title: Transcriptional analysis of blood-borne immune cells induced by VSV-based vaccines: A comparison before and after immunization

city: Shanghai

first: Fan

last: Zhao

address: 160 Baseng Road, Pudong New District

institute: R&R Therapeutic  Co., Ltd.

summary: Viral vectors are renowned for their inherent adjuvant effects. We here developed cancer vaccines based on the Vesicular Stomatitis Virus (VSV) vector using two different construction strategies.The first vaccine involved the traditional method of inserting tumor antigen genes into the VSV genome (SP vaccine). The second strategy, built upon the first, additionally displayed tumor antigens on the surface of the VSV, creating particulate antigen vaccine (CAD vaccine). To compare the early immune responses induced by these two vaccine strategies, we collected blood samples for RNA sequencing analysis at 24 hours post-immunization with both vaccines and a viral vector control group. The results indicated comparable early biological processes, including antiviral and innate immune responses, among the three groups. This confirms that the vector predominantly drives the early innate immune response.

overall_design: To compare the early immune responses induced by these  VSV-based vaccines ( SP vaccine, and CAD vaccine) and  vector control, BALB/c mice were immunized with the same dosage, and blood samples were collected for RNA sequencing analysis at 24 hours before and after immunization.

first: Chunfeng

last: Liu

first: Yilin

last: Xia


title: An Intrinsic Disorder Region Controlling Condensation of a Circadian Clock Component and Rhythmic Transcription in the Liver [RNA-seq]

status: Public on Nov 05 2024

city: Philadelphia

first: KUN

last: ZHU

state: PA

address: 3400 Civic Center Blvd, SCTR-12-196

country: USA

institute: University of Pennsylvania

type: Expression profiling by high throughput sequencing

summary: Circadian gene transcription is fundamental to metabolic physiology. Here we report that the nuclear receptor REV-ERBa, a repressive component of the molecular clock, forms circadian condensates in the nuclei of mouse liver. These condensates are dictated by an intrinsically disordered region (IDR) located in the protein’s hinge region which specifically concentrates nuclear receptor corepressor 1 (NCOR1) at the genome. IDR deletion diminishes the recruitment of NCOR1 and disrupts rhythmic gene transcription in vivo. REV-ERBa condensates are located at high-order transcriptional repressive hubs in the liver genome that are highly correlated with circadian gene repression. Deletion of the IDR disrupts transcriptional repressive hubs and diminishes silencing of target genes by REV-ERBa. This work demonstrates physiological circadian protein condensates containing REV-ERBa whose IDR is required for hub formation and the control of rhythmic gene expression.

overall_design: Comaprative gene expression profiling analysis of RNA-seq for Ctrl, REV-ERBs double-knockout and REV-ERBalpha re-expressing  mouse livers

first: Kun

last: Zhu

first: Mitchell

middle: A

last: Lazar


title: Widespread destabilization of C. elegans microRNAs by the E3 ubiquitin ligase EBAX-1 [miRNA-Seq]

status: Public on Nov 04 2024

city: Cambridge

first: Michael

middle: W

last: Stubna

state: MA

address: 455 Main St

institute: Whitehead Institute

type: Non-coding RNA profiling by high throughput sequencing

summary: MicroRNAs (miRNAs) associate with Argonaute (AGO) proteins to form a complex that directs mRNA repression. One way that miRNAs are regulated is at the level of their stability. Some animal miRNAs are destabilized through a pathway in which pairing to specialized RNA transcripts recruits the ZSWIM8 E3 ubiquitin ligase, which polyubiquitinates AGO, leading to its degradation and exposure of the miRNA to cellular nucleases. We found that 22 miRNAs in C. elegans are sensitive to loss of EBAX-1, the ZSWIM8 ortholog in nematodes, implying that these 22 miRNAs might be subject to this pathway of target-directed miRNA degradation (TDMD). These miRNAs included the embryonic miR-35–42 family, as well as other miRNAs among the least stable in the worm, suggesting that TDMD is a major pathway of miRNA destabilization in the worm. The excess miR-35–42 molecules that accumulated in ebax-1 mutants caused increased repression of their predicted target mRNAs and underwent 3′ trimming over time. In general, however, miRNAs sensitive to loss of EBAX-1 did not have a consistent pattern of either trimming or tailing. A previous study reports that in contrast to TDMD substrates in mammals and flies, the 3′ region of miR-35 is not required for its destabilization, raising the question of whether the 3′ regions of other miRNAs are dispensable for EBAX-1 sensitivity in nematodes. To address this question, we replaced the 3′ region of another EBAX-sensitive miRNA, miR-43, and found that this replacement greatly reduced EBAX-sensitivity in the worm. Together, these findings broaden the implied biological scope of TDMD-like regulation of miRNA stability in animals, and indicate that the requirement for miRNA 3′ sequences in this process is variable in the worm.

overall_design: To profile microRNAs destabilized by EBAX-1, we sequenced small RNAs from synchronized wild-type and ebax-1 mutant worms at eight different developmental time points. For mutagenesis studies of miR-43, we sequenced small RNAs from L4 larvae with or without miR-43 mutations and/or ebax-1 mutations in triplicate.

first: Aditi

last: Shukla

first: David

middle: P

last: Bartel


title: SOXC factors as suppressors modulate Rho activity to promote neuronal reprogramming of human glioblastoma cells

city: DALLAS

first: jianjing

last: Yang

state: TX

address: 6000 Harry Hines Blvd

department: Molecular Biology

institute: UT Southwestern Medical Center

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Genomic targeted genes of NGN2 in U251 cell line

overall_design: ChIP-seq for NEUROG2 (4 input datasets, 8 immunoprecipitation datasets) in U-251 Glioblastoma cells at 1,3 or 5 days post treatment of chemicals (Forskolin and Dorsomorphin) (DPT).

first: Derek

middle: K

last: Smith

first: Jianjing

first: Chun-Li

last: Zhang


title: Gene expression analysis in mesothelioma cell lines with and without K-975 treatment

city: Nagoya

first: Tatsuhiro

last: Sato

state: Aichi

address: 1-1 Kanokoden

department: Division of Cancer Biology

country: Japan

institute: Aichi Cancer Center Research Institute

summary: RNA sequencing was performed to analyze the genes whose expression was suppressed by K-975 treatment.

overall_design: Three biological replicates were analyzed for each of the 12 conditions (6 cell lines in the presence or absence of K-975).

first: Akao

last: Ken

first: Sato

last: Tatsuhiro

first: Sekido

last: Yoshitaka


title: Homology mediated end joining enables efficient non-viral targeted integration of large DNA templates in primary human T cells

city: Minneapolis

first: Lauren

last: Mills

state: MN

address: 420 Delaware St SE

department: Department of Pediatrics

institute: University of Minnesota

summary: Adoptive cellular therapy using genetically engineered immune cells holds tremendous promise for the treatment of advanced cancers. While the number of available receptors targeting tumor specific antigens continues to grow, the current reliance on viral vectors for clinical production of engineered immune cells remains a substantial bottleneck limiting translation of promising new therapies. Here, we describe an optimized methodology for efficient CRISPR-Cas9 based, non-viral engineering of primary human T cells that overcomes key limitations of previous approaches. By synergizing temporal optimization of reagent delivery, reagent composition, and integration mechanism, we achieve targeted integration of large DNA cargo at efficiencies nearing those of viral vector platforms with minimal toxicity. CAR-T cells generated using our approach are highly functional and elicit potent anti-tumor cytotoxicity in vitro and in vivo. Importantly, our method is readily adaptable to current Good Manufacturing Practices (cGMP) and clinical scale-up, offering a near-term alternative to the use of viral vectors for production of genetically engineered T cells for cancer immunotherapy.

overall_design: Peripheral Blood T cells from 3 donors were isolated, and cultured overnight.  Cells were then stimulated with anti-CD3/anti-CD28 Dynabeads for 24, 36, 48, 72, and 96 hours.  A unstimulated control was cultured for 24 hours before harvest.  RNA was isolated from samples and used for RNAseq

first: Matthew

last: Johnson

first: Tim

last: Starr

first: Jason

last: Bell

first: Xiaohong

last: Qiu

first: Walker

last: Lahr

first: Beau

last: Webber


title: Whole transcriptome sequencing using  Huh7 cells transfected with Dengue virus serotype  4 NS1 expressing plasmid.

city: Kolkata

first: UPASANA

last: RAY

state: West Bengal

address: CN-06, CN Block, Sector 5, Bidhannagar

department: Infectious Diseases and Immunology

country: India

institute: CSIR-Indian Institute of Chemical Biology

summary: To study the role of Dengue virus serotype 4  NS1 in modulation of host cell transcriptome we performed gene expression profiling using data obtained from  RNA- sequencing of  total RNA from 3  different samples: Huh7 cells (no transfection), Huh7 cells transfected with pTracer-SV40  empty vector and Huh7 cells transfected with pTracer-SV40 encoding dengue virus serotype 4 NS1  (pDENV4 NS1).

overall_design: After 24 hours of transfection, cells were lysed and total RNA was purified. RNA samples were proccessed for whole transcriptome sequencing using   Illumina Platform, followed by comparative gene expression analysis.

first: Upasana

last: Ray

first: Sandeepan

last: Das

first: Dluya

middle: S

last: Thagriki


title: Widespread destabilization of C. elegans microRNAs by the E3 ubiquitin ligase EBAX-1 [RNA-Seq]

first: Michael

last: Stubna

overall_design: To identify mRNAs regulated by EBAX-1, we sequenced poly(A)-selected RNA from synchronized wild-type and ebax-1 mutant L1 larvae in duplicate. To assay the abundances of protein-coding transcripts near piRNAs dysregulated in ebax-1 mutant adults, we sequenced poly(A)-selected RNA from wild-type and ebax-1 mutant adults in singlicate.

first: Aditi

last: Shukla

first: David

last: Bartel


title: A study of the effects of bisphenol A on dog obesity

city: Seongnam-si

first: Eun-Jung

last: Park

state: Gyeonggi-do

address: 1342 Seongnam-daero

department: Food and nutrition

country: South Korea

institute: Gachon University

summary: The transcriptomic changes caused by BPA on canine adipose-derived stem cells (cADSCs) during differentiation were investigated in this study. The transcriptomic profiles were investigated using RNA-sequencing.

overall_design: For the chemical treatment, 1 × 10^5 cADSC cells/well were seeded into six-well plates in DMEM culture medium containing 10% FBS and 1% antibiotic-antimycotic. After 2 days, cADSCs were induced with differentiation media (1 μM dexamethasone, 200 nM rosiglitazone, and 10 μg/mL insulin) with or without BPA (1 nM or 1 μM for in vitro assays whereas 1 μM for RNA-sequencing experiment ). The differentiation was induced for 14 days.

first: Hae-Jeung

last: Lee

first: Jong-Yeon

last: Kim

first: Sanghoon


title: Somatic polyploidy supports biosynthesis and tissue function by increasing transcriptional output [RNA-seq]

status: Public on Nov 03 2024

city: Stanford

first: Alexander

last: Lessenger

state: CA

address: 371 Jane Stanford Way

department: Biology

institute: Stanford University

summary: Cell size and biosynthetic capacity generally increase with increased DNA content. Somatic polyploidy has therefore been proposed to be an adaptive strategy to increase cell size in specialized tissues with high biosynthetic demands. However, if and how DNA concentration limits cellular biosynthesis in vivo is not well understood. Here, we show that polyploidy in the C. elegans intestine is critical for cell growth and yolk biosynthesis, a central role of this organ. Artificially lowering the DNA/cytoplasm ratio by reducing polyploidization in the intestine gave rise to smaller cells with dilute mRNA. Highly-expressed transcripts were more sensitive to this mRNA dilution, whereas lowly-expressed genes were partially compensated – in part by loading more RNA Polymerase II on the remaining genomes. Polyploidy-deficient animals produced fewer and slower growing offspring, consistent with reduced synthesis of highly-expressed yolk proteins. DNA-dilute cells had normal total protein concentration, which we propose is achieved by increasing expression of translational machinery at the expense of specialized, cell-type specific proteins.

sample_organism:
Caenorhabditis elegans
Pristionchus pacificus

overall_design: RNA-sequencing of dissected intestines from C. elegans with normal or reduced intestine ploidy and dissected intestines of Pristionchus pacificus used as a biological spike-in.

middle: T

first: Jan

middle: M

last: Skotheim

first: Matthew

last: Swaffer

first: Jessica

middle: L

last: Feldman


title: CEACAM1-engineered MSCs have a broad spectrum of immunomodulatory functions and therapeutic potential via cell-to-cell interaction

city: Seoul

first: Seong-Who

address: 88, Olympic-ro 43-gil

institute: University of Ulsan

summary: Mesenchymal stem cells (MSCs) have garnered attention for their regenerative and immunomodulatory capabilities in clinical trials for various diseases. However, the effectiveness of MSC-based therapies, especially for conditions like graft-versus-host disease (GvHD), remains uncertain. The cytokine IFN-γ has been known to enhance the immunosuppressive properties of MSCs through cell-to-cell interactions and soluble factors. In this study, we observed that IFN-γ-treated MSCs upregulated the expression of carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1), associated with immune evasion through the inhibition of NK cell cytotoxicity. To co-opt this immunomodulatory function, we generated MSCs overexpressing CEACAM1 and found that CEACAM1-engineered MSCs significantly reduced NK cell activation and cytotoxicity, independent of NKG2D ligand regulation. Furthermore, CEACAM1-engineered MSCs effectively inhibited the proliferation and activation of T cells along with the inflammatory responses of monocytes. In a humanized GvHD mouse model, CEACAM1-MSCs, particularly CEACAM1-4S-MSCs, demonstrated therapeutic potential by improving survival and alleviating symptoms. These findings suggest that CEACAM1 expression on MSCs contributes to MSC-mediated regulation of immune responses and that CEACAM1-engineered MSC could have therapeutic potential in conditions involving immune dysregulation.

overall_design:
To examine gene expression changes of human umbilical cord mesenchymal stem cells induced from addition of Interferon‐gamma (IFN‐γ), we treated 25 ng/ml of IFN‐γ for 7 hours or 48 hours then performed RNA-seq analysis with harvested RNA.

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Due to hospital policy regarding donor's privacy, raw data cannot be deposited
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first: Seongwho


title: Somatic polyploidy supports biosynthesis and tissue function by increasing transcriptional output [ChIP-seq]

overall_design: Chromatin immunoprecipitation and DNA sequencing (ChIP-seq) for RNA Polymerase II )Pol II) in C. elegans intestines. Worms expressing endogenous Pol II::GFP and intestine specific V5-GFP-nanobody were subjected to ChIP with an anti-V5 antibody.


title: Foxi1 regulates multiple steps of mucociliary development and ionocyte specification through transcriptional and epigenetic mechanisms

city: Freiburg

first: Peter

last: Walentek

state: BW

address: Breisacherstr. 113

department: Internal Medicine IV & IMITATE

country: Germany

institute: University Freiburg Medical Center

summary: Foxi1 is a master regulator of ionocytes (ISCs) across species and organs. Two subtypes of ISCs exist, and both a- and b-ISCs regulate pH- and ion-homeostasis in epithelia. Gain and loss of Foxi1 function are associated with human diseases, including Pendred syndrome, male infertility, kidney malfunction and cancers. Foxi1 functions were predominantly studied in the context of ISC specification, however, reports indicate additional functions in early and ectodermal development. Here, we re-investigated the functions of Foxi1 to Xenopus laevis embryonic mucociliary epidermis development and found a novel function for Foxi1 in the generation of Notch-ligand expressing mucociliary multipotent progenitors (MPPs). We demonstrate that Foxi1 has multiple concentration-dependent functions: At low levels, Foxi1 confers ectodermal competence through transcriptional and epigenetic mechanisms, while at high levels, Foxi1 induces a multi-step process of ISC specification and differentiation. We further describe how foxi1 expression is affected through auto- and Notch-regulation, how Ubp1 and Dmrt2 regulate ISC subtype differentiation, and how this developmental program affects Notch signaling as well as mucociliary patterning. Together, we reveal novel functions for Foxi1 in Xenopus mucociliary epidermis formation, relevant to our understanding of vertebrate development and human disease.

overall_design: ATAC-seq of stage 10 Xenopus laevis mucociliary organoids from uninjected or foxi1 morpholino-injected embryos

first: Sarah

last: Bowden


title: The effect of lnc-ORA knockout on muscle fiber type transformation in mouse leg muscle.

city: 咸阳市

first: HE

last: Yu

state: 陕西省

address: 陕西省 / 咸阳市 / 杨陵区 / 杨陵街道

country: China

institute: 西北农林科技大学

summary: Skeletal muscle fiber type conversion is usually regulated by coding and non-coding RNA, which is closely related to obesity, Duchenne muscular dystrophy and sarcopenia.  Furthermore, lnc-ORA KO mice were first constructed, and the results showed that lnc-ORA deficiency significantly increased exercise endurance, oxidative myofibers and mitochondrial function. Meanwhile, combining transcriptomic and proteomic data showed that lnc-ORA deficiency improved the biological processes and pathways related to the transformation of skeletal muscle fiber types. Mechanically, MYH7 is a target of lnc-ORA by multi-omics and RNA pull-down data, and it was degraded by interacting with ARIH1 through ubiquitination pathway. Importantly, lnc-ORA overexpression decreased the expression of MYH7 and enhanced the interaction of MYH7 and ARIH1, while lnc-ORA knockdown was the opposite. Overall, integrating transcriptomic and proteomic reveals the positive role of lnc-ORA deficiency on slow myofiber type formation via upregulating the level of MYH7 and inhibiting interaction between MYH7 and ARIH1. Our findings provide novel insights into the regulation of lncRNA on skeletal muscle fiber types.

overall_design: To investigate the role of lnc-ORA in skeletal muscle fiber type transformation, We generated lnc-ORA knockout (KO) mice using the CRISPR genome‐editing system with the C57BL/6 background.Subsequently, we performed RNA-seq on the gastrocnemius muscle of 15-week-old lnc-ORA knockout mice and conducted gene expression analysis.

first: Ziyi

last: Zhang

first: He


title: Genome wide map of Runx3, Brn3a, Islet1 binding, chromatin accessibility (ATAC) 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 E11.5 TrkC neurons and profiled chromatin accessibility (ATAC), Runx3, Brn3a, Islet1 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 E11.5 BacE-GFP embryos and performing Cut&Run with anti-Runx3, anti-Brn3a, anti-Islet1 and anti-H3K27Ac antibodies. Performing ATAC assay

first: Kira

last: Orlovsky

first: Elena

last: Appel

first: Shay

last: Hantisteanu

first: Joseph

last: Lotem

first: Ditsa

last: Levanon

first: Yoram

last: Groner


title: Induction of transposable element expression is central to innate sensing

city: Paris

first: Sebastian

last: Amigorena

address: 26 rue d'Ulm

country: France

institute: Institut Curie

summary: Detection of PAMPs induces transposable element expression that is subsequently detected by additional innate sensors to broaden and strengthen host defense.

overall_design: Three biological replicates were analyzed for each condition.

first: Derek

middle: C

last: Rookhuizen

first: Pierre-Emmanuel

last: Bonté


title: METTL4-mediated mitochondrial DNA N6-methyldeoxyadenosine promoting macrophage inflammation and atherosclerosis

status: Public on Nov 02 2024

city: Nanjing

first: Hong

last: Jiang

address: Longmian Road No.101

institute: Nanjing Medical University

summary: BACKGROUND: Mitochondrial dysfunction is a key factor in the development of atherogenesis. Methyltransferase-like protein 4 (METTL4) mediates N6-deoxyadenosine (6mA) methylation of mammalian mitochondrial DNA (mtDNA). However, the role of METTL4-mediated mitoepigenetic regulation in atherosclerosis is still unknown. This study aims to investigate the potential involvement of METTL4 in atherosclerosis, explore the underlying mechanism, and develop targeted strategies for treating atherosclerosis. METHODS: Expression levels of mtDNA 6mA and METTL4 were determined in atherosclerotic lesions. We explored the mechanism of METTL4 involvement in atherosclerosis using Mettl4Mac-KO-Apoe-/- and Mettl4MUT-Apoe-/- mice and cell models, as well as bone marrow transplantation. Natural compound libraries were screened to identify potent METTL4 antagonists. In addition, bioinspired proteolysis-targeting (PROTAC) technology targeting macrophages within plaques was used to increase the efficacy of the METTL4 antagonist. RESULTS: The expression levels of mtDNA 6mA and METTL4 were significantly increased in plaque macrophages. Mettl4Mac-KO-Apoe-/- mice displayed suppressed mtDNA 6mA levels and atherosclerotic progression, which were reversed by METTL4 restoration via bone marrow transplantation (n = 6). Mechanistically, elevated METTL4 expression reduces MT-ATP6 expression by suppressing its transcription, thereby impairing the activity of mitochondrial respiration chain complex V. This disruption leads to the accumulation of excess protons in the mitochondrial intermembrane space, causing mitochondrial dysfunction. Consequently, mtDNA is released into the cytoplasm, ultimately triggering inflammasome activation. All results were reversed by the mutation in the METTL4 methyltransferase active site. Mettl4MUT-Apoe-/- mice showed suppressed mtDNA 6mA levels and atherosclerotic progression, and repaired mitochondrial function of macrophage, which were reversed by METTL4 restoration via bone marrow transplantation (n = 6). Pemetrexed (PEM) was identified as the first METTL4 antagonist to effectively alleviate atherosclerotic progression. Furthermore, we generated a proteolysis-targeting chimera drug based on PEM (PROTAC-PEM) that specifically targeting METTL4 in macrophages within plaques, showing a promising therapeutic effect on atherosclerosis. CONCLUSIONS: This study revealed a novel mechanism by which mtDNA 6mA orchestrated mitochondrial function-related gene expression in macrophages, thereby promoting atherosclerosis. Through various experimental techniques, such as gene manipulation, pharmacological inhibition and PROTAC, this study demonstrated that mtDNA 6mA and its specific enzyme METTL4 hold potential as therapeutic targets for atherosclerosis.

overall_design: ox-LDL-stimulated HMDMs transfected with or without siMETTL4, three samples for each group.


title: Effect of macrophage SNO-Septin2 in aortic aneurysm development

first: Hao

address: Longmian Street

summary: We analyzed differentially expressed genes in aortas from Apoe-/- mice that received Apoe-/- bone marrow or Apoe-/-Sept2C111A bone marrow and were infused with Ang II for 28 days to explore the mechanisms of macrophage SNO-Septin2 in aortic aneurysm development.

overall_design: Identification of differentially expressed genes in aortas from Apoe-/- mice that received Apoe-/- bone marrow or Apoe-/-Sept2C111A bone marrow and were infused with Ang II for 28 days (n=3 per group).

first: Yan

first: Shuang

last: zhao

first: Yong

last: Ji


title: Effect of SNO-Septin2 in macrophage inflammation and migration

summary: We analyzed differentially expressed genes in bone marrow drived macrophages (BMDMs) from wild-type or Sept2C111A mice treated with 30ng/ml TNF-α for 24 hours to explore the mechanism that SNO-septin2 regulates inflammation and migration of macrophages.

overall_design: Identification of differentially expressed genes in BMDMs from wild-type or Sept2C111A mice treated with 30ng/ml TNF-α for 24 hours (n=4 per group).


title: Prospective pharmacotyping of urothelial carcinoma organoids for drug sensitivity prediction – feasibility and real world experience

city: Ulm

first: Michael

middle: Karl

last: Melzer

address: Albert-Einstein-Allee 23

institute: Ulm University Hospital

summary: Urothelial carcinoma (UC) of the urinary bladder has significant challenges in treatment due to its diverse genetic landscape and variable response to systemic therapy. In recent years, patient-derived organoids (PDOs) emerged as a novel tool to model primary tumors with higher resemblance than conventional 2D cell culture approaches. However, the potential of organoids to predict therapy response in a clinical setting remains to be evaluated. This study explores the clinical feasibility of PDOs for pharmacotyping in UC. Initially, we subjected tumor tissue specimens from 50 patients undergoing transurethral resection or radical cystectomy to organoid propagation, of whom 19 (38%) yielded PDOs suitable for drug sensitivity assessment. Notably, whole transcriptome-based analysis indicated that PDOs may show phenotypes distinct from their parental tumor tissue. Pharmacotyping within a clinically relevant timeframe [mean of 35.44 and 55 days for non-muscle invasive bladder cancer (NMIBC) and muscle invasive bladder cancer (MIBC), respectively] was achieved. Drug sensitivity analyses revealed marked differences between NMIBC and MIBC, with MIBC-derived organoids demonstrating higher chemosensitivity toward clinically relevant drugs. A case study correlating organoid response with patient treatment outcome illustrated the complexity of predicting chemotherapy efficacy, especially considering the rapid acquisition of drug resistance. We propose a workflow of prospective organoid-based pharmacotyping in UC, enabling further translational research and integration of this approach into clinical practice.

overall_design: Whole transcriptomes from 8 organoid lines (total RNA) and their respective parental tumors (derived from FFPE-tissue) were analyzed for phenotypic difference

first: Michael Karl

first: Yanchun

last: Ma

first: Jessica

last: Lindenmayer

first: Clara

last: Morgenstern

first: Felix

last: Wezel

first: Friedemann

last: Zengerling

first: Cagatay

last: Günes

first: Nadine Therese

last: Gaisa

first: Alexander

last: Kleger

first: Christian

last: Bolenz


title: CD4T in breast cancer

city: Guangzhou

first: Penghan

last: Huang

address: 107 Yan Jinag West Road

institute: Sun Yat-Sen University

summary: We found that a distinctive subset of IL-1+ CXCL3+ CD4 (Fer-CD4) T lymphocytes that were enriched in chemoresistant tumors and revealed the immune-reshaping capacity of neutrophil ferroptosis and its reciprocal feedback regulation with Fer-CD4 T cells, identifying promising targets for restoring anticancer immunosurveillance and reversing chemoresistance.

overall_design: Isolation and transcriptome sequencing of chemotherapy sensitive and resistant patient CD4T

first: Wenfeng

last: Zeng

first: Ruihua

last: Zhang

first: Liyan

last: Lao


title: Comparative single-cell transcriptomics of human neuroblastoma and preclinical models reveal conservation of adrenergic cell state

city: Boston

first: Shenglin

last: Mei

address: 10 Shattuck Street

institute: Harvard medical school (DMBI)

summary: Transgenic mice and organoid models, such as three-dimensional tumoroid cultures, have emerged as powerful tools for investigating cancer development and targeted therapies. Yet, the extent to which these preclinical models recapitulate the cellular identity of heterogeneous malignancies, such as neuroblastoma (NB), remains to be validated. Here we characterize the transcriptional landscape of TH-MYCN tumors by single-cell RNA sequencing (scRNA-seq) and developed novel ex vivo tumoroids. We provide a comparative analysis with murine fetal adrenal gland and human NB, to decipher the heterogeneity, cell-cell interactions, and clinical relevance of these models. Integrated analysis with fetal adrenal samples confirmed that both TH-MYCN tumors and tumoroids closely mirror the cellular profiles of normal embryonic sympathoblasts and chromaffin cells. Comprehensive comparison between tumors from NB patients and TH-MYCN mice demonstrated similarities in adrenergic tumor cell composition. Through ex vivo tumoroid cultures, we demonstrated histological resemblance, and shared transcriptional profiles with originating TH-MYCN tumors and human NB. Importantly, we identified subpopulations within tumoroids that exhibit gene expression associated with poor NB patient survival. Notably, recurrent observations of a low-proliferative chromaffin phenotype connected to the highly proliferative sympathetic phenotype may suggest that pushing sympathoblasts into a chromaffin-like state may offer an interesting therapeutic strategy for NB. Together, this study not only deepens our understanding of the widely used transgenic mouse model but also introduces a novel ex vivo model that maintains critical adrenergic cell state identity, thereby enhancing its translational potential for NB research.

overall_design: Tumors and ex vivo cultures from TH-MYCN transgenic mice were disaggregated and profiled using the Chromium Single Cell 3’ Library and Gel Bead Kit v2 and v3 (10X Genomics) .

first: BETHEL

last: EMBAIE

first: NINIB

last: BARYAWNO

first: HIRAK

last: SARKAR


title: Neratinib derivative 7A induces apoptosis in colon cancer cells via the p53 pathway

city: Lianyungang

first: wenhao

last: cheng

state: Jiangsu

address: 59 Cangwu Road, Haizhou District

department: College of Pharmacy

institute: Jiangsu Ocean University

summary: Colorectal cancer remains a significant health threat, with its incidence continuously rising, underscoring the urgent need for the development of new therapeutic agents. In our previous research, we identified 7A, a derivative of Neratinib, as having pronounced antitumor activity. However, its specific effects and mechanisms in colorectal cancer have not been thoroughly investigated. Therefore, this study employed in vivo and in vitro experiments, utilizing techniques such as RNA sequencing, Western blotting, and PCR, to provide a comprehensive analysis of 7A's mechanism of action in colorectal cancer. The results indicate that 7A induces DNA damage and activates the P53 pathway, thereby promoting apoptosis in colorectal cancer cells. Additionally, 7A treatment significantly reduced angiogenesis and tumor weight. Our findings suggest that 7A, a Neratinib derivative, holds promise as a novel candidate for colorectal cancer therapy.

overall_design: In order to investigate the role and mechanism of 7A in colon cancer, we treated HCT116 cells with 7A and without treatment. Then, we performed gene expression profiling analysis using RNA seq data from 7A and DMSO cells.

first: Zhiyu

last: Liu


title: Hyperglycemia-triggered lipid peroxidation destabilizes STAT4 and impairs anti-viral Th1 responses in type 2 diabetes

city: Hong Kong

first: Weixin

last: Chen

address: 21 Sassoon Road

country: Hong Kong

institute: The University of Hong Kong

summary: Type 2 diabetes (T2D) patients are more susceptible to severe respiratory viral infections, but the underlying mechanisms remain elusive. Here, we show that both COVID-19-infected T2D patients and influenza-infected T2D mice exhibit a defective Th1 response, which is an essential component of anti-viral immunity. This defect stems from intrinsic metabolic perturbations in CD4+T cells driven by hyperglycemia. Mechanistically, hyperglycemia triggers mitochondrial dysfunction and excessive fatty acid synthesis, leading to elevated oxidative stress and aberrant lipid accumulation within CD4+T cells. These abnormalities promote lipid peroxidation (LPO), which drives the carbonylation of STAT4, a crucial Th1 lineage-determining factor. Carbonylated STAT4 undergoes rapid degradation, causing reduced T-bet induction and diminished Th1 differentiation. LPO scavenger ameliorates these Th1 defects in T2D patients with poor glycemic control, and restores viral control in T2D mice. Thus, this hyperglycemia-LPO-STAT4 axis underpins reduced Th1 activity in T2D hosts, with important implications for managing T2D-related viral complications.

overall_design: PBMCs were isolated from healthy donors, well-controlled T2D, and poor-controlled T2D patients and stimulated with aCD3/aCD28 antibodies (biolegend) in T cell medium (RPMI 1640 medium supplemented with 10% FBS, 50M -mercapteoethanol, 1M HEPES, 1X non-essential amino acids (NEAA), 100mM sodium pyruvate, and 1% penicillin-streptomycin) for 48 hours. Untreated and stimulated PBMCs were submitted for scRNA-seq sequencing with Single Cell Gene Expression Flex (10X Genomics). Alive naï4+ T cells (CD45RA+CCR7+CD4+) were sorted from healthy donors, well-controlled T2D, and poor-controlled T2D patients through FACs with Influx (BD Biosciences). Their RNA was extracted using RNeasy Mini Kit (Qiagen) and submitted for paired-end RNA sequencing. Reads were mapped to UCSC human genome GRCh38/hg38 using STAR (version 2.6.1) and gene counts were quantified by featureCounts (version 1.6.4).

first: Victor

last: Gray

first: Rachael

middle: J

last: Tan

first: Jia

last: Teo

first: Zhihao

first: Carol

middle: H

last: Fong

first: Tommy

last: Law

first: Zi-Wei

last: Ye

first: Shuofeng

last: Yuan

first: Xiucong

last: Bao

first: Ivan

middle: F

last: Hung

first: Kathryn

first: Chi-Ho

last: Lee

first: Guang

last: Ling


title: Gene expression-based, individualized outcome prediction for surgically treated pancreatic cancer patients

city: Kamakura

first: Satoshi

last: Kondo

state: Kanagawa

address: 10-1 tebiro 6-chome

institute: Toray Industries,Inc.

summary: Individualized outcome prediction classifiers were successfully constructed through expression profiling of a total of 8,644 genes in 20 pancreatic cancer cases, which had been consecutively operated.

overall_design: In the study presented here, a consecutively operated, well-defined cohort of 20 pancreatic cancer cases, followed up more than five years, was used to acquire expression profiles of a total of 8,644 unique genes, leading to the successful construction of supervised

first: H

last: Ueda

first: M

last: Kubo

first: Y

last: Tomimaru

last: Takahashi

last: Eguchi


title: Mouse liver fxr chip-seq II

city: Wenzhou

first: yushu

last: Hou

state: Zhejiang

address: Wenzhou Medical University, University Town, Chashan, Wenzhou, Zhejiang, P.R China

department: School of Pharmaceutical Sciences

institute: Wenzhou Medical University

summary: We analyzed genome-wide FXR binding in liver of mice treated with GW4064 by ChIP-seq

overall_design: Freshly liver samples  was fixed in 1% formaldehyde for Cross-Linking of DNA and Protein. Liver was sonicated to fragment chromatin DNA.Chromatin was immunoprecipitated overnight at 4 °C using FXR antibody (Cell Signaling Technology, 72105) .The sequencing libraries of input and IP were generated by NEBNext®UltraTM DNA Library Prep Kit for Illumina.The library products corresponding to 1100-500 bps were enriched, quantified and finally sequenced on Novaseq6000 sequencer (Illumina) with PE150 model.

first: Lintao

last: Song

first: Yushu

first: Da

last: Xu

first: Yi

last: Liu


title: Decoding muscle-resident Schwann cell dynamics during neuromuscular junction remodeling

city: Ann Arbor

first: Susan

middle: V

last: Brooks

state: Michigan

address: 109 Zina Pitcher Pl

department: Molecular and Integrative Physiology

institute: University of Michigan

summary: Understanding neuromuscular junction (NMJ) repair mechanisms is essential for addressing degenerative neuromuscular conditions. Here, we focus on the role of muscle-resident Schwann cells in NMJ reinnervation. Using an accepted model of progressive NMJ degeneration, Sod1-/- mice, we identified a clear NMJ ‘regenerative window’ that allowed us to define cellular and molecular regulators of synapse remodeling and muscle fiber reinnervation. High-resolution imaging and single-cell RNA sequencing provide a detailed analysis of Schwann cell number, morphology, and transcriptome revealing multiple subtypes, including a previously unrecognized terminal Schwann cell (tSC) population expressing a synapse promoting signature. We also discovered a novel SPP1-driven cellular interaction between myelin Schwann cells and tSCs and show that it promotes tSC proliferation and reinnervation following nerve injury in wild type mice. Our findings offer important insights into molecular regulators critical in NMJ reinnervation that are mediated through tSCs to maintain NMJ function.

overall_design: S100GFPtg and S100GFPtg Sod1-/- mice were bred at the University of Michigan, were fed normal chow ad libitum, and housed 12:12 hour light-dark cycle under the UofM veterinary staff supervision. All procedures were approved by the University Committee on the Use and Care of Animals at UM and were in accordance with the U.S. National Institute of Health (NIH). We used 2-month-old male mice from each genotype (n=4 per group). We simultaneously harvested and processed hind limb muscles (Tibialis Anterior and Gastrocnemius) from both groups. Using Fluorescence-Activated Cell Sorting, we separated GFP+ and GFP- single cells. The sorted cells were then prepared for library construction and underwent single-cell RNA Sequencing, all processed in the same batch for consistency.

first: Steve

middle: D

last: Guzman

first: Ahmad

last: Abu-Mahfouz

first: Carol

last: Davis

first: Lloyd

last: Ruiz

first: Peter

last: Macpherson


title: Dynamic coupling of R-loops with transcriptional pausing modulates tissue development in Drosophila

city: Wuhan

first: Shaobo

last: Liang

state: Hubei Sheng

address: BaYi Road, LUOJIA HILL

institute: Wuhan University

summary: The R-loop in the genome participates in modulating transcription, replication and DNA damage repair, yet its regulation and biologic impact on development in vivo remain to be explored. Here, we modified CUT&Tag strategy and established R-loop profiles of Drosophila across multiple developmental stages. While lack of GC preference, the broad class of R-loops in Drosophila all highly follow the RNAPII pausing, which is also a conserved R-loop regulatory mechanism in mammals. Meanwhile, RNAPII-regulated R-loop may be regulated by multiple developmental specificity and universality transcription factors. To test the specificity and universality of transcription factor-mediated R-loop involvement in development, we knocked out the transcription elongation factor Spt6, upregulated R-loop and increased RNAPII pausing, and altered the expression of critical genes and cutin genes, thereby affecting Drosophila lifespan and cutin development. Interestingly, complementation of rnh1 could partially rescue the transcription factor deletion phenotype. We conclude that RNAPII is a conserved mechanism of R-loop regulation across species and organism specific damage caused by R-loop dysregulation is mediated by the functions of transcription factors.

overall_design: To explore the regulatory mechanism of R-loop in Drosophila and embryonic development, we constructed a multi-temporal R-loop map of embryonic development. As well as pol2-chip-seq, RNA-seq and cut&tag before and after interfering transcription factor spt6

first: Liang

last: Chen

first: Xianhong

last: Zhang


title: Transcriptomic analysis of gastric cancer cells in response to stable over-expression of circRNA derived from hnRNPU (circ-hnRNPU)

first: Qiangsong

last: Tong

state: Hubei

address: 1277 Jiefang Avenue

department: Department of Surgery

institute: Union Hospital of Tongji Medical College

summary: Circular RNA (circRNA) is a type of endogenous single-stranded and covalently closed non-coding RNA that plays emerging roles in tumorigenesis and aggressiveness. However, the functions and underlying mechanisms of circRNAs in regulating gastric cancer progression still remain elusive. We identify circ-hnRNPU, a circRNA derived from exons 5, 6, 7 and 8 of heterogeneous nuclear ribonucleoprotein U (hnRNPU), as a tumor suppressor of gastric cancer progression. To investigate the mechanisms underlying the functions of circ-hnRNPU, we employed the Illumina HiSeq X Ten as a discovery platform to analyze the transcriptome profiling changes of human gastric cancer MKN-45 and AGS cells in response to stable over-expression of circ-hnRNPU. The results showed that stable over-expression of circ-hnRNPU led to altered expression of 112 human mRNAs, including 64 up-regulated and 48 down-regulated genes, in both MKN-45 and AGS cells. Furthermore, we validated the RNA-seq results by real-time RT-PCR with high identity. Overall, our results provided fundamental information about the transcriptomic changes in response to circ-hnRNPU over-expression in human gastric cancer cells, and these findings will help us understand the pathogenesis of cancer progression.

overall_design: Total RNA of cells stably transfected with empty vector or circ-hnRNPU was extracted using the TRIzol® reagent according to the manufacturer's instructions. RNA concentration was measured using a Qubit® RNA Assay Kit with a Qubit® 2.0 Fluorometer (Life Technologies, Inc.), and integrity was assessed using the RNA Nano 6000 Assay Kit with a Bioanalyzer 2100 system (Agilent Technologies, CA). Library preparation and transcriptome sequencing on an Illumina HiSeq X Ten platform were performed by Wuhan SeqHealth Technology Co., Ltd. (Wuhan, China), and 100 bp paired-end reads were generated. HTSeq v0.6.0 was used to count the reads numbers mapped to each gene.

first: Tong

last: Qiangsong

first: Zheng

last: Liduan


title: Translational reprogramming during epithelial-mesenchymal transition is associated with fine-tuning of ribosome composition

city: Orsay

first: Olivier

last: Namy

address: Rue Gregor Mendel, bat 400

institute: CNRS, Université Paris-Sud

summary: Epithelial-mesenchymal transition (EMT) is driven by transcriptional and epigenetic reprogramming. However, it is now well established that the final step of gene expression, translation, is also key to the establishment of physiopathological phenotypes, and one of its regulators is the ribosomes themselves. Using genome-wide analyses, we report on how the translational landscape evolves and how the translational machinery rearranges itself during EMT. Ribosome profiling revealed that translation was as profoundly modulated as transcription during EMT. In addition, riboproteomics revealed an increased proportion of RPL36A-containing ribosomal particles in mesenchymal cells, indicating that ribosome composition is finely tuned during EMT. Finally, RPL36A, but not RPL19, is sufficient to induce the acquisition of mesenchymal features. Taken together, these data demonstrate that EMT is associated with both a profound translational reprogramming and changes in ribosome composition, and that a single change in ribosomal protein is sufficient to drive EMT.

overall_design: RiboSeq and RNASeq before et after induction of the EMT by overexpression of Zeb1 gene

first: Chloé

last: Morin

first: Agnès

last: Baudin-Baillieu

first: Flora

last: Long

first: Caroline

last: Isaac

first: Laure

last: Bidou

first: Hugo

last: Arbes

first: Pauline

last: François

first: Roxanne

last: Pommier

first: Annie

last: Adrait

first: Akari

last: Saku

first: Stephanie

last: Gran-Ruaz

first: Camélia

last: Machkouri

first: Frédéric

last: Catez

first: Anne-Pierre

last: Morel

first: Yohann

last: Couté

first: Sophie

last: Chat

first: Reynalt

last: Gillet

first: Alain

last: Puisieux

last: Moyret-Lalle

first: Jean-Jacques

last: Diaz

first: Virginie

last: Marcel


title: Adenine base editors induce off-target structure variations in mouse embryos and primary human T cells

status: Public on Nov 01 2024

city: Shanghai

first: Leilei

last: Wu

address: RD215 FuTe south

institute: Epigenic Therapeutics, Inc

summary:
Background
The safety of CRISPR-based gene editing methods is of the utmost priority in clinical applications. Previous studies have reported that Cas9 cleavage induced frequent aneuploidy in primary human T cells, but whether cleavage-mediated editing of base editors would generate off-target structure variations remains unknown. Here, we investigated the potential off-target structural variations associated with CRISPR/Cas9, ABE and CBE editing in mouse embryos and primary human T cells by whole-genome sequencing and single-cell RNA-seq analyses.
Results
The results showed that both Cas9 and ABE generated off-target structural variations (SVs) in mouse embryos, while CBE induced rare SVs. In addition, off-target large deletions were detected in 32.74% of primary human T cells transfected with Cas9 and 9.17% of cells transfected with ABE. Moreover, Cas9-induced aneuploid cells activated the P53 and apoptosis pathways, whereas ABE-associated aneuploid cells significantly upregulated cell cycle-related genes and arrested in G0 phase. A percentage of 16.59% and 4.29% aneuploid cells were still observable at 3 weeks post transfection of Cas9 or ABE. These off-target phenomena in ABE were universal as observed in other cell types such as B cells and Huh7. Furthermore, the off-target SVs were significantly reduced in cells treated with high-fidelity ABE (ABE-V106W).
Conclusions
This study raises urgent need for minimizing the off-target SVs of CRISPR/Cas9 and ABE.

overall_design: In this study, two target genes, PDCD1 and B2M, were edited. The same single guide RNA (sgRNA) used by Gaudelli et al. [31] to target B2M was also used in our study, successfully targeting both by ABE and Cas9 through splice site disruption. For PDCD1, we used the same sgRNA as Nahmad et al. [4] did for Cas9 and Gaudelli et al. [31] did for ABE. For high-fidelity ABE, the same sgRNA targeting PDCD1 was used. For Huh7 ,, the same sgRNA targeting B2M was used. Prior to the experiment, T Cell Medium (Lonza X vivo5 + 5% human AB serum + NAC + 10ng/ml IL7 and 10ng/ml IL15) was prepared and allowed to equilibrate at room temperature for 20 minutes within a biological safety cabinet. Cryopreserved PBMCs were then retrieved from a liquid nitrogen tank, thawed in a 37°C water bath, and transferred into the cabinet after removing any remaining ice clumps. The outer surface of the container was sterilized with 5% ethanol, dried, and transferred into the cabinet. The cells were then transferred into 9 mL of T cell medium and centrifuged at 400g for 5 minutes at room temperature. After discarding the supernatant, the cells were resuspended in 5 mL of medium and counted using a ThermoFisher Attune NxT flow cytometer. Subsequently, the cells were seeded at a density of 1e6 cells/mL in plates or culture flasks. After 6 hours of PBMC revival, CD3/28 dynabeads were added for activation. Following 3 days of cell activation, the activation beads were removed using a magnet and the culture was continued for 24 hours. For electroporation, 24-well plates were prepared by adding 900 μL of PBMC complete medium per well and incubating at 37°C for 10 minutes. The Lonza 4D Nucleofector System with the P3 Primary Cell 4D kit was used, following the kit instructions for electroporation. Each electroporation reaction consisted of 1E6 cells, 1.2 pmol of EPIREG mRNA, and 40 pmol of sgRNA for each target gene. Immediately after electroporation, 100 μL of cell-free medium was added to each well, followed by a 1-hour incubation at 37°C. The cells were then transferred to 24-well plates for further culture. To assess changes in the protein levels of PDCD1 and B2M on T cell surfaces using flow cytometry, approximately 1e5 cells were transferred to 1.5 mL Eppendorf tubes and centrifuged at 400g for 5 minutes at room temperature. The supernatant was discarded and 50 μL of the corresponding dilution ratio of antibodies was added. The tubes were incubated at 4°C in the dark for 30 minutes, followed by the addition of 500 μL of MACS buffer to each tube. After centrifuging at 400g for 5 minutes at room temperature, the supernatant was discarded and 300 μL of MACS buffer was added to each tube, gently mixed by pipetting. The samples were then analyzed using a ThermoFisher Attune NxT flow cytometer. The 10x Genomics® Cell Preparation Guide was followed for washing, counting, and concentrating cells from both abundant and limited cell suspensions (greater than or less than 100,000 total cells, respectively) in preparation for use in 10x Genomics Single Cell Protocols. The cell suspension was loaded into Chromium microfluidic chips with 3' (v2 or v3, depending on the project) chemistry and barcoded with a 10× Chromium Controller (10X Genomics). RNA from the barcoded cells was subsequently reverse-transcribed and sequencing libraries were constructed using reagents from a Chromium Single Cell 3' v2 (v2 or v3, depending on the project) reagent kit (10X Genomics) according to the manufacturer's instructions. Sequencing was performed with Illumina (HiSeq 2000 or NovaSeq, depending on the project) according to the manufacturer's instructions (Illumina). The same library construction process was used in Huh7 cells.


title: Preconditioning With Electrical Stimulation Promotes Flap Survival by Improving Blood Supply Via ROS/Nrf2/HO1 Pathway

city: Academic

first: Meijun

last: Jiang

state: Changchun

address: Changchun, 130021, People’s Republic of China

department: Department of Hand and Podiatric Surgery, Orthopedics Center

institute: The First Hospital of Jilin University

summary: We have previously demonstrated that preconditioning flap donor with electrical stimulation (ES) could increase the survival area of pedicled perforator flaps in a rat model. However, the detailed molecular mechanisms remain unclear. In this study, we first verified the effects of ES through in vivo and in vitro experiments. Then we explored the underlying mechanisms in vitro. The results indicated that preconditioning with ES could not only increase the flap survival area but mitigate human umbilical vein endothelial cells (HUVECs) dysfunction. Furthermore, low level reactive oxygen spices (ROS) were produced in HUVECs after ES, serving assignaling molecule to activate the nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HOl) pathway. Then ROS scavenger was used to attenuate thegeneration of ROS, resulting that activation of the Nrf2/HOl pathway was inhibited. These results suggest that ROS/Nrf2/HO1 pathway may play pivotal roles in the ES pretreatment induced flap survival.

overall_design: To validate this hypothesis, we utilized the modified McFarlane random flap model in rats and the human umbilical veinendothelial cells (HUVECs)oxygen-glucose deprivation (OGD) model, the study primarily measured the survival area of flaps on postoperative day 7 and assessed cell proliferation activity, both in vivo and in vitro, to confirm the protective effect of electrical stimulation preconditioning against ischemia. Subsequently,Subsequently, the ROS scavenger N-acetyl-L-cysteine (NAC) was introduced to explore potential mechanistic pathways.

first: Xin

last: Wei

first: Pengcheng

last: Liu

first: Xu

last: Gong


title: The RNA exosome maintains cellular RNA homeostasis by controlling transcript abundance in the brain

city: Los Angeles

first: Lauryn

middle: Alexis

last: Higginson

address: 1050 Childs Way Room 201B

department: Biological Sciences

institute: University of Southern California

summary: Intracellular ribonucleases (RNases) are essential in all aspects of RNA metabolism, including maintaining accurate RNA levels. Inherited mutations in genes encoding ubiquitous RNases are associated with human diseases, primarily affecting the nervous system. Recessive mutations in genes encoding an evolutionarily conserved RNase complex, the RNA exosome, lead to syndromic neurodevelopmental disorders characterized by progressive neurodegeneration, such as Pontocerebellar Hypoplasia Type 1b (PCH1b). We establish a CRISPR/Cas9-engineered Drosophila model of PCH1b to study cell-type-specific post-transcriptional regulatory functions of the nuclear RNA exosome complex within fly head tissue. Here, we report that pathogenic RNA exosome mutations alter activity of the complex, causing widespread dysregulation of brain-enriched cellular transcriptomes, including rRNA processing defects—resulting in tissue-specific, progressive neurodegenerative effects in flies. These findings provide a comprehensive understanding of RNA exosome function within a developed animal brain and underscore the critical role of post-transcriptional regulatory machinery in maintaining cellular RNA homeostasis within the brain.

overall_design: Two biological replicates of newly eclosed adult female flies were prepared for each biological replicate (Rrp40WT, Rrp40G11A, and Rrp40G146C). For each sample, 20 fly heads were pooled together, and nuclei were dissociated. Nuclei quantity and and quality were determined using propidium iodide. Single-cell libraries were prepared using Chromium Next GEM Single Cell 3' GEM, Library & Gel Bead Kit v3.1. Sequencing reads were mapped to the Drosophila melanogaster genome and CellRanger was used to create the reference.

first: Derrick

last: Morton


title: Intermittent and sustained hypoxia exposures activate distinct transcriptional responses in human aortic endothelial cells

city: Chicago

first: Rengul

last: Cetin-Atalay

address: 5841 S. Maryland Avenue

department: Medicine

institute: The University of Chicago

summary: Endothelial cell (EC) dysfunction precedes the development of cardiovascular disease in OSA; however, the mechanisms by which ECs respond to these hypoxic events is poorly understood.  To better understand EC responses to hypoxia, we examined the effects of both sustained hypoxia (SH) and intermittent hypoxia (IH) on the activation of HIF-1α in Endothelial Cells (ECs). While SH stabilized HIF-1α and led to its nuclear localization, IH didn’t activate HIF-1α and its downsteram gene expression. Using RNA-sequencing, we evaluated transcriptional responses of ECs to hypoxia. SH induced expression of HIF-1α and hypoxia response genes, while IH affected cell-cycle regulation genes.

overall_design: HAEC cells

first: Gokhan M

last: Mutlu

first: Angelo Y

last: Meliton

first: Robert S

last: Hamnaka


title: The contribution of foam cells to angiogenesis and inflammation after spinal cord injury

first: Yersen

last: Mulat

state: Shanghai

address: No. 650 Xin Songjiang Road

department: School of Medicine

institute: Shanghai Jiao Tong University

summary: After spinal cord injury (SCI), macrophages engage in the phagocytosis of myelin debris and dead cells, transforming into foam cells. Nevertheless, the ultimate fate and functional role of these foam cells remain unclear, while their inflammatory phenotype elicits ongoing controversy. We discovered that macrophages transform into foam cells through 3 stages, which were phagocytosis, digestion, and lipid overload. Macrophages possess a certain degree of antioxidant capacity. When confronted with excessive lipid pressure, they undergo lipid droplets formation and metabolic reprogramming to alleviate lipotoxicity, also promoting angiogenesis. The anti-inflammatory phenotype of macrophages gradually diminished after SCI, phagocytosing debris could assist in maintaining this phenotype. Moreover, most genes expression patterns were conserved both in the SCI mice and rats. Our study provides a detailed description of the changes in gene expression and function during foam cell formation and long-term fate after SCI, which provides a theoretical basis for further research.

overall_design: To investigate whether the genetic profile of bone marrow-derived macrophages (BMDM) changes when they engulf myelin debris (MD) under different conditions, we conducted RNA-seq sequencing, including genetic profiling of BMDM in different polarized states (M1, M2), MD ingested at different concentrations (50ug/ml, 1mg/ml) and at different times (6h, 24h, 72h), as well as genetic profiling of BMDM after they have been treated with two possible drugs that may reduce lipid content within BMDM: badoxifene and bafometrin.

first: Haodong

last: Lin

first: Ying

last: Peng

first: yersen

last: mulat

first: Zhenwu

last: Yang


title: Local Delivery of an IL-15 Superagonist Using a Replicating Retrovirus Improves Survival and Lymphocyte Infiltration in Poorly Immunogenic Mouse Models of Glioblastoma

city: San Francisco

first: Manish

last: Aghi

address: 505 Parnassus Avenue, #M774A

institute: UCSF

summary: Glioblastoma (GBM) leads to severe systemic and local immunosuppression, and immunotherapies have had limited success. One of the hallmarks of GBM immunosuppression is poor T cell infiltration and significant intratumoral T cell dysfunction. IL-15 is a promising immunocytokine with the ability to stimulate T cell activation and proliferation. In this study, we evaluate the treatment efficacy of RLI (an IL-15 superagonist) delivered to tumor cells using a tumor-selective retroviral replicating vector (RRV) in the poorly immunogenic syngeneic murine SB28 and Tu2449 models of GBM. RRV-RLI replicates and spreads effectively in cultured SB28 cells. Transduced SB28 cells express high levels of RLI (175 ng/mL), which sustained T cell growth in culture. Treatment with RRV-RLI by stereotactic injection into intracerebral murine SB28 and Tu2449 tumors significantly reduced tumor growth on bioluminescence imaging, and increased survival relative to control mice, leading to long-term survival in a subset of treated mice. RRV-RLI treated tumors showed significantly increased CD4 and CD8 T cell infiltration, without changes in immunosuppressive cell populations, including myeloid-derived suppressor cells or T regulatory cells. This study demonstrates that localized tumor-specific immunomodulatory gene therapy delivered in an RRV has the potential to safely reverse the T-cell depleted immunophenotype of GBM.

overall_design: A retroviral replicating vector was used to deliver RLI (IL-15 superagonist) in vivo to tumor cells in the SB28 murine model of GBM. Conditions tested included a (1) PBS + vehicle control, (2) PBS + TMZ, (3) RRV RLI + vehicle, and (4) RRV RLI + TMZ. CD45+ leukocytes were then isolated from these tumors using flow cytommetry and were sent for single-cell RNA sequencing (scRNA-seq) as well as single cell immune profiling of the V(D)J repertoire of T-cells.

first: Haddad

last: Alexander F

first: Kasahara

last: Noriyuki

first: Aghi

last: Manish K


title: α4 Blockade Reduces CD8 TEM Cells in the Brain and Elevates CNS Viral Loads

city: Pittsburgh

first: Smita

last: Iyer

state: Pennsylvania

address: 3550 Terrace Street

department: Pathology

institute: University of Pittsburgh School of Medicine

summary: Background. The blood-brain barrier (BBB) regulates immune cell entry into the CNS, and various strategies have been developed to target this barrier in efforts to alleviate neuroinflammation in neurodegenerative diseases. α4β1 is crucial for T cell influx into the CNS. In the context of HIV, CD4 T cells drive viral infiltration, while CD8 T cells are essential for viral clearance. This dual role complicates strategies aimed at targeting T cell entry during HIV infection, yet it remains an area that has not been thoroughly explored. Methods. We investigated the impact of disrupting α4-mediated T cell entry into the CNS in SIVmac251-infected rhesus macaques treated with an α4-blocking antibody (n=4) versus an isotype control (n=4). Our comprehensive approach included single-cell analysis of CD45+ cells in the brain and spleen, spatial transcriptomics of the hippocampus, viral load measurements, and flow cytometry on brain tissue. Results. Single-cell analysis revealed a marked reduction in the brain CD8 TEM cluster, particularly in cells expressing Granulysin, CCL5, and STAT4, while CD4 T cell clusters remained unchanged, and two monocyte clusters were enriched. This shift was accompanied by an increase in brain SIV RNA+ CD45+ CD4+ T cells in the α4-treated group (vRNA+ cells = 60) compared to controls (n=11). Spatial transcriptomics of SIV vRNA+ versus SIV vRNA- regions in the hippocampus of α4-treated macaques showed enrichment of CD4 TCM and monocyte gene signatures. Correspondingly, quantitative PCR revealed higher vRNA levels in the gray matter of the PFC, STS, and Hp. These findings suggest that α4 blockade selectively disrupts CD8 TEM populations while sparing CD4 T cells, leading to elevated viral loads and heightened neuroinflammation. The increase in SIV RNA+ CD4+ T cells and viral loads in the gray matter furthermore underscores the compartmentalized effect of α4-mediated immune modulation on viral dynamics across distinct brain regions Conclusion. These results underscore the critical need to evaluate immune-targeting therapies, with implications for strategies aimed at controlling T cell influx in the context of viral infections in the brain.

sample_organism:
Macaca mulatta
blank sample

platform_organism:
Macaca mulatta
blank sample

overall_design: Sections from the prefrontal cortex and hippocampus of six rhesus macaques were collected 3 weeks post-infection with SIVmac251. Animals were treated with either a monoclonal antibody targeting α4 integrin (n=3) or an isotype control (n=3). Morphological markers, including CD45, CD3, and NeuN, were used to define regions of interest. RNAscope was employed to further delineate SIV-positive and SIV-negative regions for transcriptomic analysis.

first: Petkov

last: Alex

first: Elizaldi

last: Sonny

first: Newhouse

last: Daniel

first: Iyer

last: Smita


title: Mast cell heterogeneity in human nasal polyps

first: Daniel

last: Dwyer

state: MA

address: 60 Fenwood Road

institute: Brigham and Women's Hospital

summary: Mast cells (MCs) expressing a distinctive protease phenotype (MCTs) selectively expand within the epithelium of human mucosal tissues during type 2 (T2) inflammation. While MCTs are phenotypically distinct from subepithelial MCs (MCTCs), signals driving human MCT differentiation and this subset’s contribution to inflammation remain unexplored. Here, we identify TGF-b as a key driver of the MCT transcriptome in nasal polyps. We find that short-term TGF-b signaling alters MC cell surface receptor expression and partially recapitulates the in vivo MCT transcriptome, while TGF-β signaling during MC differentiation upregulates a larger number of MCT-associated transcripts and inhibits the hallmark MCTC proteases chymase and cathepsin G at both the transcript and protein level, allowing selective in vitro differentiation of MCTs for functional study. We identify discrete differences in effector phenotype between in vitro-derived MCTs and MCTCs, with MCTs greatly enhancing pro-inflammatory lipid mediator generation while selectively modifying cytokine, chemokine, and growth factor production in response to both innate and adaptive stimuli, recapitulating functional features of their tissue-associated counterpart MC subsets. Thus, our findings support a role for TGF-β in promoting human MCT differentiation and identify a discrete contribution of this cell type to T2 inflammation.

overall_design:
Mast cells were isolated based on co-expression of FceR1a and CD117 from nasal polyp tissue of four donors with aspirin-exacerbated respiratory disorder (AERD) for single-cell RNA sequencing.

***************************************************************
Samples are from human patients, raw data (Fastq files) are in the process of being deposited in dbGaP
***************************************************************

first: Tahereh

last: Derakhshan


title: Decoding Aging in the Heart via Single Cell Dual Omics of Non-Cardiomyocytes (scATAC-Seq)

city: Chapel Hill

first: Yiran

last: Song

state: NC

address: 111 Mason Farm Road

institute: The University of North Carolina

summary: (scRNA and scATAC) in profiling non-myocytes (non-CMs) from young, middle-aged, and elderly mice. Non-CMs, vital in heart development, physiology, and pathology, are understudied compared to cardiomyocytes. Our analysis revealed aging response heterogeneity and its dynamics over time among non-CM cell types. Immune cells, notably macrophages and neutrophils, showed significant aging alterations, while endothelial cells displayed moderate changes. We identified distinct aging signatures within the cell type, including differential gene expression and transcription factor activity, along with motif variation. Sub-cluster analysis revealed intra-cell type heterogeneity, characterized by diverse aging patterns. The senescence-associated secretory phenotype (SASP) emerged as a key aging-related phenotype. Moreover, aging significantly influenced cell-cell communication, especially impacting a fibroblast sub-cluster with high expression of Erbb4. This study elucidates the complex cellular and molecular landscape of cardiac aging in non-CMs, highlighting their importance in cardiac aging and offering guidance for future potential therapeutic avenues to treat aging-related heart diseases.

overall_design: Male mice (C57BL/6J strain) aged at 3 months, 12 months, and 24 months each for 2 replicates of scRNA and scATAC seq were collected for non-cardiomyocyte isolation.

first: Li

last: Wang

first: Haofei

first: Hong

last: Ma

first: Jiandong

last: Liu

last: Qian


title: Decoding Aging in the Heart via Single Cell Dual Omics of Non-Cardiomyocytes (scRNA-Seq)


title: The role of LDLR in intestinal inflammation

city: Dallas

first: Hasan

last: Zaki

address: 6000 Harry Hines Blvd

institute: UT Southwestern Medical Center

summary: Low-density lipoprotein (LDL) is a risk factor for atherosclerosis, obesity, and other metabolic diseases, but its role in colitis is unknown. Here, we investigated the role of LDL in colitis by using LDL receptor deficient mice (Ldlr-/-). Wild-type and Ldlr-/- mice were fed with 3% DSS for 5 days and colons tissues were collected at day 8 post DSS treatment. We observed that Ldlr-/- mice are less susceptible to colitis. To understand the role of LDLR in colitis pathogenesis, we isolated RNA from colons of WT and Ldlr-/- mice and perfoemed RNA-seq. Our data show that the expression of inflamamtory genes are reduced in Ldlr-/- colons compared to wild-type. These data suggest that LDL is a potent endogenous trigger for intestinal inflammation.

overall_design: Wild-type and Ldlr-Ko mice (male, 8-10 weels old) were administered with 3% DSS for 5 days. Colon tissues were collected at day 8 post DSS and RNA was isolated form the colons for RNA-seq analysis.

first: Shahanshah

last: Khan

first: Yunpeng

last: Gao

first: Ram

last: Mani


title: Effects of HSV infection on melanoma cells

city: 上海

first: 玮玮

last: 代

state: 上海

address: 徐汇区东安路131号

institute: 复旦大学

summary: To investigate the effect of HSV infection on melanoma cells, we established a model of HSV infection

overall_design: Comparative gene expression profiling analysis of RNA-seq data for B16 cells and HSV-infected B16 cell

first: Minfeng

last: Shu

first: Weiwei

last: Dai


title: BACH2 Controls Seeding and Establishment of Long-Lived HIV-1 Reservoir in CD4+ T cells

city: Saint Louis

first: Hongbo

state: MO

address: 425 S Euclid Ave

institute: Washington university in St. Louis

summary: Despite antiretroviral therapy, HIV mainly persists in memory CD4+ T cells in people living with HIV. Most long-lived viral reservoir cells are infected near the time of therapy initiation. A better understanding of the early events in reservoir seeding presents opportunities for preventing latent reservoir formation. Here, we demonstrated that CD4+ T cells expressing CCR5, permissive to HIV-1 infection, are effector or terminally differentiated cells. BACH2 is expressed by a small subset of CCR5+ cells and reverses their terminal differentiation. BACH2-mediated memory differentiation is impeded due to heightened inflammation before treatment initiation. Mice with a BACH2 knockout human immune system has a reduced frequency of HIV-1 reservoir cells and do not experience virus rebound after treatment discontinuation. Our study reveals that BACH2 is essential to the seeding of long-lived HIV-1 reservoir and demonstrates the potential of targeting BACH2 at the time of treatment initiation to eliminate HIV-1 reservoirs in T cells.

overall_design: Human primary CD4+ T cells were purified by negative selection with MojoSort™ Human CD4 T Cell Isolation Kit (Biolegend). Purified CD4+ T cells (106 cells/mL) were cultured in RPMI 1640 medium supplemented with 10% FCS, penicillin (100 unit/mL) and streptomycin (100 µg/mL). To generate activated cells, CD4+ T cells were stimulated with plate-bound anti-CD3 and anti-CD28 antibodies at 1μg/mL (BioLegend) for 3 days in the presence of 20 ng/mL IL-2. Stimulated CD4+ T cells were cultured with 20 ng/mL IL-2 for 6 days. CCR5+ and CCR5- CD4+ T cells were sorted followed by bulk RNA-Seq experiments.

first: Yuhao

last: Li

first: Kolin

last: Clark

first: Liang

last: Shan


title: MiR-335-5p escaped from circKIAA0586 adsorption contributes to mechanical overloading-induced cartilage degeneration by targeting HELLS (circRNA-Seq)

city: Guangzhou

first: Haoyu

last: Xie

state: Guangdong

address: 183 Zhongshan avenue west

institute: The Third Affiliated Hospital of Southern Medical University

summary: Mechanical overload is essential to cartilage degeneration during osteoarthritis (OA) development. Recent studies have confirmed that circular RNAs (circRNAs) are mechanically sensitive and that the circRNA-associated competitive endogenous RNA (ceRNA) network regulates cartilage degeneration and OA development. However, the involvement of the circRNA-associated ceRNA network in chondrocyte senescence induced by mechanical overloading remains unestablished.Here,we examined the circRNA, miRNA and mRNA expression of chondrocytes subjected to mechanical overloading.

overall_design: circRNA profiles of human primary chondrocytes subjected to cyclic tensile strain loading (20%) and control cells without cyclic tensile strain loading.

first: Yuheng

last: Lu

first: Jianying

last: Pan

first: Hua

last: Zeng

first: Zhicheng

last: Zhang

first: Jianbin

last: Yin

first: Jinjian

last: Zhu

first: Bingsheng

last: Luo

first: Dong

last: Guo

first: Chunyu

last: Wu

first: Chun

first: Yan

last: Shao

first: Xiaochun

last: Bai

first: Haiyan

first: Daozhang

last: Cai


title: MiR-335-5p escaped from circKIAA0586 adsorption contributes to mechanical overloading-induced cartilage degeneration by targeting HELLS (miRNA-Seq)

overall_design: miRNA profiles of human primary chondrocytes subjected to cyclic tensile strain loading (20%) and control cells without cyclic tensile strain loading.


title: MiR-335-5p escaped from circKIAA0586 adsorption contributes to mechanical overloading-induced cartilage degeneration by targeting HELLS (RNA-Seq)

overall_design: mRNA profiles of human primary chondrocytes subjected to cyclic tensile strain loading (20%) and control cells without cyclic tensile strain loading.


title: Thyroid hormone receptor beta (THRβ1) is the major regulator of T3 action in human iPSC-derived hepatocytes

city: Boston

first: Pushpinder

last: Bawa

state: Massachusetts

address: 670 Albany St

department: Boston University Medical Campus

institute: Center for Regenerative Medicine

summary:
To investigate the role of THRβ1 action  in human iPSC-derived hepatocytes.
We then performed gene expression profiling analysis using data obtained from RNA-seq of  WT and THRBKO human iPSCs-derived hepatocytes in presence and absence of T3.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for BU3-10-CR2 cells (WT) and its KO derivative (THRBKO) in presence and absence of T3.

first: Lorraine

last: Soares De Oliveira


title: Bulk transcriptomic analysis of GIP-expressing cells in organoids derived from human duodenum.

city: Cambridge

first: Fiona

last: Gribble

state: Cambridgeshire

address: Box 289 Addenbrooke's Hospital, Hills Road

department: Medical Research Laboratories

country: United Kingdom

institute: University of Cambridge

summary: Glucose-dependent insulinotropic polypeptide (GIP) is an incretin hormone secreted by enteroendocrine K-cells in the proximal small intestine. This study aimed to explore the function of human K-cells at the molecular and cellular level.

overall_design: Organoids were generated using tissue from human duodenum. Venus fluorescent marker was inserted into GIP locus using CRISPR-Cas9 technology. Venus-positive cells from differentiated GIP-Venus organoids, and corresponding Venus-negative cells, were purified by flow-cytometry, and analysed by bulk RNA sequencing.

first: Christopher

last: Smith

first: Nunzio

last: Guccio


title: Epigenome analysis of MTHFD2 as a biomarker for decitabine response in MDA-MB-231 tumours

city: Halifax

first: Paola

last: Marcato

state: Nova Scotia

address: 5850 College St

country: Canada

institute: Dalhousie University

summary: Genome-wide DNA methylation profiling of MDA-MB-231 tumours with or without MTHFD2 knockdown and decitabine treatment. The Illimuna Infinium MethylationEPIC DNA  Beadchip v-1-0 B3 was used to obtain DNA methylation performed by The Centre for Applied Genomics in Toronto, Canada.

overall_design: Samples from 24 MDA-MB-231 tumours with or without MTHFD2 knockdown were implanted into NOD-SCID mice that received either control PBS or decitabine treatment were analysed. The tumours had either scramble control lentiviral vector or shRNA knockdown. There were two shRNA clones, labeled shRNA 1 or 2. Eight samples from each clone were submitted, with four each of PBS control or decitabine-treated tumours. Bisulphite converted DNA from the 24 samples were hybridized to the Illumina Infinium MethylationEPIC Beadchip v-1-0 B3.

first: Olivia

last: Walker

first: Margaret

last: Dahn


title: The master transcription factor of the Plasmodium merozoite formation. [DIP-seq]

city: tsu

first: masao

last: yuda

state: mie

address: edobashi 2-174

institute: mie university

summary: To investigate the DNA-binding property of two tandem AP2 domains of PbSIP2, DNA immunoprecipitation followed by high-throughput sequencing (DIP-seq) analysis were performed. Recombinant AP2 domains fused with maltose-binding protein (MBP) were mixed with the P. berghei genomic DNA fragmented via sonication, and protein-DNA complex was harvested using amylose resin. The obtained DNA fragments were sequenced via the next generation sequencing.

overall_design: DNA fragment encoding the AP2 domains of PbSIP2 was cloned into the expression vector pMal-c5X (NEB). Escherichia coli strain DH5α transformed with this plasmid was cultured for 12 h at 37 °C. Next, expression of the MBP-fused protein was induced by adding isopropyl β-D-thiogalactopyranoside (final concentration of 200 nM) in the culture and incubating for 9 h at 25 °C. Recombinant AP2 domains fused with MBP was purified using amylose resin, and eluted with 10 mM maltose solution. Subsequently, the MBP-fused homeodomain was incubated with P. berghei ANKA genomic DNA fragments in binding/washing buffer (10 μM ZnSO4, 2 mM MgCl2, 2 mM Tris-HCl at pH 7.4, 100 mM KCl, and 10% glycerol) for 30 min. The protein/DNA solution was further mixed with amylose resin and incubated for 30 min. After incubation, the resin was washed three times with binding/washing buffer, and bound protein-DNA complexes were eluted with 10 mM maltose solution. A sequencing library was prepared from the DNA fragments and sequenced using Illumina NextSeq. Before their use for DIP, genomic DNA fragments were also sequenced as an input.

first: Tsubasa

last: Nishi

first: Izumi

last: Kaneko

first: Masao

last: Yuda


title: Effects of high doses of cholecacyferol on gene expression in the jejunum and colon of pigs

city: Kraków

first: Alicja

last: Wierzbicka

address: ul. Sarego 2

country: Poland

institute: National Research Institute of Animal Produstion

summary: High doses of vitamin D increase antiviral response but increase expression of oncogenic genes in the intestine of pigs.

overall_design: Thirty  male fatteners, the progeny of Large White sows x Landrace boar, were assigned to one of three experimental groups (n=10) through stratified randomization. The animals were provided with isoprotein and isoenergetic feeds, differing only in the amount of vitamin D (Group A, Group B -  5,000, Group C - 10,000 IU/kg of feed).

first: Maria

last: Oczkowicz

first: Tomasz

last: Szmatoła


title: Transcriptome analysis of Arabidopsis thaliana WT, mbd1 K.O., pyl5 K.O. and mbd1 pyl5  DOUBLE K.O. mutants

city: New Delhi

first: Amrapali

last: Sharma

state: Delhi

address: Benito Juarez road

department: Department of Plant Molecular Biology

institute: University of Delhi South Campus

summary: Transcriptome profiling of WT, mbd1, pyl5 and mbd1 pyl5 knock-out mutants in response to Abscisic acid (ABA) was done using Affymetrix  GenechipTM Arabidopsis ATH1 Genome Array.

overall_design: A comparative analysis of the transcriptome profiles of WT, mbd1, pyl5 and mbd1 pyl5  in response to ABA was performed. To begin with, WT, mbd1, pyl5 and mbd1 pyl5  plants were grown in ½ MS medium either lacking or supplemented with 0.3 M ABA.  Total RNA was extracted from one-week old seedlings and the quality and quantity of  the RNA was determined by Agilent Bioanalyzer and Nanodrop spectrophotometer.  Three biological replicates per sample were used for WT and mbd1 (i.e. three for ½ MS  medium and three for ABA-containing medium) while two biological replicates per  sample were used for pyl5 and mbd1 pyl5. Preparation of labelled cRNA and array  hybridization was carried out as per manufacturer’s instructions.


title: The master transcription factor of the Plasmodium merozoite formation. [ChIP-Seq]

summary: To investigate the genome-wide binding sites of PbSIP2 in Plasmodium berghei, the chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) analyses were performed. Parasites expressing GFP-fused PbSIP2 (PbSIP2::GFP)  in cultures were harvested at 24 h post infection and subjected to ChIP-seq experiments using anti-GFP antibody. Binding sites of PbSIP2 were determined from the sequence data.

overall_design: PbSIP2::GFP parasites were synchronized by injecting cultured schizonts, which were harvested by density gradient centrifugation, into mice. Chromatins associated with GFP-fused PbSIP2 were IPed with anti-GFP antibody, and DNA fragments purified from the IPed chromoatins were sequenced by the Next Generation Sequencing. Using non-IPed samples as a control (INPUT), binding sites of PbSIP2 were determined from the sequence data.


title: Two H3K23 histone methyltransferases, SET-32 and SET-21, function synergistically to promote germline nuclear RNAi, proper endogenous siRNA expression, and germline immortality in Caenorhabditis elegans [small RNA-seq]

city: Piscataway

first: Sam

middle: Guoping

last: Gu

state: New Jersey

address: 604 Allison Road

department: Molecular Biology and Biochemistry

institute: Rutgers University

summary: Nuclear RNAi in C. elegans induces a set of transgenerationally heritable marks of H3K9me3, H3K23me3, and H3K27me3 at the target genes. The function of H3K23me3 in the nuclear RNAi pathway is largely unknown due to the limited knowledge of H3K23 histone methyltransferase (HMT). In this study we identified SET-21 as a novel H3K23 HMT. By taking combined genetic, biochemical, and genomic approaches we found that SET-21 functions synergistically with a previously reported H3K23 HMT SET-32 to deposit H3K23me3 at the native targets of germline nuclear RNAi. We identified a subset of nuclear RNAi targets become transcriptionally activated in the set-21;set-32 double mutant. SET-21 and SET-32 are also required for a robust transgenerational gene silencing induced by exogenous dsRNA. The loss of SET-21 and SET-32 leads to a mortal germline phenotype, similar to mutations of the core nuclear RNAi component. Together, these results support a model in which H3K23 HMTs SET-21 and SET-32 ensure the robustness of germline nuclear RNAi and promotes the germline immortality at a high temperature.

overall_design: small RNA-seq libaries were prepared from from wild type and mutant adult hermaphrodite C. elegans strains.

middle: G

first: Julie

middle: N

last: Ni

first: Anna

last: Zhebrun


title: Two H3K23 histone methyltransferases, SET-32 and SET-21, function synergistically to promote germline nuclear RNAi, proper endogenous siRNA expression, and germline immortality in Caenorhabditis elegans [RNA-seq]

overall_design: RNA-seq libaries were prepared from from wild type and mutant adult hermaphrodite C. elegans strains.


title: Aalpi18529 regulates vitellogenesis of Aedes albopictus via the Gadd45a-mediated JNK-dependent nurse cell apoptosis pathway [small-RNA-seq]

first: Jinbao

address: No. 1023, Shatai South Road

department: Department of Pathogen Biology, School of Public Health

institute: Southern Medical University

summary: Aedes albopictus shows a rapid global expansion and dramatic vectorial capacity for various arboviruses. Mosquitoes display distinct sexual dimorphisms，only adult females consume blood meals to complete ovarian follicle development. Therefore, cyclic reproduction in female mosquitoes serves as a foundation for the transmission of numerous disease-causing pathogens. Aedes have an expansion of the piRNA biogenesis genes, indicated that piRNA may play multiple functional roles in mosquitoes. Although the antiviral function of piRNA pathway in mosquitoes has been extensively studied, the role of piRNAs in mosquito reproduction remain to be further understood. In the present study, we first profiled the characteristics of sex-biased piRNAs in adult Ae.albopictus. Then, we identified a female biased piRNA (Aalpi18529) in adult females, that was highly expressed in ovaries at blood feeding-dependent termination, and depended on PIWI5 and ago3 mediated biogenesis. Aalpi18529 overexpression suppressed ovarian development, and reduced fertility and fecundity in adult females post-bloodmeal. Furthermore, we demonstrated that Aalpi18529 can effectively repress its direct target, growth arrest and DNA-damage-inducible protein 45a (GADD45A), and eventually regulates ovarian development via the Gadd45a-mediated JNK-dependent nurse cell apoptosis pathway. Our study is the first to report an endogenous piRNA, which trigger silencing of an important protein-coding gene by posttranscriptional regulation in mosquitoes, expanding our current understanding of the important and multiple roles of piRNAs in biological processes in Ae. albopictus.

overall_design: To explore the sex-based expression profiles of piRNAs in Ae. albopictus, small RNA libraries from 2-day-old emerged Virgin females and male adults were constructed by using Illumina Solexa high-throughput sequencing technology.

first: Lu

last: Yang

first: Yonghui

last: Gao

first: Yulan

last: Chen


title: Aalpi18529 regulates vitellogenesis of Aedes albopictus via the Gadd45a-mediated JNK-dependent nurse cell apoptosis pathway {RNA-seq]

overall_design: To better understand the effect of Aalpi-18529 on ovarian development post blood meal, we performed genome-wide transcriptional profiling analysis of ovaries that were treated with Aalpi-18529 mimics.


title: Two H3K23 histone methyltransferases, SET-32 and SET-21, function synergistically to promote germline nuclear RNAi, proper endogenous siRNA expression, and germline immortality in Caenorhabditis elegans [ChIP-seq]

overall_design: ChIP-seq libaries were prepared from from wild type and mutant adult hermaphrodite C. elegans strains.


title: Identification of a depupylation regulator for an essential enzyme in Mycobacterium tuberculosis

city: New York

first: Shoshanna

middle: Celia

last: Kahne

state: NY

address: 430 E 29th Street, Lab 324 C-D

department: Microbiology

institute: New York University Grossman School of Medicine

summary: Mycobacterium tuberculosis can use a proteasome to degrade proteins when they are post-translationally modified with prokaryotic ubiquitin-like protein (Pup). While pupylation is reversible, mechanisms regulating depupylation have not been identified. Here, we identify a depupylation regulator, CoaX, a pseudo-pantothenate kinase. Pantothenate synthesis enzymes were more abundant in a ∆coaX mutant, including PanB, a substrate of the Pup-proteasome system. Media supplementation with pantothenate decreased PanB levels in a coaX and Pup-proteasome system-dependent manner. In vitro, CoaX accelerated depupylation of Pup~PanB, while addition of pantothenate inhibited this reaction. Collectively, we propose CoaX contributes to proteasomal degradation of PanB by modulating depupylation of Pup~PanB in response to pantothenate levels.

overall_design: RNA was extracted from quadruplicate biological cultures of H37Rv pMV strep ev (WT with an empty vector, MHD761), coaX::hyg pMV strep ev (∆coaX with an empty vector, MHD1845), and coaX::hyg pMV strep coaX (complemented strain, MHD1846) of Mycobacterium tuberculosis (10 OD equivalents).

first: K

last: Darwin

first: Alejandro

last: Pironti


title: RORα is required for expansion and memory maintenance of ILC1s via a lymphnode-liver axis [naïve]

city: Hefei

first: MING

last: CHENG

address: 96 Jinzhai Road, Hefei City, Anhui Province

institute: University Of Science And Technology Of China

summary: To Explore the transcription factors with high expression in memory ILC1s.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for memory ILC1s and naïve ILC1s

first: Ming

last: Cheng

first: Xianwei

last: Wang

first: Jiarui

last: Li

first: Hui

last: Peng

first: Zhigang

last: Tian


title: An archaeal chromatin protein condenses DNA through bridging-induced phase separation

city: Qingdao

first: yunfeng

state: Shandong

address: 72 Binhaidadao, Aoshanwei, Jimo

institute: Shandong University

summary: Phase separation serves an important role in the three-dimensional chromosome organization and remodeling in eukaryotes. Whether this process is involved in archaeal chromosome organization is unknown. Here we demonstrate that archaeal DNA condensing protein1 (aDCP1) from the hyperthermophilic crenarchaeon Sulfolobus islandicus is able to bridge DNA efficiently and form large protein-DNA condensates with a droplet- or gel-like morphology in vitro. Within the condensates, aDCP1 exhibits a fast dynamic while the DNA appears in a solid-like state. At the single-molecule level, aDCP1 efficiently compacts DNA through a three-step mechanism, which presumably entails the clustering of aDCP1 on the DNA and the subsequent fusion of the clusters. Deletion of the aDCP1 gene results in noticeable changes in chromosome conformation in S. islandicus, which are characterized by enhanced interactions between the A and B compartments and reduced interactions within the self-interacting domains as well as between domains in the same compartment. Taken together, our results indicate that aDCP1 is capable of inducing DNA bridging-induced phase separation and serves a role in chromosome organization in the organism.

overall_design: Chromatin immunoprecipitation (ChIP-Seq) was performed according to Takemata et al.(39) with slight modifications. Briefly, the cells were collected , cross-linked by adding 1% formaldehyde for 15 minutes, and quenched with a final concentration of 125 mM glycine. The cells were pelleted by centrifugation at 5,000 g for 10 minutes and washed with PBS. The cells were then resuspended in TBS-TT buffer (20 mM Tris, 150 mM NaCl, 0.1% Tween-20, 0.1% Triton X-100, pH 7.5) and fragmented by sonication until the DNA fragments were of 200-500 bp. After centrifugation (10,000 g for 15 minutes), a 100 µl aliquot of the DNA-containing supernatant was kept apart to be uses as an input control and the remaining sample was divided into two aliquots. One aliquot was incubated with anti-aCcr1 antibody-coated protein A beads (Cytiva) and the other was incubated with pre-immune serum-coated protein A beads, which served as a nonspecific binding control (Mock control). After incubation at 60 °C for 10 hrs, the samples were collected and the captured DNA was purified by using the DNA Cycle-Pure Kit (Omega) according to the manufacturer's instruction. The input samples were treated as above without the addition of antiserum and beads. The purified DNA was used for ChIP-Seq library preparation. The library was constructed by Novogene Corporation (Beijing, China). Subsequently, pair-end sequencing of sample was performed on Illumina platform (Illumina, CA, USA). Library quality was assessed on the Agilent Bioanalyzer 2100 system

first: Niannian

last: Ding

first: Yunfeng

first: Zhenfeng

last: Zhang

first: Yulong

last: Shen


title: Whole Cancer Cell Vaccine Protects Ovarian Functions in Tumor-bearing Mice through Downregulating CXCL10

first: 丛蓉

last: 李

address: 安徽医科大学东校区弘毅楼1202室

institute: Nanjing Medical university

summary: Your female patients with cancer will probably become sub-fertile or infertile even though some finally overcome the cancer through various therapies. Cancer immunotherapy is recently emerging as a promising novel therapy against cancer with high malignancy and lethality, but whether or not cancer immunotherapy affect female fertility is rarely known. In the present study, we employed MCA205 cells-xenografted B6 mice as model to investigate whether two popular immunotherapies, PD-1 monoclonal antibody (PD-1) and whole cancer cell vaccine (WCA), affect ovarian function. We found that MCA205 xenograft (M) conduced to decreased follicle numbers at each stage, decreased proliferation and increased apoptosis, decreased oocyte maturation rate and disrupted estrus cycle. WCA treatment significantly rescues the upper abnormalities while PD-1 didn't. Ovary RNA sequencing revealed that multiple DEGs (differentially-expressed genes) are involved in inflammation pathways. Next, cytokine microarray characterized CXCL10 with both biggest increment in M group and best rescue in WCA group. Next, CXCL10 antibody Immunoprecipitation in ovarian lysate and LC-MS baited the only receptor IL18R1. Next, we found that CXCL10 could at least impaired ovarian function through three pathways: it can increase ovarian inflammation through CXCL10→IL18R1→p-P65, induce ovarian fibrosis through CXCL10→IL18R1→p-JNK→COL1A1, and promote primordial follicle overactivation through CXCL10→IL18R1→p-AKT. Finally, we succeeded in rescuing ovarian function in M group through blocking the CXCL10→IL18R1 binding with CXCL10 antibody or a CXCL10-IL18R1 interface peptide. This study provided both mechanical evidences and translational strategies for whole cell vaccine in achieving dual functions, suppressing tumor progression meanwhile protecting ovarian function.

overall_design: To investigate the transcription change of ovary from tumor bearing mice which were treated with two kinds of cancer immunotherapy treatment , we operated whole cell vaccine or PD-1 monoclonal antibody on MCA205 cancer cell subcutaneous injected mice and collected ovaries to carry out RNA SEQ.

first: Cong-Rong

first: Shi-Ya

last: Xie

first: Shu-Ping


title: Inhibition of RNA splicing triggers CHMP7 nuclear entry impacting TDP-43 function and leading to onset of ALS (eCLIP).

city: La Jolla

first: Brian

last: Yee

state: CA

address: 9500 Gilman Dr

department: Health

institute: UCSD

summary: Amyotrophic lateral sclerosis (ALS) has been linked to nucleoporin loss in patient neurons. In the ESCRT-III pathway, CHMP7 protein accumulation in the nucleus triggers ALS by damaging nuclear pores and disrupting cellular transport. Genes controlling CHMP7's nuclear presence remain unidentified. Our Craft-ID analysis revealed that RNA binding proteins involved in assembling small nuclear ribonucleoprotein particles control CHMP7 nuclear localization. This regulation ultimately impacts splicing processes and can lead to pore injury. Using IP-MS, we discovered that CHMP7 interacts with the survival of motor neuron (SMN) complex (SmD1-3, Gemins), mediated by U1 snRNP and direct interactions between CHMP7. Reducing expression of the core SMN complex component SmD1 was observed in ALS iPSC- Motor Neurons (MNs), affecting CHMP7 localization. However, restoring its levels can rescue the cytoplasmic localization of CHMP7 in sALS iPSC-MNs. Our discoveries hint at an early ALS pathway involving SMN core complex dysregulation, influencing disease onset.

overall_design: eCLIP of CHMP7 in motor neurons, in duplicate

first: Norah

last: Al-Azzm

first: Lena

last: Street

first: Jenny

last: To

first: Assael

last: Madrigal

first: Benjamin

last: Lee

first: Wenhao

last: Jin

first: Dylan

last: Lam

first: Vaishali

last: Gautam

first: Jack

last: Naritomi

first: Willard

last: Ford

first: Chloe

last: Nguyen

first: Gene

middle: W

last: Yeo


title: Inhibition of RNA splicing triggers CHMP7 nuclear entry impacting TDP-43 function and leading to onset of ALS (RNA-Seq).

overall_design: RNA seq of HeLa CHMP7 Helix5 and control


title: RORα is required for expansion and memory maintenance of ILC1s via a lymphnode-liver axis [Smart-seq]

first: MING

last: CHENG

summary: To explore the downstream pathways of Rora regulation of memory ILC1 during memory maintenance period.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for oil-treated ILC1s and tamoxifen-treated ILC1s

first: Ming

last: Cheng

first: Xianwei

last: Wang

first: Jiarui

last: Li

first: Hui

last: Peng

first: Rui

last: Sun

first: Zhigang

last: Tian


title: RORα is required for expansion and memory maintenance of ILC1s via a lymphnode-liver axis

summary: To explore the downstream pathways of Rora regulation of memory ILC1 during memory formation period.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for wt ILC1s and cKO ILC1s


title: Sex differences in brain transcriptomes of juvenile Cynomolgus macaques

city: Gainesville

first: Alberto

last: Riva

state: FL

address: 2033 Mowry Rd

department: ICBR

institute: University of Florida

summary: Behavioral, social, and physical characteristics are posited to distinguish the sexes, yet research on transcription-level sexual differences in the brain is limited. Here, we investigated sexually divergent brain transcriptomics in prepubertal cynomolgus macaques, a commonly used surrogate species to humans. A transcriptomic profile using RNA sequencing was generated for the temporal lobe, ventral midbrain, and cerebellum of 3 female and 3 male cynomolgus macaques previously treated with an Adeno-associated virus vector mix. Statistical analyses to determine differentially expressed protein-coding genes in all three lobes were conducted using DeSeq2 with a false discovery rate corrected P value of .05. We identified target genes in the temporal lobe, ventral midbrain, and cerebellum with functions in translation, immunity, behavior, and neurological disorders that exhibited statistically significant sexually divergent expression. We provide potential mechanistic insights to the epidemiological differences observed between the sexes with regards to mental health and infectious diseases, such as COVID19. Our results provide pre-pubertal information on sexual differences in non-human primate brain transcriptomics and may provide insight to health disparities between the biological sexes in humans.

overall_design: Total RNA sequencing was performed to compare gene expression profiles in the cerebellum, midbrain, and temporal lobe of 6 cynomolgous macaques (3 male and 3 female).

first: Nadia

last: Kabbej

first: Frederick

last: Ashby

first: Paul

last: Gamlin

first: Ronald

last: Mandel

first: Aishwarya

last: Kunta

first: Courtney

last: Rouse

first: Coy

last: Heldermon


title: BCR and TLR-induced IRF1 promotes T cell independent B cell responses by supporting Notch2 dependent Marginal Zone B cell development

first: Jessica

last: Peel

address: 1230 York Ave

institute: The Rockefeller University

summary: The prototypic interferon (IFN)-inducible transcription factor, IRF1, not only controls expression of IFN genes but also regulates T cell and macrophage fate specification and function. We show that IRF1 expression in B cells is required for marginal zone B (MZB) cell development and T cell independent antibody responses [80% B6.129S2-Ighmtm1Cgn/J (MT) + B6.129S2-Irf1tm1Mak/J (Irf1-/-), B-Irf1-/- chimeras]. While IFNs can induce IRF1 expression in MZB precursors, IFN signaling is not required for MZB cell development [C57BL/6J (B6), B6.129S7-Ifngr1tm1Agt/J (Ifngr1-/-), B6(Cg)-Ifnar1tm1Agt/J (Ifnar1-/-)].  Instead, BCR and TLR signals, which influence MZB commitment, promote IRF1 expression and nuclear translocation in MZB cell precursors. In turn, IRF1 is required for Notch-dependent gene expression in transitional B cells and development of the MZB cell compartment.  Thus, T independent MZB-driven antibody responses are regulated by BCR and TLR signals that initiate IRF1-dependent Notch programming of transitional B cells and subsequent commitment of these cells to the MZB lineage.

overall_design: RNAseq was performed for the following (in triplicate for each sample type): 1. C57BL/6 mice 2. IRF1 KO uninfected mice; Transitional 1, Transitional 2, Follicular B, Marginal Zone B cells were isolated by FACS

first: Frances

middle: E

last: Lund

first: Chris

last: Scharer

first: Alexander

last: Rosenberg