling (RNA-seq) analysis. Methods: Mouse BMDMs were generated from bone marrow cells in RPMI-1640 medium with recombinant mouse M-CSF (20ng/ml). BMDMs were stained to confirm the surface expression of CD11b and F4/80. Cells with purity >97.5% were used for subsequent experiments. BMDMs were transfected with control ASO or Acly-E14 ASO. 48 hour later, they were stimulated with LPS (100ng/ml) for 4 hours, of which RNA profiles were generated by deep sequencing, using Illumina. Results: We mapped about 10 million sequence reads per sample to the mouse genome, identified hundreds of genes with significant mRNA variation between BMDMs transfected with the indicated ASOs.

overall_design: mRNA profiles of BMDMs transfected with control ASO or Acly-E14 ASO were generated by deep sequencing.

first: Yunkai


title: Effect of deleption of BnaA06FAH and BnaC05FAH on gene expression during programmed cell death in brassica napus.L

city: yichun

first: tiantian

last: zhi

state: jiangxi

address: 576 Xuefu Road, Yichun City, Jiangxi Province

institute: College of Life Sciences and Resources and Environment, Yichun University

summary: Programmed cell death (PCD) is a genetically controlled pathway that plants can use to selectively eliminate redundant or damaged cells.The coordinated and sequential actions of lineage-specific transcription factors and epigenetic regulators are essential for the initiation of PCD. We used CRISPR/Cas9-mediated targeted mutagenesis to generate a series of transgene-free mutant lines either with single or double-null bnafah alleles. Among these mutant lines, the bnafah (deleption of BnaA06FAH and BnaC05FAH) double-null mutant line, rather than the bnaa06fah (deleption of BnaA06FAH) mutant line, exhibited PCD under short day (SD) condition. To gain deeper mechanistic understanding of PCD process, transcriptomic analyses of bnaa06fah and bnafah under LD(SD0), SD1 and SD2 were performed. Heatmap cluster of DEGs speculated that PCD may related to various redox regulation genes involved in antioxidant, ROS-responsive regulator and calcium signaling. Combined with the results of previous studies, our work showed that the expression levels of BnaC04CAT2, BnaA09/C09SAL1, BnaA08/C08ACO2, BnaA07/C06ERO1, BnaA08ACA1, BnaC04BIK1, BnaA09CRK36 and BnaA03CPK4 were significantly different and might be candidate hub genes of PCD.

overall_design: To investigate deleption of both BnaA06FAH and BnaC05FAH or single BnaA06FAH in regulating programmed cell death, We used CRISPR/Cas9-mediated targeted mutagenesis to generate a series of transgene-free mutant lines either with single or double-null bnafah alleles. To gain deeper mechanistic understanding of PCD process, transcriptomic analyses of bnaa06fah (deleption of BnaA06FAH) and bnafah (deleption of BnaA06FAH and BnaC05FAH) under different SD stages were performed.

first: Tiantian

last: Zhi

first: Zhou

last: Zhou

first: Jie

last: Zou

first: Gang

last: Chen


title: Characteristic disease defects in circulating endothelial cells isolated from patients with pulmonary arterial hypertension

city: Cleveland

first: Kulwant

middle: Singh

last: Aulak

state: Ohio

address: 9500 Euclid Ave

department: Immunology and Immunity

institute: cleveland Clinic

summary: Pulmonary arterial hypertension (PAH) is a progressive disease characterized by elevated pulmonary arterial pressures that can lead to right heart failure and death. No cure exists for this disease, but therapeutic advancements have extended its median survival from 2 to 7 years. Mechanistic research in PAH has been limited by factors including that a) animal models do not fully recapitulate the disease or provide insights into its pathogenesis, and b) cellular material from PAH patients is primarily obtained from donor lungs during autopsy or transplantation, which reflect end-stage disease. Therefore, there is a need to identify tools that can elucidate the specific mechanisms of human disease in individual patients, a critical step to guide treatment decisions based on specific pathway abnormalities. Here we demonstrate a simple method to isolate and culture circulating endothelial cells (CECs) obtained at the time of right heart catheterization in PAH patients. We tested these CECs using transcriptomics and found that they have typical traits of PAH, including those involving key treatment pathways, i.e. nitric oxide, endothelin, prostacyclin and BMP/activin pathways. CECs show important gene expression changes in other central PAH disease pathways. In summary, we present a new cellular model for the ex-vivo mechanistic evaluation of critical PAH pathways that participate in the pathogenesis of the disease and may help personalized therapeutic decisions.

overall_design: To determine whether circulating endothelial cells from the blood of PAH patients have expression profiles similar to those of endothelial cells from the lungs of PAH patients, compared to endothelial cells from control (non-PAH) lungs.

first: Lori

last: Mavarakis

first: Liping

last: Tian

first: Deborah

last: Paul

first: Suzy

middle: A

last: Comhair

first: Raed

last: Dweik

first: Adriano

middle: R

last: Tonelli


title: Tretinoin synergistically enhances the antitumor effect of combined BRAF, MEK, and EGFR inhibition in BRAFV600E colorectal cancer [RKO samples]

city: Sendai

first: Yuya

last: Yoshida

state: Miyagi

address: Seiryomachi 4-1, Aoba-ku

department: clinical oncology

country: Japan

institute: Tohoku University Hospital

summary: Patients with BRAF-mutated colorectal cancer (BRAFV600E CRC) are currently treated by a combination of BRAF inhibitor and anti-EGFR antibody with or without MEK inhibitor. A fundamental problem in treating patients with BRAFV600E CRC is intrinsic and/or acquired resistance to this combination therapy. By screening 78 compounds, we identified tretinoin, a retinoid, as a compound that synergistically enhances the antiproliferative effect of a combination of BRAF inhibition and MEK inhibition with or without EGFR inhibition on BRAFV600E CRC cells. This synergistic effect was also exerted by other retinoids. Tretinoin, added to BRAF inhibitor and MEK inhibitor, upregulated PARP, BAK, and p-H2AX. When either RARα or RXRα was silenced, the increase in cleaved PARP expression by the addition of TRE to ENC/BIN or ENC/BIN/CET was canceled. Our results suggest that the mechanism of the synergistic antiproliferative effect involves modulation of the Bcl-2 family and the DNA damage response that affects apoptotic pathways, and this synergistic effect is induced by RARα- or RXRα-mediated apoptosis. Tretinoin also enhanced the antitumor effect of a combination of BRAF inhibitor and anti-EGFR antibody with or without MEK inhibitor in a BRAFV600E CRC xenograft mouse model. Our data provide a rationale for developing retinoids as a new combination agent to overcome resistance to the combination therapy for patients with BRAFV600E CRC.

overall_design: Microarray analysis was performed to confirm whether tretinoin induced different gene expression changes depending on the presence or absence of encorafenib/binimetinib. RKO cells were incubated with DMSO, tretinoin, encorafenib/binimetinib, and tretinoin/encorafenib/binimetinib for 24 hours. After incubation, total RNA of the cells was extracted. We compared between the genes differentially expressed by DMSO- and TRE-treated cells, and the ones differentially expressed by ENC/BIN- and TRE/ENC/BIN-treated cells.

first: Masanobu

last: Takahashi

first: Sakura

last: Taniguchi

first: Ryunosuke

last: Numakura

first: Keigo

last: Komine

first: Chikashi

last: Ishioka


title: Circ-CAMTA1 regulated by Ca2+ influx inhibited pyruvate carboxylase activity and modulate T cell function in patients with systemic lupus erythematosus

city: Chia-Yi

first: Hui

middle: Chun

last: Yu

address: No. 2, Min-Sheng Rd, Dalin Town

department: Medical research

country: Taiwan

institute: Dalin Tzu Chi Hospital, Buddhist Tzu Chi Medical Foundation

summary: The expression profiles of circRNAs in Jurkats cells, co-cultured with and without ionomycin, were analyzed by next-generation sequencing and validated using real-time polymerase chain reaction. The identified Ca2+ influx-regulated circRNAs were further examined in T cells from 42 patients with SLE and 23 healthy controls. The biological function of specific circRNAs was investigated using transfection and RNA pull-down assay. After validation, we confirmed that the expression levels of circ-ERCC4, circ-NFATC2, circ-MYH10, circ-CAMTA1, circ-ASH1L, circ-SOCS7, and circ-ASAP1 were consistently increased in Jurkat cells following Ca2+ influx. The expression levels of circ-CAMTA1, circ-ASH1L, and circ-ASAP1 were significantly lower in T cells from patients with SLE, with even lower levels observed in those with higher disease activity. Interferon (IFN)-α was found to suppress the expression of circ-CAMTA1. Circ-CAMTA1 bound to pyruvate carboxylase and inhibited its biologic activity. Overexpression of circ-CAMTA1, but not its linear form, significantly decreased extracellular glucose levels. Furthermore, increased expression of circ-CAMTA1, but not its linear form, enhanced IL-2 secretion and the protein expression of NFAT1.

overall_design: For analysis the expression levels of circRNAs in Jurkat cells after stimulation, Jurkat cells were treated with either phorbol 12-myristate 13-acetate (PMA; 20 ng/mL) and inomycin (500 ng/mL) for 24 hrs.

first: HuiChun

first: HsienYu

last: Huang Tseng

first: HsienBin

last: Huang

first: MingChi

last: Lu


title: Super-enhancer driven LIF/LIFR-STAT3-SOX2 regulatory feedback loop promotes cancer stemness in head and neck squamous cell carcinoma (ATAC-seq)

city: Nanjing

first: Jin

last: Li

address: 140, Hanzhong Road

department: OMFS

institute: Nanjing Medical University

summary: Super-enhancers (SEs) have been recognized as key epigenetic regulators underlying cancer stemness and malignant traits by aberrant transcriptional control and promising therapeutic targets against human cancers. However, the SE landscape and their roles during head and neck squamous cell carcinoma (HNSCC) development especially in cancer stem cells (CSCs) maintenance remain underexplored yet. Here, we identified leukemia inhibitory factor (LIF)-SE as a representative oncogenic SE to activate LIF transcription in HNSCC. LIF secreted from cancer cells and cancer-associated fibroblasts promotes cancer stemness by driving SOX2 transcription in an autocrine or paracrine manner, respectively. Mechanistically, enhancer elements E1, 2, 4 within LIF-SE recruited SOX2/SMAD3/BRD4/EP300 to facilitate LIF transcription; LIF activated downstream LIFR-STAT3 signaling to drive SOX2 transcription, thus forming a previously unknown regulatory feedback loop (LIF-SE-LIF/LIFR-STAT3-SOX2) to maintain LIF overexpression and CSCs stemness. Clinically, increased LIF abundance in clinical samples correlated with malignant clinicopathological features and patient prognosis; higher LIF concentrations in presurgical plasma were negatively associated with patient survival and dramatically diminished following cancer eradication. Therapeutically, pharmacological targeting LIF-SE-LIF/LIFR-STAT3 significantly impaired tumor growth and reduced CSC subpopulations in HNSCC xenograft and PDX models. Our findings reveal a hitherto uncharacterized LIF-SE-mediated auto-regulatory loop in regulating HNSCC stemness and highlight LIF as a novel non-invasive biomarker and potential therapeutic target for HNSCC.

overall_design: transposase-accessible chromatin with sequencing (ATAC-seq) in Cal27-nonCSC and -CSC cells

first: Yuhan

last: Wang

first: Ziyu

first: Yuxiang

last: Wei

first: Pengfei

last: Diao

first: Yaping

last: Wu

first: Dongmiao

first: Hongbing

last: Jiang

first: Yanling

first: Jie

last: Cheng


title: Super-enhancer driven LIF/LIFR-STAT3-SOX2 regulatory feedback loop promotes cancer stemness in head and neck squamous cell carcinoma (ChIP-seq)

overall_design: BRD4 as well as EP300 ChIP-seq in human head and neck squamous cell carcinoma cell line


title: Metabolic programs drive function of therapeutic natural killer (NK) cells in low oxygen tumor microenvironments

city: Minneapolis

first: Philippa

last: Kennedy

state: Minnesota

address: 425 East River Parkway, Rm 660A

department: Medicine

institute: University of Minnesota

summary: Limited oxygen in solid tumors poses a challenge to successful immunotherapy with NK cells. NK cells have impaired cytotoxicity when cultured in hypoxia (1% oxygen), but not physiologic or atmospheric oxygen (>5%). We sought to analyze the impact of oxygen concentrations on human NK cell biology to determine what factors were driving loss of cytotoxicity and where there were opportunities to overcome hypoxia-induced dysfunction in NK cells.

overall_design:
Human NK cells were enriched from healthy blood by magnetic beads selection and then incubated in advanced incubators that were humidified and maintained at 37 degrees Celsius, 5% CO2 with different oxygen (O2) and pressure conditions. Incubators were set to model oxygen and pressure levels found (1) in arterial blood (12% O2, + 2 psi), (2) in bone marrow (5% O2, + 0.6 psi), or (3-4) within the tumor microenvironment (1% O2) at low or high pressure (+ 0.3 psi or + 2 psi). NK cells incubated in a conventional humidified, 5% CO2 incubator (20% O2) served as a control (5).
NK cells were extracted after 1, 3 or 7 days in culture.
4-5 biological replicates per condition were obtained from 6 independent donors

first: Yunmin


title: A single-cell RNA sequencing atlas of the COPD distal lung to predict cell-cell communication

city: Nashville

first: Jessica

last: Blackburn

address: 1611 21st Ave MCN

institute: VUMC

summary: In the lungs of chronic obstructive pulmonary disease (COPD) patients, numerous cell types interact in a structurally abnormal and inflammatory microenvironment. Assessing gene expression in individual cells by single-cell RNA sequencing (scRNA-seq) allows for a deeper understanding of cell types within a complex sample, like the COPD distal lung. ScRNA-seq data can be combined with cell-cell communication prediction tools to predict cellular interactions within a given sample; indeed, this approach was previously used to show evidence for increased endothelial CXCL signaling in COPD. To maximize the value of these tools, it is critical to have representation across cell lineages (e.g., epithelial, immune, endothelial, and mesenchymal). However, this can be challenging in diseases like COPD where inflammatory cells often dominate samples compared to less frequent cell types that are also perturbed in disease, such as the airway epithelium. To explore intercellular interactions in the COPD lung, we performed scRNA-seq on single cell suspensions from 6 COPD distal lung samples then combined these with previously published data to assemble a multisite COPD and control scRNA-seq dataset with increased representation of non-immune lineages.

overall_design: Distal lung tissue of 6 former smokers with COPD who underwent lung transplantation (3 male, 3 female, non-Hispanic white, mean age 58.5 years, mean pack-years of smoking 40.3) was finely minced then cryopreserved in 90%FBS and 10% DMSO until processed for sequencing. The first 3 samples and the last three samples were batched together. Aliquots from each lung sample were digested in an enzymatic cocktail (Miltenyi Multi Tissue Dissociation Kit 1) using a gentleMACS Octo Dissociator (Miltenyi, Inc.). Tissue homogenates were serially filtered through sterile gauze, then 100 μm, and  40-μm sterile cell strainers. Single-cell suspensions then were treated with ACK buffer to lyse blood cells and then underwent cell sorting using EPCAM magnetic bead based selection. The EPCAM enriched fraction was used as the input for the single cell libraries.

first: Tiffany

last: Tufenkjian

first: Bradley

last: Richmond


title: Single-cell RNA-sequencing analyses on pancreatic tumors from KPPC mice and PPSSC mice

first: Yang

last: Chen

address: 6565 MD Anderson Blvd

department: Translational Molecular Pathology

institute: MD Anderson Cancer Center

summary: Single-cell RNA-sequencing analyses were performed on pancreatic tumors from KPPC mice and PPSSC mice, so as to identify the changes of tumor immune microenvironment upon Trp53 and Smad4 deletion without KRAS mutation.

overall_design: We generated a novel transgenic mouse model (Trp53loxP/loxP;Smad4loxP/loxP;Pdx1-Cre) that harbors wild-type KRAS and loss of Trp53/Smad4 in the context of spontaneous PDAC. Pancreatic tumors from PPSSC were analyzed by 10x single-cell RNA-seq.

first: Daowei

last: Yang

first: Xinlei

last: Sun


title: Effect of resistant-derived extracellular vesicles on breast cancer cells acquired drug resistance process

city: Ribeirao PReto

first: Patrick

last: Santos

address: 3900 Bandeirantes Ave

country: Brazil

institute: USP

summary: The acquired resistance that tumors develop during the treatment is still a central cause of cancer-associated deaths. Our study investigates the transcription changes modulated by EVs from tamoxifen- and doxorubicin-resistant breast cancer cells in sensitive cells and how these changes may induce acquired drug resistance, inhibit apoptosis, and increase survivability in the sensitive cells.

overall_design: To better understand the influence of drug-resistant EVs on the sensitive MCF-7 and MDA-MB-231 transcriptomes, we performed RNA sequencing (RNA-Seq) analysis in these two cell lines after exposure to tamoxifen-resistant and doxorubicin-resistant EVs for 24 h. The transcriptomic profilling was performed using 3 biological replicates of RNA from sensitive MCF-7 and MDA-MB-231 cells in comparisson with RNA from tamoxifen and doxorubicin-resistant counterparts.

first: Fausto

last: Almeida


title: Single-nucleus RNA sequencing of human subcutaneous adipose tissue biopsies from 68 Finnish participants.

city: Los Angeles

first: Seung Hyuk

middle: T.

last: Lee

state: CA

address: 695 Charles E. Young Drive South

institute: UCLA

summary: We performed single-nucleus RNA-seq on subcutaneous adipose tissue biopsies of 68 participants from the Roux-en-Y versus one-anastomosis gastric bypass (RYSA) cohort.

overall_design:
Frozen subcutaneous adipose tissue biopsies from 68 donors were combined into batches with a pool of 8 samples per batch. We randomly assigned samples to batches multiple times and selected the set that results in a minimal correlation between batches and phenotypic traits. Nuclei were then isolated from each batch and single nucleus sequencing libraries were constructed using the Chromium Single Cell 3’ v3.1 chemistry. We sequenced all libraries together on an Illumina NovaSeq S4 with a target sequencing depth of 400 million read pairs per pool.

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Submitter states that missing raw files are due to patient privacy concerns.
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first: SeungHyuk

middle: T

first: Sini

last: Heinonen

first: Sandhya

last: Rajkumar

first: Kirsi

middle: H

last: Pietiläinen

first: Päivi

last: Pajukanta


title: Thymocytes TCR repertoire from regular B6 and Btla KO mice

city: Edmonton

first: Colin

last: Anderson

state: Alberta

department: Surgery

country: Canada

institute: University of Alberta

summary: Deletion of BTLA up regulates CD5 expression. Higher CD5 expression during thymic selection is associated with increased self-recognition, suggesting that BTLA might be needed early to establish self-tolerance. To investigate whether the regulation of CD5 by BTLA is TCR-repertoire independent, we performed single-cell TCR sequencing in SP CD4 thymocytes from regular B6 and B6 Btla Knock-Out (KO) mice.

overall_design: CD4 single positive (SP) T cells were isolated from single cell thymocyte suspensions prepared from B6.Btla WT and B6.Btla KO mice (aged 8-12 weeks). TCR sequencing was conducted using the 10x Genomics V(D)J workflow.

first: Marcos

middle: I

last: Petersen

first: Adeolu

middle: O

last: Adegoke

first: Kevin

last: Joannou

first: Troy

last: Baldwin


title: Tretinoin synergistically enhances the antitumor effect of combined BRAF, MEK, and EGFR inhibition in BRAFV600E colorectal cancer [HT29 samples]

first: Yuya

last: Yoshida

overall_design: Microarray analysis was performed to confirm whether tretinoin induced different gene expression changes depending on the presence or absence of encorafenib/binimetinib. HT29 cells were incubated with DMSO, tretinoin, encorafenib/binimetinib, and tretinoin/encorafenib/binimetinib for 24 hours. After incubation, total RNA of the cells was extracted. We compared between the genes differentially expressed by DMSO- and TRE-treated cells, and the ones differentially expressed by ENC/BIN- and TRE/ENC/BIN-treated cells.

first: Masanobu

last: Takahashi

first: Sakura

last: Taniguchi

first: Ryunosuke

last: Numakura

first: Keigo

last: Komine

first: Chikashi

last: Ishioka


title: A novel method for characterizing cell-cell interactions at single-cell resolution reveals unique immune signatures in blood T cell-monocyte complexes during infection (ATB scTCR-Seq)

first: Ningxin

last: Kang

summary: We designed a novel method to study the transcriptome of single cells forming complexes in a high-throughput fashion, without the need for bioinformatic deconvolution. We applied this method to the study of T cells and monocytes forming complexes in blood isolated from individuals with active tuberculosis, with blood collected at diagnosis and after treatment. We found that the transcriptomic signature of T cells and monocytes forming complexes was associated with TCR/MHC-II immune synaptic features, enrichment for cytotoxic clonally expanded T cells and intermediate monocytes, and - especially at diagnosis - transcriptomic signatures indicating increased immune and metabolic activity. We also found a striking enrichment of “dual-expressing” cells (i.e., co-expressing T cells and monocyte marker genes) in complexes, suggesting RNA exchange occur during T cell-monocyte interactions. Thus, using our novel method, we identified that T cells and monocyte forming complexes in blood hold unique immune signatures distinct from singlets, and that they may be a previously overlooked valuable biomarker to monitor immune synaptic interactions during infection.

first: Julie

last: Burel

first: Gregory

last: Seumois

first: Ashu

last: Chawla

first: Bjoern

last: Peters


title: A novel method for characterizing cell-cell interactions at single-cell resolution reveals unique immune signatures in blood T cell-monocyte complexes during infection (Dengue scTCR-Seq)


title: Novel mechanisms of MITF regulation identified in a mouse suppressor screen

city: Reykjavik

first: Hong Nhung

last: Vu

address: Sturlugata 8

country: Iceland

institute: University of Iceland

summary: MITF, a basic-Helix-Loop-Helix Zipper (bHLHZip) transcription factor, plays vital roles in melanocyte development and functions as an oncogene. We performed a genetic screen for suppressors of the Mitf-associated pigmentation phenotype in mice and identified an intragenic Mitf mutation that terminates MITF at the K316 SUMOylation site, leading to loss of the C-end intrinsically disordered region (IDR). The resulting protein is more nuclear but less stable than wild-type MITF and retains DNA-binding ability. Interestingly, as a dimer, it can translocate wild-type and mutant MITF partners into the nucleus, improving its own stability thus ensuring nuclear MITF supply. To further understand the effect of the suppressor mutation (MITF-sl), we performed CUT&RUN experiments to determine how MITF-sl affects the genome-wide occupancy of MITF. MITF-sl had a higher number of peaks than MITF-WT, with 10,636 peaks (p<0.01) exhibiting statistically significant differences (p < 0.01) between MITF-WT and MITF-sl. Gene ontology analysis indicates that the peaks significantly different between MITF-WT and MITF-sl are associated with genes involved in axonogenesis, axon development, cell growth, and positive regulation of MAPK cascade biological pathways. Our results suggest that the 316-419 domain is critical for selective genome occupancy and transcriptional activation of MITF.

overall_design: Cleavage Under Targets and Release Using Nuclease CUT&RUN experiments were performed as previously described (Skene and Henikoff, 2017; Zhu et al., 2019) with minor modifications. A375P cells were induced to express FLAG-tagged MITF mutant fusion proteins (MITF-WT and MITF-sl) for 48 hours.

first: Nhung

first: Eiríkur

last: Steingrímsson


title: The STAT3-SETDB2 Axis Dictates NFκB-mediated Inflammation in Macrophages during Wound Repair

city: Ann Arbor

first: Poulami

last: Dey

address: 1301 Catherine St

institute: Michigan Medicine

summary: Macrophage transition from an inflammatory to reparative phenotype after tissue injury is controlled by epigenetic enzymes that regulate inflammatory gene expression. We have previously identified the histone methyltransferase, SETDB2, in macrophages drives tissue repair by repressing NFκB-mediated inflammation. Complementary ATAC and RNA sequencing of wound macrophages isolated from mice deficient in SETDB2 in myeloid cells revealed that SETDB2 suppresses the inflammatory gene program by inhibiting chromatin accessibility at NFkB-dependent gene promoters. We found that STAT3 was required for SETDB2 expression in macrophages yet, paradoxically, it also functioned as a binding partner of SETDB2 where it repressed SETDB2 activity by inhibiting its interaction with the NFKB component, RELA, leading to increased RELA/NFKB-mediated inflammatory gene expression. Further, RNA sequencing in wound macrophages from STAT3-deficient mice corroborated this and revealed STAT3 and SETDB2 transcriptionally co-regulate overlapping genes. Finally, in diabetic wound macrophages STAT3 expression and STAT3-SETDB2 binding were increased. As such, we identify what we believe to be a novel SETDB2-STAT3 axis that modulates macrophage phenotype during tissue repair and may be an important therapeutic target for nonhealing diabetic wounds.

overall_design: For ATAC-seq, wound macrophages were harvested from mice at day 5 as described above using MACS and nuclei were isolated as described previously (45). After counting, 50K nuclei per sample were resuspended in the appropriate volume, spun down, and collected for use in downstream transposition reaction per modified ENCODE protocol (46). Data was obtained and processed according to the ENCODE pipeline and then converted to readable format as similarly described. Briefly, minimum read length prior to trimming was 45 base pairs. Paired-end sequencing was performed using NovaSeq (Illumina) and sequences were mapped to the genome. For RNA-seq, day 5 wounds were harvested from Setdb2flox/floxLyz2Cre, Stat3flox/floxLyz2Cre, and cre-negative littermate control mice, digested, and underwent negative selection for CD3, CD19, and Ly6G lineage markers. Remaining cells were surface stained and sorted by FACs into CD11b+Ly6CHi and CD11b+Ly6CLo populations. RNA isolation was performed using a RNeasy Kit (Qiagen) with DNase digestion. Library construction and analysis of reads was performed as described previously (41). Briefly, reads were trimmed using Trimmomatic and mapped using HiSAT2 (47, 48). Read counts were performed using the feature-counts option from the subRead package followed by the elimination of low reads, normalization and differential gene expression using edgeR (49, 50). Differential expression was performed on mapped reads using the taqwise dispersion algorithm in edgeR.

first: Johann

last: Gudjonsson

first: Lam

last: Tsoi

first: Kathy

last: Gallagher


title: A novel method for characterizing cell-cell interactions at single-cell resolution reveals unique immune signatures in blood T cell-monocyte complexes during infection (ATB scRNA-Seq)

overall_design:
PBMC were isolated, cryopreserved and thawed on the day of the experiment. T cell-monocyte complexes were identified by flow cytometry as live CD3+CD14+CD19- events and sorted in bulk. Cell sorting effectively disrupts the physical connection between cells forming complexes, with the vast majority of resulting cells being singlet CD3+ or singlet CD14+ cells remaining in suspension. The single-cell suspension was then used for droplet single-cell sequencing using the 10X genomics platform.
-----------------------------------
Authors state the following concerning the raw data "privacy concerns".


title: SMYD1 modulates the proliferation of multipotent cardiac progenitor cells derived from human pluripotent stem cells during myocardial differentiation through GSK3β/β-catenin/ERK signaling

city: Beijing

first: Yun

last: Chang

address: 167 Beilishi Road, Xicheng District

institute: Fuwai hospital

summary: Background: The histone-lysine N-methyltransferase SMYD1, which is specific to striated muscle, plays a crucial role in regulating early heart development. Its deficiency has been linked to the occurrence of congenital heart disease. Nevertheless, the precise mechanism by which SMYD1 deficiency contributes to congenital heart disease remains unclear. Methods: We established a SMYD1 knockout pluripotent stem cell line and a doxycyclineinducible SMYD1 expression pluripotent stem cell line to investigate the functions of SMYD1 utilizing an in vitro-directed myocardial differentiation model. Results: Cardiomyocytes lacking SMYD1 displayed drastically diminished differentiation efficiency, concomitant with heightened proliferation of cardiac progenitor cells during the early cardiac differentiation stage. These cellular phenotypes were confirmed through experiments inducing the re-expression of SMYD1. Transcriptome sequencing and small molecule inhibitor intervention suggested that the GSK3β/β-catenin/ERK signaling pathway was involved in the proliferation of cardiac progenitor cells. Chromatin immunoprecipitation demonstrated that SMYD1 acted as a transcriptional activator of GSK3β through histone H3 Lysine 4 trimethylation. Additionally, dualluciferase analyses indicated that SMYD1 could interact with the promoter region of GSK3β, thereby augmenting its transcriptional activity. Moreover, administering insulin and Insulin-like growth factor 1 can enhance the efficacy of myocardial differentiation in SMYD1 knockout cells. Conclusions: Our research indicated that the participation of SMYD1 in the GSK3β/β-catenin/ERK signaling cascade modulated the proliferation of cardiac progenitor cells during myocardial differentiation. This process was partly reliant on the transcription of GSK3β. Our research provided a novel insight into the genetic modification effect of SMYD1 during early myocardial differentiation. The findings were essential to the molecular mechanism and potential interventions for congenital heart disease.

overall_design: To investigate the function SMYD1 in human pluripotent stem cell differentiation into cardiomyocytes, we established SMYD1 knockout cell lines through the Crispr Cas9 system. We then performed gene expression analysis using data obtained from RNA-seq of 2 different cells at cardiac progenitor cells stage during myocardial differentiation. Comparative gene expression profling analysis of RNA-seq data for hESC cells and SMYD1 knockout cells at cardiac progenitor cells stage during myocardial differentiation.

first: Youxu

last: Jiang


title: A novel method for characterizing cell-cell interactions at single-cell resolution reveals unique signatures in blood T cell-monocyte complexes during infection

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

overall_design: Refer to individual Series


title: Eldecalcitol ameliorates diabetic osteoporosis and glucolipid metabolic disorder by promoting Treg cell differentiation through SOCE

city: Jinan

first: Duan

last: Gao

address: Jinan

institute: Jinan

summary: Adaptive immunity plays a key role in osteoporosis in type 2 diabetes mellitus (T2DM); eldecalcitol (ED-71) is a novel active vitamin D analog, but its specific immunological mechanisms in ameliorating diabetic osteoporosis has not been well defined. In a T2DM mouse model, ED-71 attenuated bone loss and marrow adiposity. Simultaneously, it rectified imbalanced glucose homeostasis and dyslipidemia, ameliorated pancreatic β-cell damage and hepatic glycolipid metabolism disorder. Subsequently, in T2DM mice injected with CD25, we observed that the beneficial effects of ED-71 mentioned earlier were partially contingent on the Treg subsets ratio. Mechanistically, ED-71 promoted the differentiation of CD4+ T cells into Treg subsets, facilitating Ca2+ influx and the expression of ORAI1 and STIM1, pivotal proteins in store-operated Ca2+ entry (SOCE). The SOCE inhibitor, 2-APB, partially attenuated the positive effects of ED-71 observed in the above results. Together, these findings unveil ED-71 regulates SOCE-mediated Treg cell differentiation, accomplishing the dual purpose of simultaneously ameliorating diabetic osteoporosis and glucolipid metabolic disorders.

overall_design: CD4+ T cells were purified from spleen single cell suspensions of C57BL/6J and db/db mice, total RNA was extracted from CD4+ T cells for RNA-sequencing.

first: Yujun

first: Ruihan

first: Hongrui

last: Liu

first: Tomoka

last: Hasegawa

first: Minqi

last: Li


title: Transcriptomic study of human brain organoids in Alzheimer's Disease

city: Groningen

first: Alejandro

last: Marmolejo

state: Groningen

address: Antonius Deusinglaan 2

country: Netherlands

institute: University of Groningen

summary: Alzheimer’s Disease (AD) is a neurodegenerative disorder characterized by the presence ofamyloid beta plaque deposition, tau hyperphosphorylation and decline in cognition. Mechanistically, these proteostatic changes are able to induce mitochondrial calcium overload that could lead to cell death. Conventional preclinical 2D in vitro AD models do not accurately recapitulate these hallmarks. As an effort to improve AD modeling, we have established iPSC-derived brain organoids (BO) harboring the PSEN1 Delta-E9 mutation and compared them to their CRISPR-Cas9-corrected isogenic controls. Organoids were also treated with an BACE-1 Inhibitor. As an effort to build into the complexity of organoid modeling, we co-cultured these organoids with their congenic iPSC-derived microglia-like cells.

overall_design: We have compared PSEN1 mutants and their isogenic controls. We have treated both groups with BACE-1 inhibitor, with Ferrostatin and co-cultured with their congenic microglia. We compared brain organoids with their corresponding iPSC cultures too.

last: Marmolejo-Garza

first: Amalia

middle: M

last: Dolga


title: Transcriptomic analysis of cellular senescence induced by ectopic expression of ATF6α in human breast cancer cells

city: Incheon

first: So-Hyun

last: Bae

state: Yeonsu-gu

address: 119, Academy-ro

department: Division of Life Sciences, College of Life Sciences and Bioengineering

institute: Incheon National University

summary:
To investigate the molecular mechanism that triggers endoplasmic reticulum (ER) stress-induced cellular senescence in breast cancer cells, we established MCF7 cell lines exposed to ATF6α ectopic expression.
We then performed gene expression profiling analysis using data obtained from RNA-seq of 9 different cells at three time points.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for  MCF-7 cells from three distinct time points (baseline, 6 hours, and 9 hours after ATF6α ectopic expression), using Illumina HiSeq X10 and NovaSeq 6000 platforms with paired-end 100 base sequencing.

first: Ju-Won

last: Kim

first: Yesol

last: Moon

first: Eun-Kyung

first: Youngjin

first: Yun-Gyu

last: Park

first: Mi-Ryung

last: Han

first: Jeongwon

last: Sohn


title: Epigenetic effects (H3K9ac, H3K14ac, H3K23ac and Pol II) of Kat6b loss in mouse neural stem cells (CUT&Tag)

city: Parkville

first: Waruni

last: Abeysekera

state: Victoria

address: 1G Royal Parade

department: Bioinformatics

institute: Walter and Eliza Hall Institute of Medical Research

summary: Heterozygous mutations in the histone acetyltransferase gene KAT6B (MYST4/MORF/QKF) cause cognitive disorders. Congruently, KAT6B is required for brain development, neural stem cell self-renewal and neuronal differentiation in mice. Despite the clear requirement for KAT6B in brain development, its molecular roles remain unexplored. Here we use CUT&Tag sequencing to determine the effects of loss or gain of KAT6B on H3K9ac and H3K23ac histone marks and on RNA Pol II.

overall_design:
Kat6b knock-out mice were constructed on the C57BL/6 background. Transgenic Tg(Kat6b) mice that overexpress Kat6b were also constructed and maintained on a FVBxBALB/c hybrid genetic background. The mixed genetic background was chosen because Tg(Kat6b) mice are not viable on the C57BL/6 background.
Neural stem and progenitor cells were isolated from the dorsal telencephalon of E12.5 mouse embryos. Samples were taken from Kat6b–/– knockout embryos and wild-type embryos on the C57BL/6 background, and from Tg(Kat6b) transgenic embryos and wild-type embryos on the FVB x BALB/c genetic background. All samples were also spiked with a fixed amount of DNA from Drosophila S2 cells to aid with normalization. CUT&Tag profiles were constructed for H3K9ac, H3K14ac, H3K23ac and RNA Pol II. Four biological replicates were undertaken for each genotype and CUT&Tag antibody.

first: Gordon

middle: K

last: Smyth

first: Anne

last: Voss

first: Tim

last: Thomas

first: Maria

last: Bergamasc


title: Transcriptomics Changes of Calcitonin Gene-related Peptide in Mitigating Lipopolysaccharide-induced Septic Cardiomyopathy

city: Fuzhou City

first: Shaodan

last: Feng

state: Fujian Province

address: Taijiang District

department: Emergency Department

institute: First Affiliated Hospital of Fujian Medical University

summary: Septic cardiomyopathy (SCM) is the predominant cause of death in sepsis patients with undefined mechanism to date. Our study analyzed RNA sequencing (RNA-seq) data from rat heart tissue to discover key targets and potential pharmacological actions of the calcitonin gene-related peptide (CGRP) against SCM. A lipopolysaccharide-induced SCM model was established in rats (LPS 10 mg/kg, intraperitoneal (i.p.)). Thereafter, the myocardial tissues from the three groups of rats (Ctrl group, LPS group, and CGRP group) (n=5) were extracted and underwent RNA-seq, followed by bioinformatics analyses. Our findings suggest a basis for finding potential targets for CGRP in the treatment of SCM.

overall_design: To explore the potential targets of CGRP for the treatment of septic cardiomyopathy, we established a rat model of septic cardiomyopathy. Then, we compared the gene expression profiling of RNA-seq data of rat myocardial tissues from different groups.

first: Kexin

last: Cai

first: Siming

last: Lin

first: Yuqing

last: Luo


title: Eosinophil Specialization Is Regulated by Exposure to the Esophageal Epithelial Microenvironment

city: Cincinnati

first: Marc

middle: E

last: Rothenberg

state: OH

address: 3333 Burnet Ave.

department: Allergy & Immunology

institute: Cincinnati Children's Hospital Medical Center

summary: Distinct subsets of eosinophils are reported in inflammatory and healthy tissues, yet the functions of uniquely specialized eosinophils and the signals that elicit them, particularly in eosinophilic esophagitis (EoE), are not well understood. Herein, we report an ex-vivo system wherein freshly isolated human eosinophils were cocultured with esophageal epithelial cells and disease-relevant pro-inflammatory (IL-13) or pro-fibrotic (TGF-β) cytokines.  Our results demonstrate that disease-relevant pro-inflammatory and pro-fibrotic signals present in the esophagus of EoE patients cause distinct profiles of eosinophil activation and gene expression.

overall_design: Immediately after isolation or following 96h of coculture with esophageal epithelial cells with- and without IL-13 or TGFb, eosinophils were lysed in TriPure and RNA was extracted with phenol-chloroform followed by Zymogen Quick RNA Microprep Kit. RNAseq libraries were generated as described in PMID 28041958 and 32362324.

first: Julia

middle: L

last: Dunn

first: Andrea

last: Szep

first: Emily

last: Gonzalez Galan

first: Simin

last: Zhang

first: Justin

last: Marlman

first: Julie

last: Caldwell

first: Ty

last: Troutman


title: RNA-sequencing analyses of cancer cell lines from KPPC and PPSSC pancreatic tumors

city: Houston

first: Yang

last: Chen

summary: RNA-sequencing analyses were performed on cancer cell lines from KPPC and PPSSC pancreatic tumors, so as to identify the changes of tumor immune microenvironment upon Trp53 and Smad4 deletion without KRAS mutation.

overall_design: We generated a novel transgenic mouse model (Trp53loxP/loxP;Smad4loxP/loxP;Pdx1-Cre) that harbors wild-type KRAS and loss of Trp53/Smad4 in the context of spontaneous PDAC.  Primary cancer cell lines established from pancreatic tumors were analyzed by RNA-seq.

first: Daowei

last: Yang

first: Xinlei

last: Sun


title: Myelination potential and injury susceptibility of grey versus white matter human oligodendrocytes

status: Public on Nov 08 2024

city: Montreal

first: Alyson

last: Fournier

state: QC

address: 3801 Rue University

department: Department of Neurology and Neurosurgery

institute: McGill University

summary: Increasing evidence indicates heterogeneity in functional and molecular properties of oligodendrocyte lineage cells both during development and under pathologic conditions. In multiple sclerosis, remyelination of grey matter lesions exceeds that in white matter. Here we used cells derived from grey matter versus white matter regions of surgically resected human brain tissue samples, to compare the capacities of human A2B5-positive progenitor cells and mature oligodendrocytes to ensheath synthetic nanofibers, and relate differences to the molecular profiles of these cells. For both cell types, the percentage of ensheathing cells was greater for grey matter versus white matter cells. For both grey matter and white matter samples, the percentage of cells ensheathing nanofibers was greater for A2B5-positive cells versus mature oligodendrocytes. Grey matter A2B5-positive cells were more susceptible than white matter A2B5-positive cells to injury induced by metabolic insults. Bulk RNA sequencing indicated that separation by cell type (A2B5-positive vs mature oligodendrocytes) is more significant than by region but segregation for each cell type by region is apparent. Molecular features of grey matter versus white matter derived A2B5-positive and mature oligodendrocytes were lower expression of mature oligodendrocyte genes and increased expression of early oligodendrocyte lineage genes. Genes and pathways with increased expression in grey matter derived cells with relevance for myelination included those related to responses to external environment, cell-cell communication, cell migration, and cell adhesion. Immune and cell death related genes were up-regulated in grey matter derived cells. We observed a significant number of up-regulated genes shared between the stress/injury and myelination processes, providing a basis for these features. In contrast to oligodendrocyte lineage cells, no functional or molecular heterogeneity was detected in microglia maintained in vitro, likely reflecting the plasticity of these cells ex vivo. The combined functional and molecular data indicate that grey matter human oligodendrocytes have increased intrinsic capacity to myelinate but also increased injury susceptibility, in part reflecting their being at a stage earlier in the oligodendrocyte lineage.

overall_design: RNA extracted from four distinct cell groups (GM A2B5-positive, WM A2B5-positive, GM 23 A2B5-negative and WM A2B5-negative cells) immediately after A2B5 selection. GM and WM 24 microglia RNA was extracted after 4 days in culture.

first: Abdulshakour

last: Mohammadnia


title: single cell-RNA sequencing for Immune cells from mouse brain tumor post anti PD-1 treatment

city: Daejeon

first: Keun Bon

last: Ku

address: 291 Daehak-ro

department: Biological Sciences

institute: Korea Advanced Institute of Science and Technology

summary: To evaluate the influence of ablating Fcgr2b on anti-PD-1 treatment efficacy, we administered GL261 cells to both Fcgr2b knockout and wild-type (WT) mice, followed by treatment with anti-PD-1. Afterwards, we isolated immune cells from the brains of these mice for further analysis.

overall_design: We analyzed immune cells infiltrating gliomas using single-cell RNA sequencing (scRNAseq) with 10X Genomics technology. For the administration of anti-PD-1 immune checkpoint inhibitor (ICI), wild-type (WT) and Fcgr2b knockout (KO) mice received intraperitoneal injections of 200 µg of anti-mouse PD-1 depletion antibody on days 10, 13, and 16 after inoculation with 1x10^5 GL261 cells (Day 0). Tumor-bearing brains were isolated 20 days after tumor cell inoculation, and sorted CD45-positive immune cells were subsequently analyzed.

first: Keun

middle: B

first: Chae-Won

last: Kim

first: Yumin

first: Byeong

last: Kang

first: Jeongwoo

last: La

first: Heung

last: Lee


title: Novel Mouse Model for Tissue-specific Extracellular Vesicle Isolation and Characterization

city: Rochester

first: Alexandra

last: Krivonos

state: MN

address: 200 1st St SW

department: Quantitative Health Sciences

institute: Mayo Clinic

summary: Extracellular vesicles (EVs) are key mediators of intercellular communication, with important roles in numerous physiological and pathological processes, including profound effects on bone metabolism. These small membrane-bound vesicles are produced and released in the extracellular environment by virtually all cell types, including cells in the osteogenic lineage such as bone marrow-derived mesenchymal stem cells (MSCs), osteoblasts, osteoclasts and osteocytes. EVs serve as potent carriers of bioactive molecules, such as nucleic acids, proteins, lipids and metabolites, where they can influence recipient cells through fusion with target cell membranes to deliver these functional biomolecules. However, once released, the source cell of the EV is difficult to ascertain with any certainty. To overcome this obstacle, we developed a conditional (e.g. Cre-mediated) mouse model that expresses an EV tag, containing a fusion of CD81 and multiple C-terminal tags, termed the “Snorkel-tag”. By crossing with a Cre of interest, representing a specific cell-type or tissue, the specific EV subpopulations that are released can be isolated using antibody affinity columns. We crossed the CAGS-Snorkel mouse with Prx1- and Ocn-Cre, representing cell-types in the early vs late stages of osteoblast differentiation, isolated EVs from bone marrow plasma, and treated mouse bone marrow stromal cells (mBMSCs) with Prx1-EVs, Ocn-EVs or All-EVs (isolated using a Pan EV Isolation Kit [Miltenyi Biotec]) for 3 days and performed bulk RNA-sequencing. We found unique transcriptional and pathway signatures elicited by the different EV subpopulations in the mBMSCs, suggesting that EVs from diverse sources have distinct biological activities.

overall_design: Mouse Models: We crossed the CAGS-Snorkel mouse with either the Prx1-Cre (n=5, 2 male/3 female) or Ocn-Cre (n=5, 2 male/3 female).To serve as a control EV sample, we also used wild-type (WT) mice (n=5, 2 male/3 female). For each group (Prx1-EVs, Ocn-Evs, All-EVs), bone marrow plasma was collected from the n=5 mice per group and pooled to generate a single bone marrow plasma sample for each group. All mice were in the C57BL/6N background and all mice were 6 months old at sacrifice. EV Isolation and RNA sequencing: EVs were isolated from the pooled bone marrow plasma from these mice using an HA-antibody magnetic-bead affinity column and mouse bone marrow stromal cells (mBMSCs) were treated with these EV subpopulations for 3 days. RNA was isolated from the EV-treated mBMSCs using the Rneasy Plus Kit (Qiagen) and bulk mRNA-sequencing was performed.

first: David

last: Monroe

first: Chantal

last: McCabe

first: Bryan

last: Piatkowski

first: Sundeep

last: Khosla


title: Maintenance and functional regulation of immune memory to COVID-19 vaccines in tissues

city: New York

first: Alex

last: George

state: NY

address: 960 Broadway, 2nd Floor, Room 203AC

institute: Columbia University

summary: Memory T and B cells in lymphoid and mucosal tissues maintain long-term protection, though their generation following vaccination remains challenging to assess in humans. Here, we investigated immune memory generated to COVID-19 mRNA vaccines across blood, lymphoid organs, and lungs from 42 vaccinated organ donors aged 23-86, of whom 57% were previously infected with SARS-CoV-2. Using high-dimensional profiling, we reveal that Spike (S)-reactive memory T cells distribute in lymphoid organs and lungs, variably express tissue resident markers based on infection history, and exhibit site-specific compositions of effector and regulatory memory T cells. S-reactive B cells are mostly class-switched memory cells localized to lymphoid organs correlating with circulating antibodies. Importantly, tissue memory T cells are more stably maintained post-vaccination and over age and exhibit a bias towards regulatory cell functional profiles compared to circulating populations. Our results show that mRNA vaccines can induce heterogeneous memory populations across sites for protection and controlling immune pathology.

overall_design: We performed CITE-seq on immune cell populations from human blood and different human organ donor tissues in response to SARS-COV2 and influenza peptide pool stimulation and DMSO controls.

first: Julia

last: Davis-Porada

first: Peter

last: Sims

first: Donna

last: Farber


title: TNFa has Differential Effects on the Transcriptome Profile of Selected Populations in Murine Cartilage

city: Farmington

first: Ernesto

last: Canalis

state: CT

address: 263 Farmington Avenue

department: Orthopaedic Surgery

institute: UConn Health

summary: To further our understanding of the role of tumor necrosis factor (TNFα) on the inflammatory response in cartilage, we explored its effects on the transcriptome profile of murine epiphyseal chondrocytes at the total and single cell (sc) resolution. Gene set enrichment analysis of total RNA-Seq from TNFα-treated chondrocytes revealed enhanced response to biotic stimulus, defense and immune response and cytokine mediated signaling and suppressed cartilage and skeletal morphogenesis and development. scRNA-Seq analyzed 14,239 cells and 24,320 genes and distinguished 16 cell clusters. The more prevalent ones were constituted by cells of the limb bud, chondrogenic cells and fibroblasts comprising ~73% of the cell population.  Genes expressed by joint fibroblasts were detected in 5 clusters comprising ~45% of the cells isolated.  Pseudotime trajectory finding revealed an association between fibroblast and chondrogenic clusters which was not modified by TNFα. TNFα decreased the total cells recovered by 18.5% and the chondrogenic, limb bud and mesenchymal clusters by 32%, 27% and 7%, respectively.  TNFα had profound effects on the insulin-like growth factor (IGF) axis decreasing Igf1, Igf2 and Igfbp4 and inducing Igfbp3 and Igfbp5, explaining an inhibition of collagen biosynthesis, cartilage and skeletal morphogenesis. Ingenuity Pathway Analysis of scRNA-Seq data revealed that TNFα enhanced the osteoarthritis, rheumatoid arthritis, pathogen induced cytokine storm and interleukin 6 signaling pathways and suppressed fibroblast growth factor signaling. In conclusion, epiphyseal chondrocytes are constituted by diverse cell populations distinctly regulated by TNFα to promote inflammation and suppression of matrix biosynthesis and growth.

overall_design: Chondrocyte-enriched cells were isolated from the epiphyses of long bones of the hind and fore limbs from 3- to 4-day-old C57BL/6 mice.b Cultured in the presence and absence of TNFa.

first: Canalis

last: Ernesto

first: Schilling

last: Lauren

first: Denker

last: Emily


title: Discovery and Anti-inflammatory Activity of a Cyanobacterial Macrolide Glycoside Targeting the Keap1/Nrf2 Pathway

city: GAINESVILLE

first: FAHONG

last: YU

state: FL

address: 2033 Morway RD

department: ICBR

institute: UNIVERSITY OF FLORIDA

summary: We discovered a cyanobacterial macrolide glycoside that contains an α,β-unsaturated carbonyl system a key motif required for the activation of the Keap1/Nrf2−ARE pathway that is involved in the activation of antioxidant and phase II detoxification enzymes. The compound was screened in ARE luciferase reporter gene assay which induced Nrf2 activity at 32 and 10µM. Given the established crosstalk between oxidative stress and inflammation, we aimed at investigating the antiinflammatory potential of our compound using RAW264.7 mouse macrophage. We carried RNAseq to capture the effects of the compound on transcript levels and identify additional pathways involved in inflammation that could be modulated. We further evaluated the cytotoxicity of the compound against HCT 116 and carried out RNAseq at the same cell line.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: RAW264.7 mouse macrophage cells were treated with two concentrations (32, 10 µM) of the macrolide glycoside in addition to solvent control (DMSO) for 1h followed by stimulation with Lipopolysaccharide (LPS). The RNA was collected after 12h. RNA from cells treated with DMSO without LPS stimulation were also obtained. For the HCT116, the same experimental design was carried out but without LPS stimulation.

first: Fatma

last: Al-Awadhi

first: Ranjala

last: Ratnayake

first: Hendrik

last: Luesch


title: Characterization of cuprizone mouse model at single-cell and spatial transcriptomics level

city: Cambridge

first: Baohong

last: Zhang

state: MA

address: 225 Binney St

institute: Biogen Inc.


title: TNFa has Differential Effects on the Transcriptome Profile of Selected Populations in Murine Cartilage [scRNA-seq]


title: Genome wide map of Runx3, Brn3a binding and Histone H3 acetylated on lysine 27 (H3K27Ac) during early development of DRG TrkC neurons

city: Rehovot

first: Tsviya

last: Olender

address: 234 Herzl St.

department: Molecular Genetics

country: Israel

institute: The Weizmann Institute

summary: We isolated E12.5 TrkC neurons and profiled Runx3, Brn3a binding and Histone H3 acetylated on lysine 27 (H3K27Ac) during early development of DRG TrkC neurons by Cut&Run

overall_design: Isolation of GFP expressing TrkC neurons from E12.5 BacE-GFP embryos and performing Cut&Run with anti-Runx3, anti-Brn3a and anti-H3K27Ac antibodies.

first: Elena

last: Appel

first: Kira

last: Orlovsky

first: Shay

last: Hantisteanu

first: Joseph

last: Lotem

first: Ditsa

last: Levanon

first: Yoram

last: Groner


title: Characterization of cuprizone mouse model at single-cell and spatial transcriptomics level[Spatial Transcriptomics]

summary: We generated a comprehensive dataset utilizing the cuprizone model. This dataset encompasses bulk RNA-seq, single-nucleus RNA-seq (snRNA-seq), and spatial transcriptomics, with the aim of investigating the molecular changes linked to the cuprizone-induced demyelination phenotype. Integration of these omics dataset allowed us to investigate the changes occurring at both the single-cell and spatial levels, thereby enabling a deeper understanding of the cellular dynamics and molecular interactions associated with the cuprizone-induced demyelination phenotype.

overall_design: In the cuprizone (CPZ) group, male mice aged 6-8 weeks were administered a 0.3% Cuprizone diet for 4 weeks to induce demyelination in the brain (n = 3). In the recovery group, mice were administered CPZ diet for 4-weeks and then switched to a control diet for an additional 2 weeks (n = 3). In the control group, mice received the control diet throughout the entire 6-week study period (n = 4). At 4 (CPZ) and 6 weeks (Control and Recovery), mice were euthanized by carbon dioxide (CO2) inhalation, and their brains were subsequently dissected. Each brain was cut sagittally to separate left and right hemispheres, one brain hemisphere was trimmed into three coronal segments and embedded in OCT for snRNA-seq or spatial transcriptomic study, the other hemisphere was snap frozen for bulk RNA-seq study (Figure 1). To enrich for sequencing of cells from the corpus collosum and cortex, only the top half of the coronal section was collected for snRNA-seq, which includes the dorsal cortex, CC, and hippocampus.

first: Hui-Hsin

last: Tsai

first: Sarbottam

last: Piya

first: Jing

last: Zhu


title: Characterization of cuprizone mouse model at single-cell and spatial transcriptomics level[bulk RNA-seq]


title: Sympathetic Nerve Signal is Critical for Regulating Mammary Gland Development and Mammary Stem Cell Activity

city: wuhan

first: ye

last: zi

address: donghu street

institute: wuhan university

summary: Adult stem cell activity and organ development are elaborately regulated by microenvironmental signals. However, little is known about the regulatory effects of microenvironmental sympathetic nerve signals on organ development and stem cell activity. Here, using a mouse mammary gland model, we determined the regulatory function of sympathetic nerve system (SNS) on mammary development and mammary stem cell activity. Our results indicated that depletion of sympathetic nerve signals delayed the elongation of mammary ducts during puberty and pregnancy, and lead to the loss of mammary stem cells (MaSCs). In vitro three-dimensional (3D) culture and in vivo transplantation analyses indicated that the loss of sympathetic nerve signals inhibits the self-renewal and reconstruction activity of mammary stem cells, while the activation of sympathetic nerve signals promotes the self-renewal and reconstruction activity of mammary stem cells. Mechanistically, we found that sympathetic nerve signals regulate the activity of mammary stem cells and mammary development through the PI3K/ERK pathway. Together, our study reveals the function of sympathetic nervous signals in maintaining mammary homeostasis and regulating mammary stem cell activity, providing a novel view for the nervous system's regulation of organ development.

overall_design: To explore the role of the sympathetic nervous system in regulating mammary gland development and mammary stem cells, we collected primary basal cells from 6OHDA-treated mice as well as basal clones treated with adrenergic receptor agonists in 3D Matri-gel culture. Then the four groups of cells were subjected to RNA sequencing and differential expression analysis.

first: Zi

last: Ye

first: Cheguo

last: Cai


title: Next Generation Sequencing Facilitates Quantitative Analysis of Bone Marrow-Derived Macrophages Transcriptomes for Acly-E14 siRNA effects

city: Shanghai

first: Xingguang

last: Liu

address: Xiangyin road 800

department: National Key Laboratory of Immunity & Inflammation

institute: Naval medical university

summary: Purpose: The purpose of this study is to detect activated or silenced genes during  bone marrow derived macrophages (BMDMs) transfected with control siRNA or Acly-E14 siRNA. Gene expression differences between two samples could be found using transcriptome profiling (RNA-seq) analysis. Methods: Mouse BMDMs were generated from bone marrow cells in RPMI-1640 medium with recombinant mouse M-CSF (20ng/ml). BMDMs were stained to confirm the surface expression of CD11b and F4/80. Cells with purity >97.5% were used for subsequent experiments. BMDMs were transfected with control siRNA or Acly-E14 siRNA. 48 hour later, they were stimulated with LPS (100ng/ml) for 4 hours, of which RNA profiles were generated by deep sequencing, using Illumina. Results: We mapped about 10 million sequence reads per sample to the mouse genome, identified hundreds of genes with significant mRNA variation between BMDMs transfected with the indicated siRNAs.

overall_design: mRNA profiles of BMDMs transfected with control siRNA or Acly-E14 siRNA were generated by deep sequencing.

first: Yunkai