title: Identification of an anergic BND cell-derived activated B cell population (BND2) in young-onset type 1 diabetes

city: Aurora

first: Kristen

last: Wells

address: 1775 Aurora Ct,

department: Barbara Davis Center

institute: University of Colorado Anschutz

summary: Recent evidence suggests a role for B cells in the pathogenesis of type 1 diabetes (T1D), particularly in individuals who develop T1D at a younger age and demonstrate rapid progression and beta cell loss. However, little is known regarding the specificity, phenotype, and function of B cells in young-onset T1D. We performed a cross-sectional analysis comparing insulin-reactive to tetanus-reactive B cells in the blood of T1D and controls using mass cytometry. Unsupervised clustering revealed the existence of a highly activated B cell subset, we term BND2, that falls within the previously defined anergic BND subset. We found a specific increase in the frequency of insulin-reactive BND2 cells in the blood of young-onset T1D donors, which was further enriched in the pancreatic lymph nodes of T1D donors. The frequency of insulin-binding BND2 cells correlated with anti-insulin autoantibody levels in T1D subjects. We demonstrate BND2 cells phenotypically and functionally are pre-plasma cells and can likely act as antigen-presenting cells to T cells. These findings suggest that activation of insulin-reactive anergic B cells may play a role in the rapid progression of young-onset T1D and warrants further investigation in other autoimmune conditions.

overall_design:
Single cell RNA and CITE seq on B cells isolated from PBMCs from a non-diabetic donor. CITE seq antibodies included IgM, IgD, CD21, CD27 and CXCR5. All scripts for analysis are available on github https://github.com/CUAnschutzBDC/Smith_210825_b_cell_analysis
Filtered output files from cellranger are provided as supplementary files as well as a metadata file that contains cell statistics (nFeature, nCount, percent.mt), cell type labels, final clusters that align with the paper, and UMAP coordinates that also align with the paper. A description of how to analyze this data is on the github linked above.

first: Zachary

middle: C

last: Stensland

first: Mia

last: Smith


title: Modeling and therapeutic targeting of inflammation-induced hepatic insulin resistance using human iPSC-derived hepatocytes and macrophages

status: Public on May 09 2023

city: San Francisco

first: Marko

last: Groeger

state: CA

address: 35 Medical Center Way

country: USA

institute: University of California San Francisco

type: Expression profiling by high throughput sequencing

summary: RNAseq was used to identify inflammation-related pathways activated in iPSC-derived hepatocytes by co-culture with iPSC-derived macrophages, thereby allowing for making inferences about cytokine mediators.

overall_design: Comparison of gene expression profile by RNAseq of iPSC-derived hepatocytes (n=3 per condition) after 24-hour co-culture with activated (M1) iPSC-derived macrophages or non-activated (M0) iPSC-derived macrophages or after mon-culture (C).

first: Cindy

last: Pino

first: Edward

last: Hsiao

first: Holger

last: Willenbring


title: Co-option of endogenous retroviruses through genetic escape from TRIM28 repression

status: Public on May 10 2023

city: London

first: Helen

middle: M.

last: Rowe

state: --- Select a state ---

address: 4 Newark St, Whitechapel

department: Blizard Institute, Barts and The London School of Medicine and Dentistry

country: United Kingdom

institute: Queen Mary University of London

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Endogenous retroviruses (ERVs) have rewired host gene networks. To explore the origins of co-option, we employed an active murine ERV, IAPEz, and an embryonic stem cell (ESC) to neural progenitor cell (NPC) differentiation model. Epigenetic silencing via TRIM28 maps to a 190bp sequence encoding the IAP signal peptide, which confers retrotransposition activity. A subset of ‘escapee’ IAPs (~15%) exhibit significant genetic divergence from this sequence. Canonical repressed IAPs succumb to a previously undocumented demarcation by H3K9me3 and H3K27me3 in NPCs. Escapee IAPs, in contrast, evade epigenetic repression in both cell types, resulting in their transcriptional derepression, particularly in NPCs. We validate the enhancer function of a 47bp sequence within the U3 region of the LTR and show that escapee IAPs convey an activating effect on nearby neural genes. In sum, co-opted ERVs stem from genetic escapees that have lost vital sequences required for both TRIM28 restriction and autonomous retrotransposition.

overall_design: Identification of how sequence divergence from a functional ER-targeting signal, which conferred the active murine endogenous retrovirus IAPs with the ability to retrotranspose, can lead to escape from epigenetic repression and paves the way to co-option of these elements as enhancers.

first: Rocio

last: Enriquez-Gasca

first: Poppy

middle: A

last: Gould

first: Hale

last: Tunbak

first: Lucia

last: Conde

first: Javier

last: Herrero

first: Alexandra

last: Chittka

first: Christine

middle: R

last: Beck

first: Robert

last: Gifford

middle: M


title: Microarray gene expression profiles from female ER-positive HER2-normal breast cancer patients

city: Copenhagen Ø

first: Maria

last: Rossing

address: Blegdamsvej 9

department: Center for Genomic Medicine

country: Denmark

institute: Rigshospitalet

summary: Diagnostic samples from female breast cancer patients with ER-positive and HER2-normal tumors selected for neoadjuvant chemotehrapy.

overall_design: Tumor specimens from female breast cancer patients were collected as part of routine diagnostics. Expression patterns were analyzed using the Affymetrix HG-U133 Plus 2.0 platform as well as IHC and FISH.

first: Christina

middle: B

last: Pedersen


title: Dysregulation of amino acid metabolism upon rapid depletion of cap-binding protein eIF4E

city: Berkeley

first: Paige

middle: D

last: Diamond

address: Barker Hall

department: MCB

institute: University of California - Berkeley

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

overall_design: Refer to individual Series


title: Ccr2+ monocyte-derived macrophages influence trajectories of acquired therapy resistance in Braf-mutant melanoma

city: ithaca

first: Luye

last: An

address: 618 tower road

department: biomedical sciences

institute: cornell university

summary: Therapies targeting oncogene addiction have had a tremendous impact on tumor growth and patient outcome, but drug resistance continues to be problematic. One approach to deal with the challenge of resistance entails extending anti-cancer treatments beyond targeting cancer cells by additionally altering the tumor microenvironment. Understanding how the tumor microenvironment contributes to the evolution of diverse resistance pathways could aid in the design of sequential treatments that can elicit and take advantage of a predictable resistance trajectory. Tumor associated macrophages are often the most abundant immune cell found in tumors. Here, we used clinically relevant in vivo Braf-mutant melanoma models with fluorescent markers to track the stage-specific changes in macrophages under targeted therapy with Braf/Mek inhibitors and assessed the dynamic evolution of the macrophage population generated by therapy pressure-induced stress. During the onset of a drug-tolerant persister state, Ccr2+ monocyte-derived macrophage infiltration rose, suggesting that macrophage influx at this point could facilitate the onset of stable drug resistance after several weeks of treatment. Comparison of melanomas that develop in a Ccr2-proficient or deficient microenvironment demonstrated that lack of melanoma infiltrating Ccr2+ macrophages delayed onset of resistance and shifted melanoma cell evolution towards unstable resistance. Unstable resistance was characterized by sensitivity to targeted therapy when factors from the microenvironment were lost. Importantly, this phenotype was reversed by co-culturing melanoma cells with Ccr2+ macrophages. Overall, this study demonstrates that the development of resistance may be directed by altering the tumor microenvironment to improve treatment timing and the probability of relapse.

overall_design: Comparative gene expression profiling analysis of isolated macrophages (F4/80+) from resistant tumor tissue in control and Ccr2 KO mice using bulk RNA-seq data

first: Dahihm

last: Kim

first: Jiwon

last: Moon

first: Viviana

middle: I

last: Maymi

first: Alexander

last: McGurk

first: Brian

last: Rudd

first: Deborah

last: Fowell

first: Andrew

last: White


title: Parallels and contrasts between the cnidarian and bilaterian maternal-to-zygotic transition are revealed in Hydractinia embryos

city: Pittsburgh

first: Miler

middle: T

last: Lee

state: PA

address: 4249 Fifth Ave

department: Biological Sciences

institute: University of Pittsburgh

summary: Embryogenesis requires coordinated gene regulatory activities early on that establish the trajectory of subsequent development, during a period called the maternal-to-zygotic transition (MZT). The MZT comprises transcriptional activation of the embryonic genome and post-transcriptional regulation of egg-inherited maternal mRNA. Investigation into the MZT in animals has focused almost exclusively on bilaterians, which include all classical models such as flies, worms, sea urchin, and vertebrates, thus limiting our capacity to understand the gene regulatory paradigms uniting the MZT across all animals. Here, we elucidate the MZT of a non-bilaterian, the cnidarian Hydractinia symbiolongicarpus. Using parallel poly(A)-selected and non poly(A)-dependent RNA-seq approaches, we find that the Hydractinia MZT is composed of regulatory activities analogous to many bilaterians, including cytoplasmic readenylation of maternally contributed mRNA, delayed genome activation, and separate phases of maternal mRNA deadenylation and degradation that likely depend on both maternally and zygotically encoded clearance factors, including microRNAs. But we also observe massive upregulation of histone genes and an expanded repertoire of predicted H4K20 methyltransferases, aspects thus far unique to the Hydractinia MZT and potentially underlying a novel mode of early embryonic chromatin regulation. Thus, similar regulatory strategies with taxon-specific elaboration underlie the MZT in both bilaterian and non-bilaterian embryos, providing insight into how an essential developmental transition may have arisen in ancestral animals.

overall_design: RNA-seq with poly(A)+ selection or rRNA-depletion was performed on a timecourse of development in Hydractinia symbiolongicarpus. Replicates are named A-G according to collection group


title: Spatiotemporal resolution of germinal center Tfh cell differentiation and divergence from central memory CD4+ T cell fate

city: Birmingham

first: Fangming

last: Zhu

state: AL

address: 845 19th Street South

department: Microbiology

institute: University of Alabama at Birmingham

summary: Transcriptome analysis of shPlekho1 and shCD19 GC-Tfh cells responding to influenza infection.

overall_design: Purified CD45.1+CD45.2+ and CD45.2+ OT-II T cells were activated with 0.5 μg/ml anti-CD3 and 1 μg/ml anti-CD28 antibodies in plates pre-coated with 20 μg/ml goat anti-hamster IgG under Th0 condition. The activated cells were transduced at 18 and 48 hours with shCD19 and shPlekho1 virus-containing media, respectively. After the second transduction, cells were expanded for 2 days in complete T cell medium containing 20 U/ml recombinant human IL-2. 2 × 104 retrovirally infected shPlekho1 OT-II cells were co-transferred with shCD19 control OT-II cells intravenously into CD45.1+ SMARTA recipient mice followed by intranasal infection with PR8-OVA 2 days after cell transfer. The donor OT-II cells from the medLN of the recipient mice were analyzed by RNA-seq at day 14 p.i.

first: Ryan

middle: J

last: McMonigle

first: Diego

middle: O

last: Sialer

first: Braxton

last: Greer

first: Edahi

last: Gonzalez Avalos

first: Hui

last: Hu


title: Class 3 PI3K participates in nuclear gene transcription and co-activates the circadian clock to promote de novo purine synthesis [RNA-seq]

city: Paris

first: Ganna

last: Panasyuk

address: 156-160 Rue Vaugirard, Faculté de Médecine Paris Descartes,, 4th Floor - Lab-408/Office-404,, 4th Floor - Lab-408/Office-404,

country: France

institute: Inserm U1151/CNRS UMR 8253, Necker Enfants Malades Institute (INEM)

summary: Metabolic demands fluctuate rhythmically and rely on coordination between the circadian clock and nutrient sensing signaling pathways, yet mechanisms of their interaction remain not fully understood. Here, surprisingly, we find that class 3 Phosphatidylinositol-3-kinase (PI3K), known best for its essential role as a lipid kinase in endocytosis and lysosomal degradation by autophagy, has an overlooked nuclear function in participating to gene transcription as a co-activator of heterodimeric transcription factor and circadian driver BMAL1-CLOCK. Canonical pro-catabolic functions of class 3 PI3K in trafficking rely on the indispensable complex between lipid kinase Vps34 and regulatory subunit Vps15. We find that although both subunits of class 3 PI3K interact with RNA Pol2 and co-localize with active transcription sites, exclusive loss of Vps15 blunted BMAL1-CLOCK transcriptional activity. Thus, we established non-redundancy between nuclear Vps34 and Vps15 reflected in a persistent nuclear pool of Vps15 in Vps34-depleted cells and ability of Vps15 to co-activate BMAL1-CLOCK independently of its complex with Vps34. In physiology, we find Vps15 is required for metabolic rhythmicity in liver and, unexpectedly, it promoted pro-anabolic de novo purine nucleotide synthesis. We show Vps15 activated transcription of Ppat, a key enzyme for production of inosine monophosphate, a central metabolic intermediate for purine synthesis. Finally, we demonstrate that in fasting, which represses clock transcriptional activity, BMAL1 recruitment to chromatin is unmodified but it coincides with depletion of Vps15 on the promoters of its targets, Nr1d1 and Ppat. Our findings suggest that nuclear pool of class 3 PI3K Vps15 subunit could couple transcriptional activity of the circadian clock for temporal regulation of energy homeostasis and it opens novel avenues for establishing the complexity for nuclear class 3 PI3K signaling.

overall_design: RNA-seq of  WT and Vps15 liver KO at ZT6.

first: Nathaniel

middle: F

last: Henneman

first: Chantal

last: Alkhoury

first: Volodymyr

last: Petrenko

first: Yui

last: Shibayama

first: Arianna

last: Segaloni

first: Alexis

last: Gadault

first: Ivan

last: Nemazanyy

first: Edouard

last: Le Guillou

first: Amare

last: Wolide

first: Konstantina

last: Antoniadou

first: Xin

last: Tong

first: Teruya

last: Tamaru

first: Takeaki

last: Ozawa

first: Muriel

last: Girard

first: Karim

last: Hnia

first: Dominic

last: Lutter

first: Charna

last: Dibner


title: Sperm chromatin structure and reproductive fitness are altered by substitution of a single amino acid in mouse Protamine 1 [scRNA-Seq]

city: Ann Arbor

first: Qianyi

last: Ma

state: MI

address: 1141 E Catherine St

department: Human Genetics

institute: University of Michigan

summary: Conventional dogma presumes that protamine-mediated DNA compaction in sperm is achieved by electrostatic interactions between DNA and the arginine-rich core of protamines. However, phylogenetic analysis reveals several non-arginine residues conserved within, but not across species. The significance of these residues or post-translational modifications are poorly understood. Here, we investigated the functional role of K49, a rodent-specific lysine residue in mouse protamine 1 (P1) that is acetylated early in spermiogenesis and retained in sperm. In vivo, alanine substitution (P1 K49A) results in loss of programmatic histone retention, decreased sperm motility, decreased male fertility, and in zygotes, premature P1 removal from paternal chromatin, altered DNA replication, and embryonic arrest. In vitro, P1 K49A decreases protamine-DNA binding and alters DNA compaction/decompaction kinetics. Hence, a single amino acid substitution outside the P1 arginine core is sufficient to profoundly alter protein function and developmental outcomes, suggesting that protamine non-arginine residues are essential for reproductive fitness.

overall_design: Two-cell embryos were collected from mice between 26 and 28 hours post fertilization and each embryo was immediately placed into a single well of a plexWell Rapid Single Cell plate, which was then frozen until sequencing.

first: Saher

middle: S

last: Hammoud

first: Jun

middle: Z

last: Li


title: Tumor-intrinsic ST3GAL3 Activity Blunts Anti-tumor Immune and Drives Immunotherapy Resistance in Ovarian Cancer

city: Shanghai

first: Haiou

last: Liu

address: No.128, Shengyang road, Yangpu distinct, Shanghai, P. R. China.

country: China

institute: Fudan University

summary: ST3GAL3 is a α2,3-sialyltransferase and involves in α2,3 sialylation in tumors; however, its role in the tumorigenic capacity of malignant cells and anti-tumor immune remains largely unexplored. Here, HM1 cells were stably transfected using lentivirus expressing a shRNA targeting ST3GAL3 (HM1-shST3GAL3) or scramble shRNA as control (HM1-shControl). Then, HM1-shControl or HM1-shST3GAL3 cells were injected into the right flank of B6C3F1 mice using a 27 g needle. ST3GAL3 knockdown significantly compromised tumor growth at a immune-dependant manner. Mechanistically, ST3GAL3 knockdown enhances pro-inflammatory phenotype of macrophages, leading ultimately to activation and recruitmentof CD8+ T cells. Moreover, we identified ST3GAL3 as a potential novel biomarker for bevacizumab or immune checkpoint blockade (ICB) intervention. ST3GAL3 knockdown in tumors also be useful to improve cancer immunotherapies.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for shControl tumor tissues (shControl tumors, n = 3) and  shST3GAL3 tumor tissues (shST3GAL3 tumors, n = 3) of early tumor model.

first: Kankan

last: Cao


title: Genome-wide CRISPR screen to identify CD58 regulators

city: New York

first: Patricia

last: Ho

state: NY

address: 650 West 168th Street

country: Canada

institute: Columbia University

summary: The cell autonomous balance of immune-inhibitory and -stimulatory signals is a critical yet poorly understood process in cancer immune evasion. Using patient-derived co-culture models and humanized mouse models, we show that an intact CD58:CD2 interaction is necessary for anti-tumor immunity. Defects in this axis lead to multi-faceted immune evasion through impaired CD2-dependent T cell polyfunctionality, T cell exclusion, impaired intra-tumoral proliferation, and concurrent protein stabilization of PD-L1. We performed genome-scale CRISPR-Cas9 and CD58 co-immunoprecipitation mass spectrometry screens identifying CMTM6 as a key stabilizer of CD58, and show that CMTM6 is required for concurrent upregulation of PD-L1 in CD58 loss. Single-cell RNA-seq analysis of patient melanoma samples demonstrates that most TILs maintain strong CD2 expression, thus providing an opportunity to mobilize a so far therapeutically untapped pool of TILs for anti-tumor immunity. We identify two potential therapeutic avenues, including rescued activation of human CD2-expressing TILs using recombinant CD58 protein, and targeted disruption of PD-L1/CMTM6 interactions. Our work identifies an underappreciated yet critical axis at the nexus of cancer immunity and evasion, uncovers a fundamental mechanism of co-inhibitory and -stimulatory signal balancing, and provides new approaches to improving cancer immunotherapies.

overall_design: A375 Cas9-expressing cells were transduced with a whole genome targeting sgRNA library and cultured for 10 days. Cells were sorted by FACS for CD58-low expressing cells alongside a CD58-mid expressing control population. This experiment was performed in two biological replicates, with two technical replicates each. sgRNA sequences from each population were amplified and sequenced. sgRNA sequences were also captured from the original sgRNA library virus as well from Day 6 and 10 after transduction from each biological replicate to capture sgRNA dropout.

first: Chris

last: Frangieh

first: Johannes

last: Melms

first: Benjamin

last: Izar


title: Class 3 PI3K coactivates the circadian clock to promote rhythmic de novo purine synthesis

first: Ganna

last: Panasyuk


title: Sperm chromatin structure and reproductive fitness are altered by substitution of a single amino acid in mouse Protamine 1 [MNase-Seq]

type: Methylation profiling by high throughput sequencing

overall_design: MNase-Seq was performed on four wild type C57BL6 mice biological replicates and two mutant K49A mice biological replicates.


city: Wuhan

first: Li-quan

last: Zhou

state: Hubei

address: 13 Hangkong road

department: Tongji Medical College

institute: Huazhong University of Science and Technology

summary: We demonstrat that Mdivi1 treatment impacts early embryo transcriptome.

overall_design: We treated embryos with Mdivi-1 to inhibit DRP1 activity and examined embryo transcriptome.

first: Shi-meng

last: Guo

first: Yi-ran

last: Zhang

first: Ying

last: Yin

first: Ximiao

last: He


title: Sperm chromatin structure and reproductive fitness are altered by substitution of a single amino acid in mouse Protamine 1


title: PIK3CA mutation in endometrial epithelia promotes Viperin-dependent inflammatory response to insulin 2

city: Grand Rapids

first: Ronald

middle: L

last: Chandler

state: Michigan

address: 400 Monroe Ave NW

institute: Michigan State University

summary: Endometrial epithelia (EE) are known to harbor cancer driver mutations in the absence of any pathologies, including mutations in PIK3CA. Insulin plays an important role in regulating uterine metabolism during pregnancy, and hyperinsulinemia is associated with conditions impacting fertility. Hyperinsulinemia also promotes cancer, but the direct action of insulin on mutated EE is unknown. Here, we treated EE cells carrying the PIK3CAH1047R oncogene with insulin and examined differential transcriptomes by RNA-seq. While cells responded to insulin, the magnitude of differential gene expression (DGE) was 9 times greater in PIK3CAH1047R cells, representing a synergistic effect between insulin signaling and PIK3CAH1047R. Interferon signaling and the unfolded protein response (UPR) were enriched pathways among affected genes. Insulin treatment in normal cells activated normal endoplasmic reticulum stress (ERS) response programs, while PIK3CAH1047R cells activated programs necessary to avoid ERS-induced apoptosis. PIK3CAH1047R expression alone resulted in fewer differentially expressed genes, however, top among these was overexpression of Viperin (RSAD2), which is involved in viral response and upregulated in the endometrium during early pregnancy. The transcriptional changes induced by insulin in PIK3CAH1047R cells were rescued by knockdown of Viperin, while overexpression alone was insufficient to induce a DGE response to insulin, suggesting that Viperin is necessary but not sufficient for the synergistic effect of PIK3CAH1047R and insulin treatment. We identified interferon signaling, viral response, and protein targeting pathways that are induced by insulin but dependent on Viperin in PIK3CAH1047R mutant cells. These results suggest a differential response to insulin signaling in mutated EE.

overall_design: 12Z endometriotic epithelial cells transfected with a combinaton of siRSAD2 or non-targeting siRNA control and pBabe-PIK3CA*H1047R or empty vector, and were treated either with 100 nM insulin or vehicle. Cells were assayed 72 hours post-transfection via RNA-seq.

first: Mike

last: Wilson


title: PIK3CA mutation in endometrial epithelia promotes Viperin-dependent inflammatory response to insulin 3

overall_design: 12Z endometriotic epithelial cells transfected with wither pCAG Viperin plasmid or pCAG empty vector, and were treated either with 100 nM insulin or vehicle. Cells were assayed 24 hours post-insulin or vehicle treatment for RNA-seq.

first: Shannon

last: Harkins


title: Site-specific development and progressive maturation of human tissue resident memory T cells over infancy and childhood

first: Thomas

last: Connors

state: New York

address: 3595 Broadway CHN 10-24

institute: Columbia University Medical Center

summary: Infancy and childhood are critical life stages for the generation of immune memory for protection against pathogens. This process has been challenging to elucidate in humans as most memory T cells reside in tissues. Here, we define localization, and developmental pathways of memory T cells in mucosal sites, lymphoid tissues, and blood from pediatric donors aged 0-10 years. We reveal that memory T cells preferentially localize in mucosal sites during infancy and accumulate more rapidly over childhood compared to blood and lymphoid organs. Mucosal memory T cells in infancy exhibit stem-like transcriptional profiles, but progressively adopt signatures of mature tissue resident memory T cells over childhood. Our results show that exposures during the formative years establish the landscape of immunity through tissue-specific generation and adaptation of memory T cells

overall_design: Resident memory T cells over infancy and childhood

first: Rei

last: Matsumoto

first: Shivali

last: Verma

first: Peter

last: Szabo

first: Rebecca

last: Guyer

first: Joshua

last: Gray

first: Zicheng

last: Wang

first: Puspa

last: Thapa

first: Pranay

last: Dogra

first: Maya

last: Poon

first: Ksenia

last: Rybkina

first: Marissa

last: Bradley

first: Emma

last: Idzikowski

first: James

last: McNichols

first: Masaru

last: Kubota

first: Kalpana

last: Pethe

first: Yufeng

last: Shen

first: Maigan

last: Brusko

first: Todd

first: Andrew

last: Yates

last: Sims

first: Mark

last: Atkinson

first: Donna

last: Farber


title: Convergence of YAP/TAZ, TEAD and P63 activity directs premalignant lung gene expression  [ChIP-seq]

city: BOSTON

first: Boting

last: Ning

state: Massachusetts

address: 72 E Concord St

institute: BOSTON UNIVERSITY

summary: Bronchial premalignant lesions (PMLs) are composed of expanding bronchial basal cells that can progress to lung squamous cell carcinoma (LUSC) by evading immune responses. Despite ongoing efforts that have mapped gene expression and cell diversity across bronchial PML pathologies, signaling and transcriptional events driving malignancy are poorly understood. Evidence has suggested key roles for the Hippo pathway effectors YAP and TAZ and associated TEAD and TP63 transcription factor families in bronchial basal cell biology and LUSC. In this study we mapped target genes regulated by these factors by combined RNA-seq and ChIP-seq in primary human bronchial epithelial cells, revealing a converged transcriptional network that is strongly associated with bronchial PML progression. Our observations suggest that YAP/TAZ-TEAD-TP63 associate to cooperatively promote basal cell proliferation and repress signals associated with interferon responses and immune cell communication. Directly repressed targets include MHC Class II factors expressed in bronchial epithelium that correlate with adaptive immune Th1 cell activities known to contribute to the immunosuppressive microenvironment in lung cancers. Our findings provide a molecular mechanism for gene regulation in progressive PMLs that contributes to immune evasion, offering potential new avenues for lung cancer interception.

overall_design: HBECs for ChIP were cultured in Pneumocult EX Plus media and cross-linked in 1mM EGS in PBS for 30min followed by a 1% formaldehyde treatment for 10 min. Fixation was subsequently neutralized with 0.125M glycine in PBS. Harvested chromatin was isolated as single samples from each patient line, sonicated using the Bioruptor UCD- 200 and the incubated with the following antibodies at 4 °rnight: Rabbit anti-Yap (Abcam,  Cat# ab52771, 3ug), Rabbit anti-TEAD (AvivaSysBio, Cat# ARP38276, 1ug), and Mouse anti-P63 (Biocare # CM163, 5ug). Immunoprecipitated complexes were collected by Protein A/G Magnetic beads (Pierce, 8802). Samples were washed with low salt buffer (20mM Tris, 140mM NaCl, 1mM EDTA, 0.1% NaDeoxycholate, 0.1% SDS, 1% Triton X-100), followed by a high salt buffer (20mM Tris, 500mM NaCl, 1mM EDTA, 0.5% NaDeoxycholate, 1% Triton X-100), and a LiCl buffer (20mM Tris, 1mM EDTA, 0.1% NaDeoxycholate, 1% Triton X-100, 250mM LiCl). Chromatin was de-crosslinked overnight at 65C and purified using the Qiaquick PCR purification kit (Qiagen, 28104). For ChIP-seq, the purified DNA was ligated to specific adaptors and sequenced using DNB-seq, performed by BGI, to a depth of 40 million reads.

first: Julia

last: Berthet

first: Varelas

last: Xaralabos


title: Convergence of YAP/TAZ, TEAD and P63 activity directs premalignant lung gene expression [RNA-seq]

overall_design: For generating the YAP/TAZ, TEAD and TP63 regulated gene expression signature in human airway cells, HBECs were cultured in Pneumacult EX plus and transfected with control, pan-TEAD targeting, TP63 targeting and LATS1/2 targeting siRNAs and RNA was extracted for quality assessment and library prep for RNA-sequencing. 3 unique siRNA control sequences were used, and all siRNA transfected samples were collected in triplicate, 48hours after transfection. RNA quality for all samples was assessed by BioAnalyzer before proceeding with library preparation for sequencing. Sequencing libraries were prepared from total RNA samples using Illumina TruSeq RNA Sample Preparation Kit v2. The libraries from individual samples were pooled sequencing. HBEC samples were sequenced on the Illumina HiSeq 2500 platform to generate single end 50bp reads.

last: Tilston-Lunel


title: Tumor-intrinsic ST3GAL3 Activity Blunts Anti-tumor Immune and Drives Immunotherapy Resistance

first: Haiou

last: Liu

overall_design: Comparative gene expression profiling analysis of RNA-seq data for HM1 cells (HM1-shControl, n = 3) and its ST3GAL3 knockdown derivatives (HM1-shST3GAL3, n = 3). Comparative gene expression profiling analysis of RNA-seq data for shControl tumor tissues (shControl tumors, n = 3) and  shST3GAL3 tumor tissues (shST3GAL3 tumors, n = 3).

first: Kankan

last: Cao


title: PIK3CA mutation in endometrial epithelia promotes Viperin-dependent inflammatory response to insulin 1

first: Ronald

last: Chandler

overall_design: 12Z endometriotic epithelial cells transfected with non-targeting siRNA and pBabe-PIK3CA*H1047R or empty vector, and were treated either with 100 nM insulin or vehicle. Cells were assayed 72 hours post-transfection via RNA-seq.

first: Mike

last: Wilson


title: Co-transcriptional genome surveillance by HUSH is coupled to termination (PRO-Seq)

city: Stanford

first: Tomek

last: Swigut

address: 265 Campus Dr

department: Chemical and Systems Biology

institute: Stanford University

summary: The HUSH complex recognizes and silences foreign DNA such as viruses, transposons, and transgenes without prior exposure to its targets. Recent studies revealed that HUSH recruitment is dependent on transcription, but a unifying molecular mechanism by which HUSH identifies its targets as ‘non-self’ DNA and selects them for silencing remains unknown.  Harnessing a de novo silencing event in ESCs, we uncover distinct modes of HUSH function – one involves co-transcriptional surveillance in the absence of silencing, while the other is associated with H3K9me3 deposition and repression. We demonstrate that HUSH travels with elongating RNAPII, interacts with the transcriptional termination machinery, and accumulates on chromatin in a manner dependent on the termination factor WDR82. We further show that perturbation of endogenous termination signals triggers the switch from HUSH surveillance to silencing. Together, our results uncover an RNAi-independent, co-transcriptional gene silencing mechanism in mammals that senses aberrant termination to distinguish self from non-self.

overall_design: Precision nuclear run-on sequencing (PRO-seq) in  R1 mouse embryonic stem cells

last: Spencley

first: Shiran

last: Bar

first: Ryan

last: Flynn

first: Cameron

last: Lee

first: Liang-Fu

last: Chen

first: Michael

last: Bassik

first: Joanna

last: Wysocka


title: Co-transcriptional genome surveillance by HUSH is coupled to termination (CLIP-Seq)

overall_design: MPP8 irCLIP-seq in R1 mouse embryonic stem cells

first: Andrew


title: Co-transcriptional genome surveillance by HUSH is coupled to termination

overall_design: Refer to individual Series


title: Co-transcriptional genome surveillance by HUSH is coupled to termination (RNA-Seq)

overall_design: rRNA-depleted RNA-seq in R1 mouse embryonic stem cells


title: Intrinsically determined turnover underlies broad heterogeneity in plasma-cell lifespan

city: Melbourne

first: Zhoujie

last: Ding

state: VIC

address: 89 Commercial Road

country: Australia

institute: Monash University


title: Transcriptomic profiling of induced steatosis in human precision-cut liver slices

city: Biberach a.d. Riss

first: Eric

last: Simon

address: Birkendorfer Str. 65

country: Germany

institute: Boehringer Ingelheim Pharma GmbH & Co KG

summary: Progression of non-alcoholic fatty liver disease (NAFLD) to non-alcoholic steatohepatitis (NASH) is a major cause of end-stage liver diseases. There is a high need for predictive human ex vivo models to translate preclinical findings for potential new NAFLD drugs to human disease. About a decade ago, precision cut liver slices (PCLSs) have been established as an ex vivo assay for humans and other organisms. The molecular and functional baseline characteristics of PCLSs have been extensively described previously. In the present study, we provide a comprehensive RNA-Seq based molecular characterization of a new assay for NAFLD that induces steatosis in human and mouse PCLSs culture by incremental supplementation of sugars (glucose and fructose), insulin, and fatty acids (palmitate, oleate). Independent of the triglyceride levels of the human donor liver organs at baseline, the nutrient over-supplementation leads to an increase of triglyceride synthesis and cytokine release to the culture supernatant and thus recapitulates hallmark features of steatosis and inflammation in NAFLD. The mirrored design of human vs. mouse liver slices cultured at the exact same conditions allows a comprehensive assessment of donor variability and the contribution of time vs. nutrient factors to the overall observed molecular pattern and the investigation of conserved and species-specific transcriptional pathways that are involved in steatosis and inflammation of the liver.

overall_design: 99 samples from human precision cut liver slices treated incrementally with sugar, insulin and lipids. Small human liver wedges were collected from 7 human donors following partial resection, donor liver resizing or when livers were unsuitable for transplantation. These experiments were approved by the Medical Ethical Committee of the University Medical Centre Groningen. Liver tissue was stored in University of Wisconsin preservation solution (UW, 4 °C).  We prepared eight different media i.e. CTR: Control with 25mM Glucose, G: 36mM Glucose, F: 25mM Glucose + 5mM Fructose, GF: 36mM Glucose + 5mM Fructose, GFI: GF + 1nM Insulin, GFIO: GFI + 480uM Oleic acid, GIP: GFI + 240uM Palmitic acid, GFIPO: GFI + 480uM Oleic acid + 240uM Palmitic acid) and cultured PCLSs from each donor for 24h and 48h. From the 112 biological samples, 13 samples did not provide sufficient RNA & data quality for further processing.

first: E

first: Maciej

last: Motyka

first: Grietje

middle: H

last: Prins

first: Peter

last: Olinga

first: Anouk

last: Oldenburger


title: Transcriptomic profiling of induced steatosis in precision-cut liver slices

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


summary: Antibody persistence frequently determines the longevity of immunity, yet the mechanisms that set antibody-producing plasma cell turnover rates are unestablished. While it is proposed that new plasma cells displace existing ones in competition for specialised survival niches, it is also possible that the spontaneous loss of existing plasma cells frees niches thereby creating opportunities for recruitment, possibilities with quite different implications for maximising antibody production and persistence. Here, using a genetic reporter to timestamp plasma cells, we define the distribution, phenotype, transcriptional program and population structure of long-lived plasma cells and calculate turnover dynamics. We show that the attributes of longevity are acquired and that turnover of long-lived plasma cells at homeostasis is unaffected by the absence of B cells, indicating that the mechanism of turnover is independent of competition. We propose that understanding plasma cell population dynamics may allow their manipulation to achieve specific outcomes.

overall_design: BLTcre.mcl1fl/+ reporter mice were given tamoxifen to trigger the expression of human CD4 reporter gene. Single cell whole transcriptome profiles of human CD4+ and human CD4- plasma cells sorted from BLTcre.mcl1fl/+ reporter mice 128 days after tamoxifen treatment

first: Marcus

last: Robinson

first: Craig

last: McKenzie

first: Nick

last: Wong

first: David

last: Tarlinton


overall_design: BLTcre.mcl1fl/+ reporter mice were given tamoxifen to trigger the expression of a reporter human CD4 gene. Whole transcriptome RNA sequencing of human CD4+ and human CD4- plasma cells sorted from BLTcre.mcl1fl/+ reporter mice 60 days and 270 days after tamoxifen treatment


title: Transcriptomic profiling of induced steatosis in mouse precision-cut liver slices

overall_design: Animal Study   Male C57/BL6 (Centrale Dienst Proefdieren, UMCG, Groningen, The Netherlands) were housed under standard conditions with chow and water ad libitum. Experiments were approved by the Animal Ethical Committee of the University of Groningen (Permit number 171290-01-001). At the time of sacrifice, mice were aged 10 weeks and their mean body weight was 26.4g. Livers were harvested after exsanguination under isoflurane/O2 anesthesia and stored in UW. Liver organs from five mice have been used for PCLS preparation.  we incubated 48 PCLSs per animal (n=5), yielding a total of 80 samples. All mouse samples have been included. We prepared eight different media i.e. CTR: Control with 25mM Glucose, G: 36mM Glucose, F: 25mM Glucose + 5mM Fructose, GF: 36mM Glucose + 5mM Fructose, GFI: GF + 1nM Insulin, GFIO: GFI + 480uM Oleic acid, GIP: GFI + 240uM Palmitic acid, GFIPO: GFI + 480uM Oleic acid + 240uM Palmitic acid) and cultured PCLSs from each donor for 24h and 48h


title: Co-transcriptional genome surveillance by HUSH is coupled to termination (ChIP-Seq)

first: Tomek

last: Swigut

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) in R1 mouse embryonic stem cells

last: Spencley

first: Shiran

first: Ryan


title: Accelerated replicative senescence of ataxia-telangiectasia skin fibroblasts is retained at physiologic oxygen levels, with unique and common transcriptional patterns

city: Tel-Aviv

first: Amit

last: Levon

address: Ramat Aviv

country: Israel

institute: Tel-Aviv University

summary: Genome instability enhances the pace of aging. Therefore, genome instability syndromes are usually associated with accelerated segmental aging. Genome integrity in the face of ongoing DNA damage is maintained primarily by the DNA damage response (DDR) – a complex signaling network that is induced by DNA lesions. Among the strongest DDR inducers are double-strand breaks (DSBs) – extremely cytotoxic lesions. The chief mobilizer of the DSB response is the ATM protein kinase. ATM loss in humans leads to a multisystem genome instability syndrome, ataxia-telangiectasia (A-T). Part of the complex A-T phenotype is segmental premature aging. Cellular senescence is a cell fate that includes cell cycle arrest, marked alterations in chromatin organization and metabolic circuits, and a multi-faceted senescence-associated secretory phenotype (SASP). This process plays a role in development and tissue homeostasis, as well as age-related degenerative and malignant diseases. Senescent cells increase with age in all tissues studied, and consequently, the SASP can disrupt the tissue microenvironment. Early studies demonstrated a rapid decline in the proliferation of cultured primary skin fibroblasts from A-T patients, which was attributed to premature senescence.  Comparative RNA-sequencing analysis of A-T and control fibroblasts revealed unique transcriptional alterations in A-T cells and their dynamics over time in culture. Cluster analysis of differentially expressed genes (DEGs) allowed for the detection and categorization of these transcriptomic patterns. Functional enrichment analyses provided insight into the molecular pathways and cellular components associated with the premature senescence of A-T cells. The results were further corroborated by analyzing alterations in gene expression profiles using ‘gene set enrichment analysis’ (GSEA). Altogether, the analyses point to engagement of interferon signaling and extracellular matrix remodeling pathways, which are presumably caused by an accumulation of DNA damage. Notably, these pathways are known to be involved in cellular senescence and various age-related pathologies, including fibrosis and cancer. Our data provide a molecular dimension to the segmental premature aging observed in A-T patients

overall_design:
Total number of sequenced samples= 33. Samples from both AT and WT cells (4 replicates each genotype) were collected at different passage levels until they ceased to divide and underwent senescnece. Irradiated WT cells with 10Gy were used as a positive control of senescent cells. AT replicates = 4 , WT replicates=4 ,IR replicates=4.

Please note that each sample was collected during a specific time frame, indicated by the R# in the sample titles;
R1 collected between passage levels 4-6, R2 between 12-14, R3 between 20-22, R4 between 35-38 and R5 at passage 42 (only one cell line reached this level).

first: Majd

last: Haj

first: Yann

last: Fret

first: Judith

last: Campisi

first: Ran

last: Elkon

first: Yael

last: Ziv

first: Yosef

last: Shiloh

first: Noa

last: Hourvitz

first: Yehuda

last: Tzfati


title: Spatiotemporal resolution of germinal center Tfh cell differentiation and divergence from central memory CD4+ T cell fate

first: Fangming

last: Zhu

summary: Transcriptome analysis of CD4+ T cells responding to influenza infection.

overall_design: Purified naïve OT-II T cells were transferred into CD45.1+ SMARTA recipient mice followed by intranasal infection with PR8-OVA. The donor OT-II cells from the medLN of the recipient mice were analyzed by RNA-seq at day 21 p.i.

last: McMonigle

first: Diego

last: Sialer

first: Braxton

last: Greer

first: Edahi

last: Gonzalez Avalos

first: Hui

last: Hu


title: Single-cell dissection of aggression in honeybee colonies

city: Princeton

first: Ian

middle: Michael

last: Traniello

state: NJ

address: Princeton University

department: Lewis-Sigler Insitute for Genomic Biology

institute: Princeton University

summary: RNA-Sequencing performed on 177 honey bee whole-brains, divided into "soldier" and "forager" groups from Puerto Rican honey bee colonies.

overall_design:
Sample phenotyping and collection:
Thirteen colonies from eight locations were sampled across Puerto Rico.  All colonies except the one residing there were transported to the University of Puerto Rico’s Gurabo Agricultural Field Station. Colonies were set up so that no one colony was less than 1 meter apart from another. Following transportation, colonies were allowed a period of at least two weeks to acclimate to the new location before undergoing our phenotyping assay and sample collection.
Phenotype was measured using two assays applied to each colony individually. In the first assay we closed the colony entrance and collected returning foraging bees (naïve foragers, NFr group), flash freezing them with liquid nitrogen as they were aspirated from the colony entrance using a modified collection vacuum The entrance was then re-opened, and immediately after, a string of black suede patches was placed 21 cm from the entrance. Once in place we disrupted the colony by rhythmically striking the top cover. This elicited a defensive response from bees which streamed out of the colony to sting the black suede patches. For two minutes from the beginning of the disruption, we aspirated and flash-froze emerging bees in the act of stinging (aggressors, Agg group). At the end of the two minutes we rapidly collected the suede patches, placing them in a sealable plastic bag and removing them from the area. A researcher then stepped ~3 meters from the colony, and rapidly collected and flash froze bees that followed and remained hovering (pursuers, Pur group). Quickly after we restricted the entrance to the colony and used talcum powder to dust bees that continued to emerge for 30 seconds more. The colony was then left alone for 30 minutes, after which the entrance was again sealed, and we collected those bees returning to the colony that had obvious signs of foraging (pollen loads) and retained the talcum powder (returning foragers, For group). This last group were bees present during disturbance, i.e. perceived the disrupting stimulus, but did not respond with a defensive action.
The method simultaneously provided the individual level phenotype and one of the two components of the colony phenotype. Colony phenotype from this assay was measured as the number of stings counted across the set of black suede patches. Only two colonies were sampled each day using this protocol, observing that the two colonies sampled were no closer than 3 meters from each other. Our scheduling also assured that colonies adjacent to the those phenotyped were themselves undisturbed for at least 48 hours when possible. Upon completion of collection, samples were transported in liquid nitrogen to the University of Puerto Rico, Rio Piedras, and transferred to a -80 °C freezer.
The second phenotyping assay took place two weeks after the last colony was phenotyped by our first method. The process examined colony aggression using a behavioral ranking assay which assigns intensity to four discrete behaviors for each colony then sums the total. This approach is a variation of a standard practice in assessing colony-level defensive behavior. A Kendall’s tau non-parametric correlation test between the scores showed a significant positive correlation between the metrics (Kendall’s tau, n = 13, z = 0.38, p-value = 0.044.  Our final phenotype score was derived by combining these ranks using a multi-dimensional scaling (MDS) analysis. The MDS provided two dimensions, one highly correlated with colony defense in both scores (Dimension 1 vs. Mean of Rank Scores, Kendall’s Tau, n = 13, z = 4.76, p-value < 0.001; Dimension 2 vs. Mean of Rank Scores, Kendall’s Tau, n = 13, z = -0.24, p-value = 0.807), and the second dimension correlated with consistency between the scores (Dimension 1 vs. Mean of Rank Scores, Kendall’s Tau, n = 13, z = 0.18, p-value = 0.855; Dimension 2 vs. Mean of Rank Scores, Kendall’s Tau, n = 13, z = -4.71, p-value < 0.001).
Immediately after phenotyping a colony in this way, we collected and flash-froze 12 queens from all 13 colonies. Two of these queens came from one colony that was poised to split (colony number 6). At the end of the study, samples were individually assessed and sorted into microcentrifuge tubes while on dry ice in preparation for shipping. Samples were later transported to the facilities at the Carl R. Woese Institute for Genomic Biology inside a dry-shipper (CXR500 Cryogenic Shipper, Taylor-Wharton America, Baytown, TX).
Sample selection for sequencing:
Of the thirteen sampled colonies, we excluded three from consideration for sequencing. For two of these, colonies number 5 and 13, we were unable to secure the queen, and thus had no representative parental genotype. The remaining colony number 8, showed signs of swarming at the point of queen collection, likely to have occurred at some point after the first phenotyping effort. Due to uncertainty in sample and indeed colony phenotype, we decided not to consider this colony. Colony 6 was not excluded, as both queens were collected, and the mother-queen was identified through ovary dissections. For the remaining ten colonies we retained over one hundred individuals for each of the four behavioral groups. A total of twenty Agg and For individuals were selected for sequencing along with their corresponding queens. Honey bees from both these groups would have experienced the colony disturbance but responded in markedly different ways.

first: Arian

last: Avalos

first: Amy

last: Ahmed

first: Gene

middle: E

last: Robinson


summary: Each sample includes 5 pooled whole-brains from adult worker honey bees collected across two behavioral groups: soldiers and foragers. We used the Chromium Single Cell 3' Reagent Kits with the Chromium Controller (10x Genomics) and performed sequencing on the Illumina NovaSeq 6000.

overall_design: We dissected and pooled 5 whole-brains from soldiers and forager adult worker honey bees. We collected one soldier and one forager sample (5 brains per sample) from four colonies (eight samples total, two per colony) headed by naturally mated queens at the University of Illinois at Urbana-Champaign Bee Research Facility in Urbana, IL. All samples were collected in August 2019.


summary: Chromatin accessibility analysis of CD4+ T cells responding to influenza infection.

overall_design: Purified naïve OT-II T cells were transferred into CD45.1+ SMARTA recipient mice followed by intranasal infection with PR8-OVA. The donor OT-II cells from the medLN of the recipient mice were analyzed by ATAC-seq at day 14 p.i.

first: Ryan


title: Transcriptome of E18.5 CHD4-M195I mouse hearts

status: Public on May 08 2023

city: Chapel Hill

first: Wei

last: Shi

state: NC

address: 221 Fordham Hall

department: Biology

institute: UNC-Chapel Hill

summary: Purpose: The goal of this study is to compare the cardiac transcriptome profiling (RNA-seq) of WT and CHD4-M195I hearts at E18.5 to conclude genes affected by this CHD4 mutation. Methods: mRNA profiles of E18.5 WT and CHD4-M195I mouse hearts were generated by deep sequencing, n=4 for each genotype, using Illumina HiSeq2500. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows–Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. Results: RNA-sequencing (RNA-seq) analyses on E18.5 WT and CHD4-M195I hearts and identified 323 genes that were differentially expressed [adjusted P value <0.05, |log2(Fold Change)| > 0.5], 113 upregulated and 210 downregulated in CHD4-M195I hearts.

overall_design: WT and CHD4-M195I hearts at E18.5, n=4 for each genotype.

first: Frank

last: Conlon


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


title: ChIP-seq of holocentric Chionographis japonica to determine the centromere distribution and genome organization

city: Seeland OT Gatersleben

first: Anne

last: Fiebig

address: Correnstraße 3

department: BIT

institute: IPK Gatersleben

summary: To determine the distribution of centromere units in the genome of holocentric Chionographis japonica, we performed CENH3-ChIPseq using the customized species-specific CENH3 antibody. We mixed the chromatins of C. japonica and Secale cereal (inbred line Lo7) to dilute the highly abundant centromeric Chio satellite repeats (16%) in the C. japonica genome before immunoprecipitation. In addition, to determine the large-scale genome organization, we performed ChIPseq by targeting the evolutionarily conserved eu- and heterochromatin-specific histone marks H3K4me2 and H33K9me2

sample_organism:
Secale cereale
Chionographis japonica

platform_organism:
Secale cereale
Chionographis japonica

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for chromatin repeat CENH3 as well as conserved eu- and heterochromatin-specific histone marks H3K4me2 and H33K9me2. Each CENH3-, H3K4me2-, and H3K9me2-ChIPseq experiments was performed for two biological replicates.

first: Yi-Tzu

last: Kuo

first: Himmelbach

last: Axel

first: Andreas

last: Houben

first: Takayoshi

last: Ishii

first: Amanda

last: Câmara

first: Veit

last: Schubert

first: Pavel

last: Neumann


title: Single cell RNAseq of co-cultured mouse iPSC-derived lung mesenchyme and mouse iPSC-derived lung epithelial progenitors in distal and proximal differentiation media

city: Boston

first: Pushpinder

last: Bawa

address: 670 Albany St

department: Boston University Medical Campus

institute: Center for Regenerative Medicine

summary: Co-cultures of lung epithelium and mesenchyme are useful tools to study epithelial-mesenchymal crosstalk in lung development and disease. However, many previous attempts to generate such co-cultures have yielded poor juxtaposition between the epithelial and the mesenchymal lineage. In addition, induced pluripotent stem cell (iPSC)-derived co-cultures often contain generic mesenchyme that is not necessarily lung-specific. We sought to establish co-cultures of purified mouse iPSC-derived lung-specific mesenchyme and iPSC-derived lung epithelial progenitors. We used a mouse iPSC line carrying a lung mesenchyme-specific reporter/tracer (Tbx4-LERGFP) to generate lung mesenchymal progenitors by directed differentiation via a lateral plate mesodermal progenitor state (induced lung mesenchyme, iLM). In parallel we differentiated a mouse embryonic stem (ES) cell line carrying a Nkx2-1mCherry reporter into lung epithelial progenitor cells using our established directed differentiation protocol. We then combined the purified lung epithelial and mesenchymal progenitor cells and co-cultured them in distal or proximal differentiation media for 1 week on Matrigel. We find that cells self-organize into complex 3-dimensional organoids with closely juxtaposed epithelial and mesenchymal cells. Furthermore, co-culture affects the molecular phenotype of both lineages. Our iPSC-derived co-culture model can provide an inexhaustible source of cells for studying lung development, modeling diseases, and developing therapeutics.

overall_design: Global transcriptomic profiling at single cell resolution of ESC-derived lung epithelial progenitors cultured alone or with iPSC-derived lung mesenchyme (iLM) in distal or proximal differentiation media: 1) “e pre”: ESC-derived lung epithelium before co-culture carrying a Nkx2-1mCherry reporter, sorted based on Epcam+/Nkx2-1mCherry+, 2) “iLM pre” iPSC-derived lung mesenchyme before co-culture carrying a Tbx4-LERGFP reporter, sorted based on Tbx4-LERGFP+, 3) e-dist: ESC-derived lung epithelium carrying a Nkx2-1mCherry reporter cultured in distal differentiation medium for 7 days, sorted based on Epcam+/Nkx2-1mCherry+. 4) e-prox: ESC-derived lung epithelium carrying a Nkx2-1mCherry reporter cultured in proximal differentiation medium for 7 days, sorted based on Epcam+/Nkx2-1mCherry+. 5) me-dist: iPSC-derived lung epithelium carrying a Nkx2-1mCherry reporter and iPSC-derived lung mesenchyme carrying a Tbx4-LERGFP reporter co-cultured in distal differentiation medium for 7 days, sorted based on Epcam+/Nkx2-1mCherry+ and Tbx4-LERGFP+ and recombined at a 1/1 epithelial/mesenchymal ratio. 6) me-prox: iPSC-derived lung epithelium carrying a Nkx2-1mCherry reporter and iPSC-derived lung mesenchyme carrying a Tbx4-LERGFP reporter co-cultured in proximal differentiation medium for 7 days, sorted based on Epcam+/Nkx2-1mCherry+ and Tbx4-LERGFP+ and recombined at a 1/1 epithelial/mesenchymal ratio.

first: Andrea

last: Alber

first: Feiya

last: Wang

first: Darrell

middle: N

last: Kotton


title: Perfluoroalkyl substances (PFAS) impact inflammation in lung cells and tissues

city: Burlinton

first: Arti

last: Sukla

state: VT

address: 89 Beaumont Avenue

department: Pathology and Laboratory Medicine

institute: University of Vermont

summary: Using mouse lungs from perfluorooctanoic acid (PFOA) exposed mice, we examine effects of exposure to short and long chain PFAS alone or in a mixture on NLRP3 inflammasome activation and cytokine productions.

overall_design: Using mouse lungs from perfluorooctanoic acid (PFOA) exposed mice, we examine effects of exposure to PFOA on lung gene expression.

first: Julie

last: Dragon

first: Michael

last: Hoaglund

first: Appala

last: Badireddy

first: Greylin

last: Nielsen

first: Jennifer

last: Schlezinger

last: Shukla


title: Pbp1 associates with Puf3 and promotes translation of its target mRNAs involved in mitochondrial biogenesis

city: Dallas

first: Benjamin

last: Tu

state: TEXAS

address: 5323 Harry Hines Blvd.

department: Biochemistry

institute: UT Southwestern Medical Center

summary: Pbp1 (polyA-binding protein - binding protein 1) is a stress granule marker and  polyglutamine expansions in its mammalian ortholog ataxin-2 have been linked to neurodegenerative conditions. Pbp1 was recently shown to form intracellular assemblies that function in the negative regulation of TORC1 signaling under respiratory conditions. Furthermore, it was observed that loss of Pbp1 leads to mitochondrial dysfunction. Here, we show that loss of Pbp1 leads to a specific decrease in mitochondrial proteins whose encoding mRNAs are targets of the RNA-binding protein Puf3, suggesting a functional relationship between Pbp1 and Puf3. We found that Pbp1 stabilizes and promotes the translation of Puf3-target mRNAs in respiratory conditions, such as those involved in the assembly of cytochrome c oxidase. We further show that Pbp1 and Puf3 associate through their respective low complexity domains, which is required for target mRNA stabilization and translation. Our findings reveal a key role for Pbp1-containing assemblies in enabling the translation of mRNAs critical for mitochondrial biogenesis and respiration under metabolically challenging conditions. They may further explain prior associations of Pbp1/ataxin-2 with stress granule biology and RNA metabolism.

overall_design: Two biological replicates of wild type or ∆pbp1 cells grown in YPD media or switched from YPD to YPL media and grown for an additional 3 hrs were collected for RNA-seq.

first: Floortje

last: van de Poll

last: Sutter

first: Michelle

middle: G

last: Acoba

first: Daniel

last: Caballero

first: Samira

last: Jahangiri

first: Chien-Der

last: Lee

first: Yu-San

last: Yang

middle: P


title: Classification of IRF3- vs ISGF3-dependent gene induction in murine fibroblasts

city: Würzburg

first: Florian

last: Erhard

address: Versbacher Str. 7

department: Institut für Virologie und Immunbiologie

institute: Julius-Maximilians-Universität Würzburg

summary:
To investigate the dependency on gene expression on the PRR-activated transcription factor IRF3 versus the type I IFN receptor (IFNAR)-activated transcription factor ISGF3, we extracted primary murine embryonic fibroblasts (MEF) of wild-type, IRF3-knockout or IFNAR-knockout mice (strain C57BL/6J, protocol of Podlech et al., 2002).
We stimulated the 3 different cell lines by transfection of immunostimulatory DNA (ISD, 4 hours) for activation of IRF3, or by treatment with murine IFNbeta (3 hours) for activation of the IFNAR-ISGF3 cascade. Mock-transfected (4 hours) or untreated cells served as respective controls.
In the last 2 hours before harvest, cells were additionally treated with 200µM 4-thiouridine (4sU) for metabolic labelling of nascent transcripts, and total mRNAs were processed accordingly by SLAM-sequencing procedures for gene expression profiling of newly synthesized transcripts.

overall_design: Comparative gene expression profiling analysis of nascent transcripts from SLAM-seq data for gene induction by PRR-IRF3 vs. by IFNAR-ISGF3 signalling.

first: Melanie

middle: M

last: Brinkmann

first: Hella

last: Schwanke

first: Emanuel

last: Wyler

first: Caroline

last: Friedel


title: The nutrient-sensing GCN2 signaling pathway is essential for circadian clock function by regulating histone acetylation under amino acid starvation

city: Beijing

first: Xiaolan

last: Liu

address: NO.1 Beichen West Road, Chaoyang District

institute: Institute of Microbiology, Chinese Academy of Sciences

summary: Circadian clocks are evolved to adapt to the daily environment changes under different conditions. The ability to maintain circadian clock functions in response to various stress and perturbations is important for organismal fitness. Here, we show that the nutrient sensing GCN2 signaling pathway is required for robust circadian clock function under amino acid starvation in Neurospora. The deletion of GCN2 pathway components disrupts rhythmic transcription of clock gene frq by suppressing WC complex binding at the frq promoter due to its reduced histone H3 acetylation levels. Under amino acid starvation, the activation of GCN2 kinase and its downstream transcription factor CPC-1 establish a proper chromatin state at the frq promoter by recruiting the histone acetyltransferase GCN-5. The arrhythmic phenotype of the GCN2 kinase mutants under amino acid starvation can be rescued by inhibiting histone deacetylation. Finally, genome-wide transcriptional analysis indicates that the GCN2 signaling pathway maintains robust rhythmic expression of metabolic genes under amino acid starvation. Together, these results uncover an essential role of GCN2 signaling pathway in maintaining robust circadian clock function in response to amino acid starvation and the importance of histone acetylation at the frq locus in rhythmic gene expression.

overall_design: For RNA-seq analyses, strains cultured in 2% glucose in constant light  at 25 °C for 18h were havested. Total RNAs were extracted using Trizol reagents.The purity of the sample was determined by NanoPhotometer ® (IMPLEN, CA, USA)；The concentration and integrity of RNA samples were detected by Agilent 2100 RNA nano 6000 assay kit (Agilent Technologies, CA, USA). Sequencing libraries were generated using VAHTS Universal V6 RNA-seq Library Prep Kit for Illumina ® (NR604-01/02) (Annoroad Gene Technology Co., Ltd, Beijing)  following the manufacturer’s recommendations and index codes were added to attribute sequences to each sample.Libraries were prepared according to the manufacturer’s instructions and analyzed using 150bp paired-end Illumina sequencing (Annoroad Gene Technology, Beijing).

first: Yulin

first: Xiao


title: Effect of MCMV M35 on global gene expression during PRR signalling in murine fibroblasts

summary:
To investigate if the protein M35 of murine cytomegalovirus (MCMV) affects transcription of genes beside IFNbeta, we immortalised murine embryonic fibroblasts (MEF) isolated from wild-type C57BL/6J mice with SV40 Large T antigen to generate iMEF, and transduced these with retrovirus to stably express M35-HAHA or carry a control vector (EV).
We stimulated the 2 different cell lines by transfection of Alexa488-labelled immunostimulatory DNA (ISD) for 2, 4 or 6 hours to activate the host transcription factor IRF3. Mock-transfected and untreated cells served as controls.
In the last 90 min before harvest, cells were additionally treated with 200µM 4-thiouridine (4sU) for metabolic labelling of nascent transcripts, and total mRNAs were processed accordingly by SLAM-sequencing procedures for gene expression profiling of newly synthesized transcripts.

overall_design: Comparative gene expression profiling analysis of total transcripts from SLAM-seq data for differential gene expression in the presence of M35 compared to control cells at different time points of stimulation.


title: An integrated multiorgan feed-forward metabolic response exacerbates muscle wasting and lipid accumulation in human mitochondrial myopathies

first: Nneka

last: Southwell

address: 407 E 64th St Fl 5

department: Brain and Mind Research Institute

institute: Weill Cornell Graduate School

summary: Mitochondrial diseases are a heterogeneous group of monogenic disorders that result from impaired oxidative phosphorylation (OXPHOS). As neuromuscular tissues are highly energy-dependent, mitochondrial diseases often affect skeletal muscle. Although genetic and bioenergetic causes of OXPHOS impairment in human mitochondrial myopathies are well established, there is a limited understanding of metabolic drivers of muscle degeneration. This knowledge gap contributes to the lack of effective treatments for these disorders. Here, we discovered fundamental muscle metabolic remodeling mechanisms shared by mitochondrial disease patients and a mouse model of mitochondrial myopathy. This metabolic remodeling is triggered by a starvation-like response that evokes accelerated oxidation of amino acids through a truncated Krebs cycle. While initially adaptive, this response evolves in an integrated multiorgan catabolic signaling, lipid store mobilization, and intramuscular lipid accumulation. We show that this multiorgan feed-forward metabolic response involves leptin and glucocorticoid signaling. This study elucidates systemic metabolic dyshomeostasis mechanisms that underlie human mitochondrial myopathies and identifies potential new targets for metabolic intervention.

overall_design: Libraries were sequenced with single-end 86 bps on an Illumina NextSeq500 sequencer

first: Marilena

last: D'Aurelio


title: DDT-RELATED PROTEIN4-IMITATION SWITCH alters nucleosome distribution to relieve transcriptional silencing in Arabidopsis [MNase-seq]

first: Zejia

last: Wang

address: Yanyuan Street

institute: Peking University

summary: DNA methylation is a conserved epigenetic modification that usually functions to repress gene expression. However, not all DNA methylated loci are faithfully silenced, and genes bearing promoter DNA methylation often show varying levels of transcription. The flexibility in transcription allows for greater adaptability in response to environmental and developmental cues, the mechanisms of which are not fully understood. In this work, we demonstrate that a plant-specific ISWI complex, consisting of CHR11, CHR17, DDR4, and DDR5, functions to partially de-repress DNA methylated genes and TEs through chromatin relaxation. This action requires the known transcriptional activator DNAJ proteins, providing a mechanistic link between nucleosome remodeling and transcriptional activation. We also show that the anti-silencing effects of enhanced chromatin accessibility can be genetically uncoupled from that of DNA demethylation, and is antagonized by the chromatin compactor, MORC proteins. Our work reveals a mechanism for balancing between the needs of transcriptional flexibility and faithful silencing of DNA methylated loci. As both ISWI and MORC family genes are distributed widely across plant and animal species, the findings may represent a conserved eukaryotic mechanism for fine-tuning gene expression under epigenetic repression.

overall_design: The dataset contain 37 RNAseq samples, 7 ChIPseq samples, 4 WGBS samples, 10 MNase-seq samples and 58BSPCR samples. Plants are of different genetic backgrounds, which include col-0, morc1 mutant, morc1/ddr4 mutant, ZF-DDR4 mutant, met1 mutant, drm3/cmt3 mutant, drm2/cmt3/ddr4 mutant, ZF-DDR4/chr11/17 mutant.

first: Qikun

first: Qiyan

last: Zhang


title: Mitochondrial energetic stress triggers AMPK/FNIP1-dependent TFEB nuclear translocation, inducing transcriptional waves of lysosomal and mitochondrial biogenesis.

city: La Jolla

first: Jingting

last: Yu

address: 10010 N Torrey Pines Rd

institute: Salk Institute for Biological Studies

summary: Cells respond to mitochondrial energetic stress with rapid activation of the AMP-activated protein kinase (AMPK), which acutely inhibits anabolism and stimulates catabolism.  AMPK also induces sustained transcriptional reprogramming of metabolism.  The TFEB transcription factor is a major effector of AMPK signals, inducing lysosome genes following energetic stress. Yet the molecular mechanism underlying how AMPK activates TFEB remains unresolved.  We demonstrate here that AMPK directly phosphorylates five conserved serine residues in FNIP1, which suppresses the function of the FLCN/FNIP1 RagC GAP complex, in turn controlling TFEB lysosomal localization.  We demonstrate that FNIP1 phosphorylation is required for AMPK to induce nuclear translocation of TFEB, which is fully separable from AMPK control of canonical mTORC1 signaling.  Using a non-phosphorylatable allele of FNIP1, we show that in parallel to lysosomal biogenesis, AMPK induces mitochondrial biogenesis via TFEB-dependent induction of PGC1a mRNA.  This signaling from mitochondrial stress is also independent of amino-acid control of the Rags and TFEB, which still proceed normally in cells bearing the AMPK-non phosphorylatable allele of FNIP1.  Taken together, mitochondrial energetic stress triggers AMPK/FNIP1-dependent TFEB nuclear translocation, inducing transcriptional waves of lysosomal and mitochondrial biogenesis.

overall_design: We generated  FNIP1 knock out HEK 293T cells, which were reconstituted with WT FNIP1 or mutant FNIP1 cDNA, in which all 5 AMPK phosphosites had been mutated to alanine (SA5). Following treatment with the synthetic AMPK activator 991 for 0-16 hours, RNAseq was   RNAseq was carried out to examine mitochondrial and lysosomal gene transcription under these conditions. Additionally, WT or AMPK KO HEK 293T cells were treated with 991 or the mitochondrial poisons CCCP, Phenformin or Rotenone to observe effects on global gene transcription and more specifically on lysosomal and mitochondrial gene sets. Similarly, RNAseq was carried out following 991 treatments in WT and TFEB/TFE3 double knock out. HEK 293T; or WT and ERRa knock out HEK 293T cells to examine the role of TFEB/TFE3 and ERRa on lysosomal and mitochondrial gene transcription.  RNAseq was also carried out following PGC1A knockdown by siRNA and compared with control siRNA conditions following 991 treatments for 0-16 hours to assess. effects on mitochondrial gene transcription.

first: Nazma

last: Malik

first: Nasun

last: Hah

first: Reuben

last: Shaw


title: PIK3CA mutation in endometrial epithelia promotes Viperin-dependent inflammatory response to insulin

first: Ronald

last: Chandler

address: 400 Monroe Ave NW

institute: Michigan State University

overall_design: Refer to individual Series


title: DDT-RELATED PROTEIN4-IMITATION SWITCH alters nucleosome distribution to relieve transcriptional silencing in Arabidopsis [Bisulfite-seq]


title: DDT-RELATED PROTEIN4-IMITATION SWITCH alters nucleosome distribution to relieve transcriptional silencing in Arabidopsis

relation:
SuperSeries of: GSE202204
SuperSeries of: GSE202205
SuperSeries of: GSE202270
SuperSeries of: GSE227074


title: Diurnal changes in bacterial settlement on the Peyer’s patch and surrounding mucosa in the rat ileum and its effect against the intestinal immune system

status: Public on May 07 2023

city: Kobe

first: Youhei

last: MANTANI

state: Hyogo

address: Rokkodai-cho 1-1, Nada-ku

department: Graduate school of Agricultural Science

country: Japan

institute: Kobe University

summary:
To investigate how stimulation from indigenous bacteria for a day affects the local transcriptome of both Peyer's patch and surrounding ordinary mucosa in the rat ileum, we treated rats with a nonabsorbable antibiotics cocktail (vancomycin, polymyxin B and sucrose) for antibiotics-treatment group or sucrose solution for control group in drinking water 1 day before euthanasia.
We then  performed gene expression profiling analysis using data obtained from RNA-seq of ileal Peyer's patch and surrounding ordinary tissue from rats with or without antibiotics treatment.

overall_design: Comparative gene expression profiling analysis of RNA-seq data from Peyer's patch and ordinary ileal tissue between control group and antibiotics treated group.

first: Naoto

last: KUBOTA


title: Development and validation of urinary exosomal microRNA biomarkers for the diagnosis of acute rejection in kidney transplant recipients

city: Seong-nam

first: Yu Ho

last: Lee

address: 59 Yatap-ro, Bundang-gu

country: South Korea

institute: CHA

summary: Acute rejection prediction classifiers were successfully constructed through expression profiling of a total of 개수 urinary exosomal microRNAs in 108 kidney transplant recipients.

overall_design: In the study presented here, 108 kidney transplant recipients (normal pathology on protocol biopsy (NP, n=19), long-term graft survival (LTGS, n=29)], acute T cell-mediated rejection (TCMR, n=40) and active antibody-mediated rejection (aABMR, n=20)).

first: Jung-Woo

last: Seo

first: Sang-Ho


title: DDT-RELATED PROTEIN4-IMITATION SWITCH alters nucleosome distribution to relieve transcriptional silencing in Arabidopsis (ChIP-seq)


title: NanoSTRING analysis of sorted intratumoral CD8+ T cells following radiotherapy

first: Juliana

last: Idoyaga

state: California

address: 299 Campus drive, 3RD FLOOR RM D330 & D345

department: Microbiology and Immunology

institute: Stanford University

summary:
The recruitment of monocytes and their differentiation into immunosuppressive cells is associated with the negative outcome of non-conformal radiotherapy (RT). However, non-conformal RT is irrelevant in the clinic, and little is known about the role of monocytes following radiotherapy modes used in patients, such as conformal radiotherapy (CRT). Here, we investigated the acute immune infiltration post-CRT. Contrary to non-conformal RT approaches, we found that CRT induces a rapid and robust recruitment of monocytes to the tumor that minimally differentiate into tumor-associated macrophages (TAM) or dendritic cells (DC), but instead upregulate major histocompatibility complex II (MHCII) and costimulatory molecules. We establish that these large numbers of infiltrating monocytes are responsible for increasing type I interferon in the tumor microenvironment (TME), activation of CD8+ T cells and the reduction in tumor burden. Importantly, we demonstrate that rapid monocyte infiltration to the TME is hindered when RT inadvertently affects healthy tissues. Our results unravel a positive role of monocytes during clinically relevant modes of RT and demonstrate that limiting exposure of healthy tissues to radiation has a positive therapeutic effect on the overall immune response within the tumor.
The objective of this experiment was to evaluate the transcriptional changes in CD8+ T cells following different modes of radiation at 3 days post treatment

overall_design: Three different modes of radiatherapy in triplicate. Control samples are marked as Nontreated

first: Derek

last: Clements

first: Sirimuvva

last: Tadepalli


title: NanoSTRING analysis of sorted monocytes/monoACT cells following radiotherapy

summary:
The recruitment of monocytes and their differentiation into immunosuppressive cells is associated with the negative outcome of non-conformal radiotherapy (RT). However, non-conformal RT is irrelevant in the clinic, and little is known about the role of monocytes following radiotherapy modes used in patients, such as conformal radiotherapy (CRT). Here, we investigated the acute immune infiltration post-CRT. Contrary to non-conformal RT approaches, we found that CRT induces a rapid and robust recruitment of monocytes to the tumor that minimally differentiate into tumor-associated macrophages (TAM) or dendritic cells (DC), but instead upregulate major histocompatibility complex II (MHCII) and costimulatory molecules. We establish that these large numbers of infiltrating monocytes are responsible for increasing type I interferon in the tumor microenvironment (TME), activation of CD8+ T cells and the reduction in tumor burden. Importantly, we demonstrate that rapid monocyte infiltration to the TME is hindered when RT inadvertently affects healthy tissues. Our results unravel a positive role of monocytes during clinically relevant modes of RT and demonstrate that limiting exposure of healthy tissues to radiation has a positive therapeutic effect on the overall immune response within the tumor.
The objective of this experiment was to evaulate the transcriptional changes in monocytes following different modes of radiation at 3 days post treatment

overall_design: Two different modes of radiotherapy in triplicate for each sorted population.


title: Radiation-induced Circulating microRNAs predict cardiac dysfunction after radiotherapy

city: Łódź

first: Zuzanna

last: Nowicka

address: Mazowiecka 15

department: Department of Biostatistics and Translational Medicine

country: Poland

institute: Medical University of Łódź

summary: Patients treated with radiotherapy to the chest region are at risk of cardiac sequelae, however, identification of those with greatest risk of complications remains difficult. Here, we sought to determine whether short-term changes in circulating miRNA expression are related to measures of cardiac dysfunction in follow-up. miRNA expression analysis in the primary group showed marked changes in serum miRNAs immediately after RT compared to baseline. miRNAs with increased expression correlated positively with cardiopulmonary dose-volume histogram metrics, while those with decreased expression exhibited negative correlations.

overall_design: Serum samples from 28 patients with left-sided breast cancer were collected before RT, 24h after the last dose and 1 month after RT; the patients were followed up for 3 years after completion of RT and cardiac echocardiography at 2.5-3 year follow-up was performed. All patients underwent adjuvant chemotherapy prior to RT, if indicated. Concomitant treatment with systemic therapies other than chemotherapy was allowed, including treatment with trastuzumab, tamoxifen or aromatase inhibitors.

first: Wojciech

last: Fendler

first: Justyna

last: Chałubińska-Fendler


title: Evolutionarily divergent mTOR remodels translatome for tissue regeneration

city: Palo Alto

first: Maria

last: Barna

state: CA

address: 240 Pasteur Drive, 4500

department: Genetics

summary: An outstanding mystery in biology is why some species, such as the axolotl, can regenerate tissues whereas mammals cannot1. Here, we demonstrate that rapid activation of protein synthesis is a unique feature of the injury response critical for limb regeneration in the axolotl (Ambystoma mexicanum). By applying polysome sequencing, we identify hundreds of transcripts, including antioxidants and ribosome components that are selectively activated at the level of translation from pre-existing messenger RNAs in response to injury. By contrast, protein synthesis is not activated in response to non-regenerative digit amputation in the mouse. We identify the mTORC1 pathway as a key upstream signal that mediates tissue regeneration and translational control in the axolotl. We discover unique expansions in mTOR protein sequence among urodele amphibians. By engineering an axolotl mTOR (axmTOR) in human cells, we show that these changes create a hypersensitive kinase that allows axolotls to maintain this pathway in a highly labile state primed for rapid activation. This change renders axolotl mTOR more sensitive to nutrient sensing, and inhibition of amino acid transport is sufficient to inhibit tissue regeneration. Together, these findings highlight the unanticipated impact of the translatome on orchestrating the early steps of wound healing in a highly regenerative species and provide a missing link in our understanding of vertebrate regenerative potential.

overall_design: Axolotl limbs were amputated and tissue was harvested from the plane of amputation at 0 h and 24 h post-amputation. The tissue was lysed and part of the lysate was retained as the "input" sample for RNA-Seq analysis. The remaining lysate was subjected to sucrose gradient fractionation followed by pooling and RNA-Seq of mRNAs associated with the free/RNP, light polysome and heavy polysome fractions.

first: Olena

last: Zhulyn

first: Hannah

last: Rosenblatt

first: Leila

last: Shokat

first: Shizhong

last: Dai

first: Duygu

last: Kuzuoglu-Öztürk

first: Zijian

first: Davide

last: Ruggero

first: Kevan


title: DDT-RELATED PROTEIN4-IMITATION SWITCH alters nucleosome distribution to relieve transcriptional silencing in Arabidopsis [RNA-Seq]

first: Zejia

last: Wang

overall_design: The dataset contain 37 RNAseq samples, 7 ChIPseq samples, 4 WGBS samples, 10 MNase-seq samples and 58BSPCR samples. Plants are of different genetic backgrounds, which include col-0, morc1 mutant, morc1/ddr4 mutant, ZF-DDR4 mutant, met1 mutant, drm3/cmt3 mutant, drm2/cmt3/ddr4 mutant, ZF-DDR4/chr11/17 mutant.!Series_overall_design =

first: Qikun

last: Liu

first: Qiyan

last: Zhang


title: Lp(a) and 17K recombinant apo(a) treatment of THP-1 macrophages

city: London

first: Michael

last: Boffa

state: ON

address: 1151 Richmond Street North

institute: The University of Western Ontario

summary: Lp(a) and its distinguishing apolipoprotein constituent apo(a) have been shown to elicit proinflammatory responses from monocytes and macrophages. To obtain a global picture of the effect of Lp(a) and apo(a) on the transcriptome of these cells, we treated THP-1 macrophages with 250 nM Lp(a) or apo(a) for 6 hours, harvested RNA from the cells, and then performed RNA-seq using an Illumina NextSeq instrument. Analysis of the data using GO and KEGG strategies revealed a series of proinflammatory genes upregulated by apo(a) and even more dramatically by Lp(a).

overall_design: Duplicate wells of THP-1 macrophages were treated for 6 hours with 250 nM Lp(a) or recombinant 17-kringle apo(a), or vehicle alone (phosphate-buffered saline).

first: Matthew

last: Borrelli

first: David

last: Carter

first: Marlys

last: Koschinsky


title: Single Cell and Spatial Sequencing Define Processes by which Keratinocytes and Fibroblasts Amplify Inflammatory Responses in Psoriasis

city: Los Angeles

first: Feiyang

last: Ma

address: 610, Charles Young Dr East, TLSB 3000C

department: Molecular Biology Institute

institute: UCLA

summary: The immunopathogenesis of psoriasis, a common chronic inflammatory disease of the skin, is incompletely understood. Here we demonstrate, using a combination of single cell and spatial RNA sequencing, IL-36 dependent amplification of IL-17A and TNF inflammatory responses in the absence of neutrophil proteases, which primarily occurred within the supraspinous layer of the psoriatic epidermis. We further show that a subset of SFRP2+ fibroblasts in psoriasis contribute to amplification of the immune network through transition to a pro-inflammatory state. The SFRP2+ fibroblast communication network involves production of CCL13, CCL19 and CXCL12, connected by ligand-receptor interactions to other spatially proximate cell types: CCR2+ myeloid cells, CCR7+ LAMP3+ dendritic cells, and CXCR4 expressed on both CD8+ Tc17 cells and keratinocytes, respectively. The SFRP2+ fibroblasts also express cathepsin S, further amplifying inflammatory responses by activating IL-36G in keratinocytes. These data provide an unprecedented view of psoriasis pathogenesis, which expands our understanding of the critical cellular participants to include inflammatory fibroblasts and their cellular interactions.

overall_design: Visium spatial sequencing comparing psoriasis skin to healthy control skin samples

first: Olesya

last: Plazyo

first: Johann

last: Gudjonsson


title: HSFA2 and HSFA3 binding after heat acclimation

city: Potsdam

first: Isabel

last: Bäurle

address: Karl-Liebknecht-Str. 24-25

department: Institut für Biochemie und Biologie

institute: Universität Potsdam

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


title: HSFA2 and HSFA3 binding after heat acclimation [DAP-seq]

summary: Transcriptional regulation is a key aspect of environmental stress responses. Heat stress (HS) induces transcriptional memory that allows plants to respond more efficiently to a recurrent HS. In light of more frequent temperature extremes due to climate change, improving heat tolerance in crops is an important breeding goal. However, not all HS-inducible genes show sustained induction/transcriptional memory and it is unclear what distinguishes memory and non-memory genes. To address this issue and understand the (epi-) genome architecture in transcriptional memory after HS, we investigated genome-wide target genes of the two key memory heat shock transcription factors, HSFA2 and HSFA3. We determined the binding kinetics of these factors to their target genes and asked whether genes that show sustained induction of transcription carry specific features that allow prediction and potentially engineering of memory gene behaviour. HSFA2 and HSFA3 show near identical binding patterns. In vitro binding strength as determined by DAP-seq analysis correlates strongly with in vivo binding strength, confirming the importance of sequence features. However, no single distinctive sequence motif appears to be required for memory behaviour. Instead, HS memory genes are characterized by a combination of features: low expression levels in the absence of HS, chromatin environment and an enrichment of H3K4 methylation after HS. Our findings are confirmed by an orthogonal transcriptomic data set using both de novo clustering and an established definition of memory genes. In summary, our findings provide an integrated view of HSF-dependent transcriptional memory and shed light on its sequence and chromatin determinants. They will contribute to the prediction and engineering of genes with transcriptional memory.

overall_design: Identification of binding sites for HSFA1b, HSFA2 and HSFA3 in Arabidopsis thaliana using DAP-seq.

first: Li

last: Jiang

first: Christian

last: Kappel

last: Lenhard


title: Central amygdala cell types and their transcriptional regulation by fasting

status: Public on May 06 2023

city: Munich

first: Songwei

last: He

address: Am Klopferspitz 18

institute: Max Planck Institute of Neurobiology

summary: A complete cell type census based on their full molecular profiles at subclass level and their subnuclei distribution in the central amygdala (CeA), linked with their behavior relevance, is still lacking. Here, we performed single nuclei RNA sequencing in adult mice and compared the results with known genetic and functionally defined populations. We identified 9 transcriptionally and spatially discrete cell types (3 in the lateral division CeL; 5 in the medial division CeM; and 1 in the capsular division CeC). In the CeL, we found one PKCδ expressing cluster (CeL PKCδ ) and two clusters marked by Sst: the CeL Sst cluster which contains the largest and most highly expressing population of Sst-positive neurons, and the CeL NTS/Tac2 cluster which contains low expressing Sst-positive cells. This two Sst population are also Htr2a positive. The CeC contains one cluster marked by the expression of Calcrl and partially expressed PKCδ. In the CeM, the largest cluster was marked by Il1Rapl2, which contains the CeM subsets of Pnoc and Htr2a, and is likely appetitive. Another Sst population, the CeM Tac1/Sst, remains largely unexplored. Moreover, we discovered three previously uncharacterized CeM clusters: CeM Vdr, CeM Drd2/Rai14, and CeM Dlk1. To begin investigating whether these cell types show individual responses in appetitive behavior, we compared their transcriptomes in food-deprived and satiated mice. We found a cell type specific transcriptional response across neuronal populations, particularly in one of the appetitive clusters in the CeL.

overall_design: Mouse CeA were dissected out and single nuclei suspension were prepared. Two data sets (biological replicate) were obtained from 8-week old naïve mice. A third data set came from Htr2a-Cre transgenic mice expressing AAV-DIO-mCherry to determine the CeAHtr2a population. Another two datasets (biological replicate) were obtained from mice that were fasted for 20 hours.

first: Ruediger

last: Klein


title: Effects of a novel HBP inhibitor, FR054, and Gemcitabine in pancreatic cancer cells

city: Milan

first: Ferdinando

last: Chiaradonna

address: Piazza della Scienza 2

department: Biotechnology and Biosciences

country: Italy

institute: University of Milano Bicocca

summary: In order to define the role of the Hexosamine Biosynthetic Pathaway (HBP) in pancreatic cancer cells proliferation, survival and gemcitabine resistance, we treated MiaPaCa2 and BxPC3 cells with FR054, a novel HBP inhbitor, and gemcitabine either alone or in combination for 48h

overall_design: Comparative gene expression profiling analysis of RNA-seq data for MiaPaCa2 and BxPC3 cells treated with an HBP inhbitor, FR054, and Gemcitabine alone or in combination. The analysis has been performed by using the biological duplicate of all samples.

first: Barbara

last: Zerbato


title: Single-cell atlases of two lophotrochozoan larvae highlight their complex evolutionary histories

status: Public on May 05 2023

first: Laura

last: Piovani

address: Room B02, Anatomy Building, Gower Street

department: GEE

institute: UCL

summary: Single cell atlases of platyhelminth Müller’s larva and mollusc trochophore larva reveal homologous cell types and phylum specific novelties

sample_organism:
Magallana gigas
Prostheceraeus crozeri

platform_organism:
Magallana gigas
Prostheceraeus crozeri

overall_design: 10x genomics 3' GEX single cell sequencing was performed on trochophore larvae of Crassostrea gigas (3'v2) and 1-day old Muller's larvae of Prostheceraeus crozieri (3'v3.1). Raw fastqs and filtered matrix counts, barcodes and features of libraries Cg1 (Crassostrea gigas), Pc3 and Pc4 (Prostheceraeus crozieri repeat 3 and 4) are made available. For details see linked publication.


title: Allele-specific gene regulation, phenotypes, and therapeutic vulnerabilities in estrogen receptor alpha mutant endometrial cancer (xenograft RNA-seq)

city: Salt Lake City

first: Jason

last: Gertz

state: UT

address: 2000 Circle of Hope

department: Oncological Sciences

institute: University of Utah

summary: Activating estrogen receptor alpha (ER) mutations are present in primary endometrial and metastatic breast cancers, promoting estrogen-independent activation of the receptor. Functional characterizations in breast cancer have established unique molecular and phenotypic consequences of the receptor, yet the impact of ER mutations in endometrial cancer has not been fully explored. In this study, we used CRISPR-Cas9 to model the clinically prevalent ER-Y537S mutation and compared results to ER-D538G to discover allele-specific differences between ER mutations in endometrial cancer. We found that constitutive activity of mutant ER resulted in changes in the expression of thousands of genes, stemming from combined alterations to ER binding and chromatin accessibility. The unique gene expression programs resulted in ER mutant cells developing increased cancer associated phenotypes, including migration, invasion, anchorage independent growth, and growth in vivo. To uncover potential treatment strategies, we identified ER associated proteins via Rapid Immunoprecipitation and Mass Spectrometry of Endogenous Proteins (RIME) and interrogated two candidates, CDK9 and NCOA3. Inhibition of these regulatory protein resulted in decreased growth and migration, representing potential novel treatment strategies for ER mutant endometrial cancer. Taken together, this study provides insight into ER mutant activity in endometrial cancer and identifies potential therapies for women with ER mutant endometrial cancer.

overall_design: RNA-seq was performed on xenografted Ishikawa ER wildtype or ER mutant cell line tumors to assess effects of the ER mutations in vivo.

first: Zannel

last: Blanchard

first: Craig

last: Rush


title: HSFA2 and HSFA3 binding after heat acclimation [ChIP-seq]

overall_design: Identification of HSFA2 and HSFA3 binding sites in Arabidopsis thaliana seedlings 4, 28 and 56 hours after heat acclimation and in non-heat stressed control plants using ChIP-seq. Three replicates for each sample.

first: Thomas

last: Friedrich

first: Vicky

last: Oberkofler

first: Michael


title: Hexokinase 2-mediated gene expression via histone lactylation is required for hepatic stellate cell activation and liver fibrosis (HSC-RNA-Seq)

city: Chicago

first: Nissim

last: Hay

state: IL

address: 900 South Ashland Ave

department: BMG

institute: University of Illinois Chicago

summary: Lactate was implicated in activation of hepatic stellate cells (HSCs). However, the mechanism by which lactate exerts its effect remains elusive. We show that hexokinase 2 (HK2) is sufficient to change gene expression by histone lactylation but not histone acetylation. Using RNA-seq and CUT&Tag chromatin profiling, we found that induction of HK2 expression in activated HSCs is required for the induced gene expression by elevating histone lactylation. Inhibiting histone lactylation by Hk2 deletion or pharmacological inhibition of lactate production diminishes HSC activation, whereas exogenous lactate but not acetate supplementation rescues the activation phenotype. Thus, lactate produced by activated HSCs determines the HSC fate via histone lactylation. We found that histone acetylation competes with histone lactylation, which could explain why class I HDAC inhibitors impede HSC activation. Finally, HSC-specific or systemic deletion of HK2 inhibits HSC activation and liver fibrosis in vivo. Therefore, we provide evidence that HK2 may be an effective therapeutic target for liver fibrosis.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for quiescent (QS), activated (ACT), activated HK2-deleted (KO), KO with lactate-treated (LAC) mouse primary HSCs.

first: Hyunsoo

last: Rho


title: VEGFR3 signaling restrains the neuron-macrophage crosstalk during neurotropic viral infection

city: Shanghai

first: Linlin

last: Qi

state: Shanghai

address: 333 Nanchen Road, Baoshan District

institute: Shanghai University

summary: To investigate the transcriptomic characteristics of CD11b+F4/80+ cells in the brain of JEV- infected WT and Vegfr-3ΔLBD/ΔLBD mice, we sorted the CD11b+F4/80+ cells from  the brain of  WT and Vegfr-3ΔLBD/ΔLBD mice post 5 days of JEV infection by FACS. We then performed gene expression profiling analysis using data obtained from RNA-seq of 2 different cells.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for CD11b+F4/80+ cells in the brain of JEV- infected WT and Vegfr-3ΔLBD/ΔLBD mice.

first: Dan

last: Yang

first: Hongyan

last: Wang

first: Bin

last: Wei


title: Hexokinase 2-mediated gene expression via histone lactylation is required for hepatic stellate cell activation and liver fibrosis (CUT&Tag)

overall_design: Genomic occupancy profiling of H3K18la and H3K18ac for quiescent (QS), activated (ACT), activated HK2-deleted (KO), KO with lactate-treated (LAC) mouse primary HSCs.


title: Low Dose Gamma Irradiation of Trypanosoma evansi Parasites Identifies Gene Transcripts Involved in Establishing Disease in Mice Post Irradiation

city: Seibersdorf

first: Richard

middle: Thiga

last: Kangethe

address: Friedenstrasse 1

country: Austria

institute: IAEA

summary:
Transcriptome analysis of irradiated T evansi parasites
The protozoan parasite Trypanosoma evansi is responsible for causing Surra in a variety of mammalian hosts over a wide geographical area.  In order to identify which genes and processes are required to establish disease in mice, parasites were irradiated over a range using a Cobalt60 gamma source. A custom Trypanosome spp. array that covers the genomes of three trypanosome species, T. brucei, T. evansi and T. congolense was designed by Affymetrix with an  average of 9300 whole gene transcripts from all three species were targeted.  Irradiation differentially affected the abundance of gene transcripts in a dose-dependent trend. We present these genes as necessary for repair from irradiation damage, and essential for disease establishment in mice post irradiation.

overall_design: We analyzed RNA from 67 samples of T. evansi irradiated samples with non irradiated controls using the Affymetrix TrypMS array. Data was processed by Affymetrix Exon Array Computational Tool. A minimum of 3 techinical replicates were performed for each dose and time delay sample point.

first: Eva

last: Winger

first: Tirumala

middle: B

last: Settypalli

first: Sneha

last: Datta

first: Viskam

last: Wijewardana

first: Charles

last: Lamien

first: Hermann

last: Unger

first: Theresa

last: Coetzer

first: Giovanni

last: Cattoli

first: Adama

last: Diallo


title: Hexokinase 2 and lactate

sample_organism:
Cricetulus griseus
Mus musculus

platform_organism:
Cricetulus griseus
Mus musculus

overall_design: Refer to individual Series


title: Multiomics analysis of HIF-stabilization in primary renal tubular cells

city: Aachen

first: René

last: Krüger

address: Pauwelsstraße 20

department: Institute for Stem Cell Biology

institute: RWTH Aachen University Medical School


title: Hexokinase 2 and lactate mediated gene expression via histone H3 lysine 18 lactylation (CHO CUT&Tag)

first: Nissim

last: Hay

summary: Hexokinase catalyzes the first committed step in glucose metabolism by phosphorylating glucose to produce glucose-6-phosphate. Highly glycolytic proliferating cells such as cancer cells take advantage of HK2 expression to accelerate glucose metabolism even in the presence of oxygen. This acceleration not only provides sufficient glycolytic intermediates to support the anabolic demands of the cells but also inevitably accompanies increased formation of metabolic end products such as lactate. Currently, the effect of lactate caused by HK2-mediated metabolic alteration is largely unknown. A recent study found that lactate plays a role in an epigenetic alteration known as histone lactylation. Here, using RNA-seq and CUT&Tag chromatin profiling, we study the effect of HK2 and lactate on gene expression via histone lactylation (H3K18la).

overall_design: Genomic occupancy profiling of H3K18la and H3K18ac for hexokinase null (EV), wildtype HK2 (WTHK2), kinase-dead mutant HK2 (SAHK2) expressing cells.


title: Hexokinase 2 and lactate mediated gene expression via histone H3 lysine 18 lactylation (CHO RNA-Seq)

overall_design: Comparative gene expression profiling analysis of RNA-seq data for hexokinase null (EV), wildtype HK2 (WTHK2) expressing cells, and EV with 10 mM lactate treatment (LAC) for 24 h.

first: Hyunsoo

last: Rho