title: The expression data of wt, nlp6, nlp7, and nlp6nlp7

city: Taipei

first: Yi-Fang

last: Tsay

address: 128 Academia Rd, Sec. 2, Nankang

department: Institute of Molecular Biology

country: Taiwan

institute: Academia Sinica

summary:
Transciptome profileing analysis of wt, nlp6, nlp7 and nlp6nlp7 before nitrate treatment and after 35 minutes of 25mM nitrate induction. NLP7 (NIN-LIKE-PROTEIN 7)  (NLP7) is the major transcriptional factor responsible for the primary nitrate response (PNR), but the role of its closest homologue , NLP6, in nitrogen signaling and the interplay between NLP6 and NLP7 remain to be elucidated. In this study, we show that, like NLP7, nuclear localization of NLP6 via a nuclear retention mechanism is nitrate-dependent, but nucleocytosolic shuttling of both NLP6 and NLP7 is independent of each other. Compared to single mutants, the nlp6 nlp7 double mutant displays a synergistic growth retardation phenotype in response to nitrate. The transcriptome analysis of primary nitrate response (the PNR) showed that NLP6 and NLP7 govern ~50% of nitrate-induced genes, with cluster analysis highlighting two distinct patterns. In the A1 cluster, NLP7 plays the major role, whereas in the A2 cluster, NLP6 and NLP7 are partially functionally redundant. Interestingly, comparing the growth phenotype and PNR under high and low nitrate conditions demonstrated that NLP6 and NLP7 exert a more dominant role in the response to high nitrate. Apart from nitrate signaling, NLP6 and NLP7 also participated in ammonium conditions . Growth phenotypes and transcriptome data uncovered revealed that NLP6 and NLP7 are completely functionally redundant, and may acting as repressors in response to ammonium. Other NLP family members also participated in the PNR, with NLP2 and NLP7 acting as broader regulators and NLP4, -5, -6, and -8 regulating PNR in a gene-dependent manner. Thus, our findings indicate that multiple modes of interplay exist between NLP6 and NLP7 that, which differ depending on nitrogen sources and gene clusters.
To determine the genome-wide contributions of NLP7 and NLP6 to the overall nitrate response, we performed a transcriptomics analysis on nlp6, nlp7 and nlp6nlp7  for comparison to wild-type (Col-0) plants. A similar approach has been applied previously for NLP7 (Marchive et al., 2013), but not for NLP6. We collected root tissue before nitrate induction and 35 min after nitrate induction and assessed alterations in gene expression.

overall_design: We analyzed the genes expression level of Col-0, nlp6 nlp7 and nlp6nlp7 using Affymetrix ATH1 GeneChip according to a standard Affymetrix protocol (Affymetrix Inc.). Experiments comprised three biological replicates, with ~200 seedlings for each replicate. Roots were collected at time 0 (before induction, which is under 12.5mM (NH4)2succinate for 9 days) and at 35 min (after 25 mM KNO3 induction).

first: Fu-Chiun

last: Hsu

first: Yu-Hsuan

last: Cheng


title: Whole-body gene expression atlas of an adult metazoan

status: Public on Apr 12 2023

city: Charlottesville

first: Eyleen

last: O'Rourke

state: VA

address: 90 Geldard Dr

department: Department of Biology

country: USA

institute: University of Virginia

type: Expression profiling by high throughput sequencing

summary: Animals are integrated organ systems composed of interacting cells whose structure and function are in turn defined by their active genes. Understanding what distinguishes physiological and disease states therefore requires systemic knowledge of the gene activities that define the distinct cells that make up an animal. Towards this goal, this study reports the first single-cell resolution transcriptional atlas of a fertile multicellular organism: Caenorhabditis elegans.

overall_design: We dissociated young adult N2 wild type C. elegans into single cells and analyzed them by scRNA-Seq

first: Abbas

last: Ghaddar

first: Erick

last: Armingol

first: Chau

last: Huynh

first: Louis

last: Gevirztman

first: Nathan

middle: E

last: Lewis

first: Robert

last: Waterston

middle: J


title: The Mediator complex regulates enhancer-promoter interactions [Micro-Capture-C]

status: Public on Apr 13 2023

city: Goettingen

first: Shyam

last: Ramasamy

address: Am Fassberg 11

department: Genome Organization and Regulation

country: Germany

institute: Max Planck Insitute for Multidisciplinary Sciences

summary: Enhancer-mediated gene activation generally requires physical proximity between enhancers and their target gene promoters. However, the molecular mechanisms by which interactions between enhancers and promoters are formed are not well understood. Here, we investigate the function of the Mediator complex in the regulation of enhancer-promoter interactions, by combining rapid protein depletion and high-resolution MNase-based chromosome conformation capture approaches. We show that depletion of Mediator leads to reduced enhancer-promoter interaction frequencies, which are associated with a strong decrease in gene expression. In addition, we find increased interactions between CTCF-binding sites upon Mediator depletion. These changes in chromatin architecture are associated with a re-distribution of the Cohesin complex on chromatin and a reduction in Cohesin occupancy specifically at enhancers. Our results indicate that enhancer-promoter interactions are dependent on an interplay between the Mediator and Cohesin complexes and provide new insights into the molecular mechanisms by which communication between enhancers and promoters is regulated.

overall_design: Micro-Capture-C (MCC). Details are described in the Methods section of the paper. Experiments were performed for multiple technical replicates (DMSO - 14 replicates, dTAG - 12 replicates and Triptolide - 18 replicates) that were derived from three biologically independent samples per experimental condition.

first: A

middle: M

last: Oudelaar


title: Chemoproteomic Mapping of Glycolytic Targetome in Cancer Cells

city: NANJING

first: zhang

last: hanqing

address: tong jia xiang 24

country: China

institute: china pharmaceutical university

summary: Hyperactivated glycolysis is a metabolic hallmark of most cancer cells. Although sporadic information has revealed that glycolytic metabolites possess non-metabolic functions as signaling molecules, it remains largely elusive how these metabolites interact with and functionally regulate their binding targets. Here we introduce a Target Responsive Accessibility Profiling (TRAP) approach that measures ligand binding-induced accessibility changes for target identification through globally labeling reactive proteinaceous lysines. With TRAP, we mapped 913 responsive target candidates and 2,487 interactions for 10 major glycolytic metabolites in a model cancer cell line. The wide targetome depicted by TRAP unveils diverse regulatory modalities of glycolytic metabolites involving direct perturbation of carbohydrate metabolism enzymes, intervention of an orphan transcriptional protein’s activity, and modulation of targetome-level acetylation. These results deepen our understanding of how glycolysis orchestrates signaling pathways in cancer cells in support of their survival and inspire future exploitation of the glycolytic targetome for cancer therapy development.

overall_design: We performed RNA sequencing to understand lactate administration would influence transcriptional activity of TRIM28 and induce transcript level changes in a TRIM28-dependent manner

first: Yang

last: Tian

first: Ning

last: Wan

first: Hanqing

last: Zhang

first: Chang

last: Shao


title: Single-cell multi-omics of mitochondrial DNA disorders reveals dynamics of purifying selection across human immune cells

city: New York

first: Caleb

last: Lareau

state: New York

address: 417 E 68th St, Zuckerman - ZRC 1132

institute: Memorial Sloan Kettering

type: Genome binding/occupancy profiling by high throughput sequencing

summary: This SuperSeries is composed of the SubSeries listed below to reproduce the data and analyses from Lareau et al. 2023 Nature Genetics

overall_design: Refer to individual Series


title: Dual gene activating and repressive functions of TET1 in germ layer lineage bifurcation distinguished by genomic context and dependence on 5-methylcytosine oxidation [CUT&RUN]

city: Leuven

first: Bernard

middle: Klaas

last: van der Veer

state: Vlaams-Brabant

address: Herestraat 49 - box 804

department: Department of Development and Regeneration

country: Belgium

institute: KU Leuven

summary: Gastrulation initiates when the epiblast differentiates into either definitive ectoderm or primitive streak. During the lineage bifurcation, the DNA dioxygenase TET1 plays dual roles in both transcriptional activation and repression, but how it exerts this bipartite control via 5-methylcytosine (5mC) oxidation-dependent and independent activities remains unclear. Here, we perform a monolayer differentiation of mouse embryonic stem cells (ESCs) into neuronal precursors to define at single-cell resolution how Tet1-/- cells undergo a lineage switch to primitive streak and subsequently form mesoderm and endoderm. We identify the Wnt repressor Tcf7l1 as a direct target of TET1 that controls a signaling cascade of Wnt/β-catenin upstream of Nodal. Disrupting the endogenous catalytic site of Tet1 in ESCs contributes to activation of Nodal and subsequently Wnt/β-catenin signaling, promoting tri-lineage differentiation into ectoderm, mesoderm and endoderm. Catalytically dead TET1 is sufficient in sustaining open neuroectoderm enhancers, by which gene expression is uncoupled from enhancer DNA demethylation, and indirectly keeping primitive streak enhancers inaccessible to Wnt regulators and effectors. DNA hypermethylation caused by TET1 catalytic dysfunction instead promotes precocious primitive streak gene activation when associated with CpG islands overlapping bivalent gene promotors. Moreover, hypermethylated regions amplify in numbers in the absence of TET1 through differentiation to affect genes associated with neurological functions. Our results reveal two-way safeguarding activities of TET1 separable by genomic features, where at CpG-poor distal enhancers TET1 maintains accessible chromatin permissive for neural fate independently of 5mC oxidation; at CpG-rich bivalent promoters it prevents premature gene activation inducing alternative fates by harnessing 5mC oxidation in Polycomb gene repression.

overall_design: WT, Tet1 MUT and Tet1 KO mESCs were differentiated to antNPCs.

middle: K


title: Dual gene activating and repressive functions of TET1 in germ layer lineage bifurcation distinguished by genomic context and dependence on 5-methylcytosine oxidation [RNAseq]


title: Expression of retrotransposons contributes to aging in Drosophila

city: Bronx

first: Blair

middle: Kathleen

last: Schneider

address: 1300 Morris Park Ave., 809 Ullmann

department: Genetics

institute: Albert Einstein College of Medicine

summary: We aimed to elucidate the mechanism by which retrotransposon (RT) knockdown extends female lifespan in drosophila. To do so we compared two, single RT (412 and Roo) shRNA knockdowns (KD) to a scrambled hairpin control.

overall_design: We performed paired end RNA-sequencing on pooled thoraces from day 20 old female flies (F1-3) in triplicate. We used a scramble (scr) hairpin as a control to our 412 and Roo shRNA ubiqutious (Act5C) knockdown animals

first: Julie

last: Secombe


title: Dual gene activating and repressive functions of TET1 in germ layer lineage bifurcation distinguished by genomic context and dependence on 5-methylcytosine oxidation [ATACseq 2]


title: Dual gene activating and repressive functions of TET1 in germ layer lineage bifurcation distinguished by genomic context and dependence on 5-methylcytosine oxidation [ATACseq]


title: Dual gene activating and repressive functions of TET1 in germ layer lineage bifurcation distinguished by genomic context and dependence on 5-methylcytosine oxidation [10x scRNAseq]


title: IL15 Deficiency in Rats

city: London

first: Stephen

last: Renaud

state: Ontario

address: 1151 Richmond St, MSB428

country: Canada

institute: University of Western Ontario

summary: NK cells are a major immune cell in the uterus during pregnancy. IL15 is required for the maturation of these cells, therefore a mutation in the IL15 gene results in a lack of NK cells. In this study, implantation sites were collected from gestational day 9.5 pregnant IL15 deficient or WT Sprague-Dawley Holtzman rats. RNA was collected, and a Clariom S microarray was performed to elucidate differences in gene expression at the developing placenta and uterus between mutants and WT dams.

overall_design: WT dams and IL15Δ/Δ dams were mated with a WT male. On GD 9.5 implantation sites (containing developing placenta and embryo) were collected and RNA extracted using a TRIzol-RNeasy column hybrid protocol including on-column DNase I digestion, and purified. Each sample represents RNA pooled from 3 implantation sites a single dam. Normalization and output file generation was performed with Affymetrix Gene Expression Console (Thermo) using the Robust Multi-Chip Analysis (RMA) algorithm [RMA-Gene-full].

first: Kelly

last: Baines


title: Cancer-associated fibroblasts in T1 colorectal cancer

status: Public on Apr 11 2023

city: Leiden

first: Hailiang

last: Mei

address: Einthovenweg 20

department: Sequencing Analysis Support Core

country: Netherlands

institute: Leiden University Medical Center

summary: Characterization of gene expression profiles of primary human patient-derived T1 colorectal cancer-associated fibroblasts (T1CAFs) and patient-matched normal fibroblasts (NFs) by bulk RNA sequencing

overall_design: Bulk RNA sequencing of 10 primary patient-matched NF-T1CAF pairs

first: Hao

last: Dang

first: Tom

last: Harryvan

first: Szymon

last: Kielbasa

first: James

middle: C

last: Hardwick

first: Jurjen

last: Boonstra

first: Noel

middle: F

last: de Miranda

first: Lukas

last: Hawinkels


title: Dual gene activating and repressive functions of TET1 in germ layer lineage bifurcation distinguished by genomic context and dependence on 5-methylcytosine oxidation

type: Methylation profiling by high throughput sequencing

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


title: MiR-29a-deficiency Causes Thickening of the Basilar Membrane and Age-related Hearing Loss by Upregulating Collagen Ⅳ and Laminin

city: Yantai

first: peng

last: ma

address: Marina Road Street

institute: Binzhou Medical University

summary: Age-related hearing loss (ARHL) is the most common sensory degenerative disease and can significantly impact the quality of life in elderly people. A previous study using GeneChip miRNA microarray assays showed that the expression of miR-29a changes with age, however, its role in hearing loss is still unclear. In this study, we characterized the cochlear phenotype of miR-29a knockout (miR-29a-/-) mice and found that miR-29a-deficient mice had a rapid progressive elevation of the hearing threshold from 2 to 5 months of age compared with littermate controls as measured by the auditory brainstem response. Stereocilia degeneration, hair cell loss and abnormal stria vascularis were observed in miR-29a-/- mice at 4 months of age. Transcriptome sequencing results showed elevated extracellular matrix (ECM) gene expression in miR-29a-/- mice. Both GO annotation and KEGG pathway enrichment analysis revealed that the key differences were closely related to ECM. Further examination with a transmission electron microscope showed thickening of the basilar membrane in the cochlea of miR-29a-/- mice. Five Col4a genes (Col4a1-a5) and two laminin genes (Lamb2 and Lamc1) were validated as miR-29a direct targets by dual luciferase assays and miR-29a inhibition assays with a miR-29a inhibitor. Consistent with the target gene validation results, the expression of these genes was significantly increased in the cochlea of miR-29a-/- mice, as shown by RT-PCR and Western blot. These findings suggest that miR-29a plays an important role in maintaining cochlear structure and function by regulating the expression of collagen and laminin and that the disturbance of its expression could be a cause of progressive hearing loss.

overall_design: Morphological experiments reveal hearing loss. Cochlear mRNA profiles of 21 day old wild type and miR-29a-/- mice were generated by deep sequencing.

first: Peng

last: Ma


title: CD34+ HSPC  CITE-seq for day 0 and Day 1 and Day 4 under VPA

first: Tiphaine

middle: Christiane

last: Martin

address: 1470 Madison Ave

department: Oncological Sciences

institute: Icahn School of Medicine at Mount Sinai, Tisch Cancer Institute

summary: The regenerative potential of human hematopoietic stem cells (HSCs) is well established by to their ability of life-long blood cell production and cure of a wide range of hematological diseases upon transplantation. This regenerative potential depends on HSC self-renewal and the coordinated adaptation to metabolic stress conditions. This is especially critical during ex vivo culture/manipulation of HSCs that is frequently accompanied with loss of self-renewal potential resulting in stem cell exhaustion. We have previously reported that CD34+ human hematopoietic stem and progenitor cells (HSPC) can be efficiently reprogrammed and expanded to phenotypic HSCs with long-term repopulation capacity in the presence of cytokines and valproic acid (VPA). Here, we present evidence that the SIRT1-SIRT3 axis maintains the mitochondrial activity below a critical threshold by coordinating and retaining a transcriptional and metabolic landscape of human HSCs with long-term self-renewal properties during ex vivo HSC reprogramming.

overall_design: We performed CITE-sequencing to obtain measurement at the protein and transcriptome level in order to measure these data simultaneously in before (day 0) and after cytokines and valproic acid (VPA) (day 1 and day 4).

first: Luena

last: Papa

first: Christoph

last: Schaniel

first: Mansour

last: Djedaini

first: Ronald

last: Hoffman


title: Genome wide expression analysis of Caenorhabditis elegans mdt-15(tm2182) null mutants (vs. WT N2 worms)

city: Vancouver

first: Stefan

last: Taubert

state: BC

address: 950 W 28th Ave, CMMT Room 2024

department: Medical Genetics

institute: UBC

summary: MDT-15/MED15 is a subunit of the Mediator complex and is known to regulate specific gene programs. We had previously profiled gene expression in C. elegans following mdt-15 depletion by RNAi. To complement this, we also performed gene expression profiling of the mdt-15(tm2182) hypomorhic mutant to identify MDT-15 regulated genes and developmental and/or physiological gene programs.

overall_design: We grew developmentally synchronized wild-type and mutant worms to the L4 larval stage and harvested them for RNA extraction. L4 was chosen to avoid contamination from embryonic mRNA present in adult worms and to allow best comparison to previous Mediator subunit expression profiling done at the same stage. Please note that this study is an extension of the study profiled in GSE68520; animals were grown, harvested, and processed at the same time. RNA was extracted, purified and processed at the same time, and microarrays were run and analyzed at the same time. The set of wild-type samples for these one-color arrays is identical to the set used in GSE68520.

first: Rebecca

last: Barbeau

first: Chris

last: Eisley

first: Andrea

last: Barczak

first: David

last: Erle


title: Neuropilin-1 Knockout and Rescue Confirm its Role to Promote Metastasis in MDA-MB-231 Breast Cancer Cells

city: Muscat

first: Noura

middle: Salim

last: Al-Zeheimi

address: Muscat

country: Oman

institute: Sultan Qaboos University

summary: To investigate the function of Neuropilin-1 (NRP-1) in breast cancer MDA-MB-231 cells. CRISPR-Cas9 gene editing was used to knockout (KO) the NRP-1 gene in MDA-MB-231 human triple-negative breast cancer cells. Differentially expressed genes (DEGs) were determined in NRP-1 KO and parental MDA-MB-231 cells using whole transcriptome next-generation sequencing.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for MDA-MB-231 cells and its KO derivatives (NRP-1 KO).

last: Al Zeheimi

first: Sirin

middle: A

last: Adham

first: Yan

last: Gao

first: Peter

last: Greer


title: Dual gene activating and repressive functions of TET1 in germ layer lineage bifurcation distinguished by genomic context and dependence on 5-methylcytosine oxidation [oxWGBS]

first: Bernard

last: van der Veer


title: H4K16ac activates retrotransposons and contributes to their cis regulatory function [RNA-seq]

first: Pradeepa

last: Madapura

state: UK

address: 4 Newark Street, Whitechapel

country: United Kingdom

institute: Queen Mary University of London

summary: Mammalian genomes harbour a large number of transposable elements (TEs) and their remnants. Most TEs are incapable of retrotransposition, however, they have evolved as cis-regulatory elements (CREs), enabling them to recruit host-encoded factors. Understanding the contribution of TEs in the regulation of the mammalian genome is an active area of research. Here we show that the male-specific lethal (MSL) complex-mediated acetylation of histone H4 lysine 16 (H4K16ac) regulates the transcription of TEs. Furthermore, H4K16ac marked TEs function as a rich source of cis regulatory elements in the human genome, wherein H4K16ac acts by maintaining the permissive chromatin structure and promoting the transcription of these TEs.

overall_design: RNA-sequencing (RNA-Seq) for H9 hPSCs stable cell lines expressing non-targeting shRNA (control/Scr.) and MSL3 knockdown (KD, two independent shRNAs). Each sample was processed in four biological replicates.

first: Debosree

last: Pal

first: Manthan

last: Patel

first: Madapura

last: Pradeepa


title: Targeting KDM6A-KMT2D-p300 axis as a therapeutic strategy for diverse coronaviruses [RNAseq]

status: Public on Apr 10 2023

city: New Haven

first: Qin

last: Yan

email: qin.yan@yale.edu, craig.wilen@yale.edu

address: 310 Cedar St.

department: Pathology

institute: Yale University

summary: We demonstrate that the histone demethylase KDM6A promotes infection of diverse coronaviruses, including SARS-CoV-1, SARS-CoV-2, MERS-CoV and mouse hepatitis virus (MHV) in a demethylase activity-independent manner

overall_design: RNA-seq of Vero-E6 cell line

first: Wesley

middle: L

last: Cai

first: Wei

last: Jin

first: Wilen

last: Craig


title: linc-mipep and linc-wrb encode micropeptides that regulate chromatin accessibility in vertebrate-specific neural cells

first: Antonio

last: Giraldez

state: CT

address: 333 Cedar street

institute: Yale

summary: Thousands of long intergenic non-coding RNAs (lincRNAs) are transcribed throughout the vertebrate genome. A subset of lincRNAs enriched in developing brains have recently been found to contain cryptic open reading frames and are speculated to encode micropeptides. However, systematic identification and functional assessment of these transcripts have been hindered by technical challenges caused by their small size. Here we show that two putative lincRNAs (linc-mipep, also called lnc-rps25, and linc-wrb) encode micropeptides with homology to the vertebrate-specific chromatin architectural protein, Hmgn1, and demonstrate that they are required for development of vertebrate-specific brain cell types. Specifically, we show that NMDA receptor-mediated pathways are dysregulated in zebrafish lacking these micropeptides and that their loss preferentially alters the gene regulatory networks that establish cerebellar cells and oligodendrocytes - evolutionarily newer cell types that develop postnatally in humans. These findings reveal a key missing link in the evolution of vertebrate brain cell development and illustrate a genetic basis for how some neural cell types are more susceptible to chromatin disruptions, with implications for neurodevelopmental disorders and disease.

overall_design: Flash-frozen pooled brains were prepared based on Protocol CG000366 – Rev D (Protocol 2) from 10x Genomics (available at https://www.10xgenomics.com/support/single-cell-multiome-atac-plus-gene-expression/documentation/steps/sample-prep/nuclei-isolation-from-embryonic-mouse-brain-tissue-for-single-cell-multiome-atac-plus-gene-expression-sequencing). It is critical to keep samples cold and/or on ice for all steps. Briefly, all samples were processed identically and simultaneously, to minimize batch effects. Chilled 0.1X Lysis Buffer (500μl) was immediately added to frozen samples, and samples were homogenized using a glass dounce tissue grinder with glass pestle. Samples were incubated on ice for 5 minutes, gently pipetted 10x, then incubated again for 5 minutes. Chilled Wash Buffer (500μl) was gently added to samples. After pipetting the mix 5x, the samples were passed through 70μm-porosity Flowmi tips into new ice-cold low-bind 1.5ml tubes. Each suspension was subsequently passed through a 40μm-porosity Flowmi tip into a new ice-cold low-bind 1.5ml tube. Samples were centrifuged at 500 rcf 5 minutes at 4°C. The supernatant was gently removed, without disturbing the nuclei pellet. Chilled Wash Buffer (1ml) was added, and the nuclei were gently resuspended 5x. This wash and resuspension step was repeated one more time. On the final step, nuclei were resuspended in Diluted Nuclei Buffer. Quality and number of nuclei (as assessed by >90% Trypan Blue staining) for each sample was assessed using a hemocytometer, and were immediately used for tagmentation step using the 10x Genomics platform.

first: Valerie

last: Tornini

first: Liyun

last: Miao

first: Ho-Joon

last: Lee

first: Timothy

last: Gerson

first: Sarah

last: Dube

first: Valeria

last: Schmidt

first: Francois

last: Kroll

first: Yin

last: Tang

first: Katherine

last: Du

first: Manik

last: Kuchroo

first: Charles

last: Vejnar

first: Ariel

last: Bazzini

first: Smita

last: Krishnaswamy

first: Jason

last: Rihel


title: Transcriptome profiling of phagocytic or non-phagocytic microglia from Ifngr1 cKO mice.

city: Boston

first: Zhuoran

last: Yin

state: Massachusetts

address: 60 Fenwood Road

department: Neurology

institute: Brigham and Women's Hospital

summary: Investigate the effect of depleting Ifngr1 in microglia on their response to acute neurodegeneration

overall_design: Apoptotic neurons were stereotaxically injected into the cortex and hippocampus of WT and Ifngr1 cKO mice. After 16 hours, phagocytic/non-phagocytic microglia were sorted for RNAseq.

last: Zhuoran

first: Herron

last: Shawn

first: Sebastian

last: Silveira

first: Christian

last: Gauthier

first: Dania

last: Mallah

first: Kilian

last: Kleemann

first: Caterina

last: Cheng

first: Milica

last: Margeta

first: Kristen

last: Pitts

first: Jen-Li

last: Barry

first: Hannah

last: Shorey

first: Wesley

last: Brandao

first: Ana

last: Durao

first: Jean

last: Christophe Delpech

first: Charlotte

last: Madore

first: Mark

last: Jedrychowski

first: Amrendra

last: Ajay

first: Gopal

last: Murugaiyan

first: Samuel

middle: W

last: Hersh

first: Seiko

last: Ikezu

first: Tsuneya

first: Oleg

last: Butovsky


title: Effect of blocking IFNgamma on the transcriptomics of phagocytic or non-phagocytic microglia from miR-155 cKO mice.

first: Zhuoran

last: Yin

summary: Investigate whether miR-155 deletion activates microglia through interferon-dependent mechanism.

overall_design: WT and miR-155 cKO mice were treated with IFNgamma blocker or IgG control i.p. and intracerebral injection. Apoptotic neurons were stereotaxically injected into the cortex and hippocampus. After 16 hours, phagocytic/non-phagocytic microglia were sorted for RNAseq.

first: Yin

last: Zhuoran

first: Herron

last: Shawn

first: Sebastian

last: Silveira

first: Christian

last: Gauthier

first: Dania

last: Mallah

first: Kilian


title: Gene expression profile at single cell level of CD11b+/CD45+ myeloid cells from the brain (scRNA_male)

summary: We used single cell RNA sequencing (scRNA-seq) to analyze the subsets of myeloid cells affected by deletion of miR-155 in microglia.

overall_design: CD11b+/CD45+ myeloid cells were isolated by Fluorescence-activated cell sorting (FACS) and analyzed using scRNAseq.

last: Kleemann

first: Caterina

last: Cheng

first: Milica

last: Margeta

first: Kristen

last: Pitts

first: Jen-Li

last: Barry

first: Hannah

last: Shorey

first: Wesley

first: Ana


title: Gene expression profiles between the palbociclib monotherpy versus palbociclib alternating with ICI182,780 treatment in breast cancer cells at 10 weeks.

city: Washington

first: Ayesha

last: Shajahan-Haq

state: District of Columbia

address: 3970 Reservoir Road, NW, NBR E404

institute: Georgetown University Medical Center, Lombardi Comprehensive Cancer Center

summary: We examined differential gene expression profiles between MCF7 human breast cancer cells treated with vehicle alone (control), palbociclib monotherapy, palbociclib alternating with ICI182,780 treatment, and palbociclib resistant cells. For this, we used gene microarray expression profiling to identify significantly changed genes under different treatment conditions.

overall_design: Gene expression signatures for four conditions of MCF7 cells were: (1) parental cells; vehicle treated (0.04% DMSO); control, (2) cells treated with palbociclib monotherapy at 750nM for 10 weeks, (3) alternating treatment  with palbociclib at 750nM for 1 weeek  and then switched to ICI182,780 at 750nM for 1 week, for 10 weeks total, and (4) cells cultured for >6 months in palbociclib (500nM). Four biological replicated were collected for each treatment condition.


title: Hypoxia extends lifespan and neurological function in a mouse model of aging

first: Vamsi

last: Mootha

state: MA

address: 185 Cambridge Street 6th Floor

institute: Massachusetts General Hospital

summary:
To investigate the effect of chronic continuous hypoxia (11%) oxygen on lifespan and neurological function in Ercc1 ∆/- mice.
We performed bulk RNA-seq of the cerebellum from mice that were Ercc1 ∆/- normoxia, Ercc1 ∆/- hypoxia, wildtype normoxia, wildtype hypoxia.

overall_design: Differential gene expression analysis of cerebellar bulk RNA-seq from the 4 groups above, taken at the time when the normoxic mutant of a birth cohort reached euthanasia criteria, ages 15-19 weeks.

first: Robert

middle: S

last: Rogers

first: Hong

last: Wang

first: Timothy

last: Durham

first: Jonathan

last: Stefely

first: Norah

last: Owiti

first: Andrew

last: Markhard

first: Lev

last: Sandler

first: Tsz-Leung

last: To


title: Charting the impact of supraphysiologic MDM2 on gene regulatory circuits in sarcoma

first: Salvador

last: Casani

address: 450 Brookline Avenue

department: Cancer Biology

institute: Dana Farber Cancer Institute

summary: Amplification of the ubiquitin ligase MDM2 is a common mechanism of P53 inactivation across human tumors. Here we investigated the impact of supraphysiologic MDM2 expression on chromatin topology, gene expression and cellular phenotypes in liposarcoma tumors and models. We identified three independent gene regulatory circuits that predominate in aggressive, dedifferentiated tumors. RUNX and AP-1 family transcription factors bind a shared set of enhancers associated with mesenchymal lineage genes that underlie the developmental state of these tumors. P53 and MDM2 co-occupy enhancers and promoters associated with P53 pathways. When highly expressed, MDM2 also binds an additional set of P53-independent promoters that engage in multi-way physical interactions and regulate genes involved in cell division, RNA splicing, translation and stress responses. We find that liposarcoma cells with low to moderate MDM2 levels are sensitive to P53 inhibitors and their combination with pro-apoptotic agents, but that cells with very high-level MDM2 amplification and expression resist treatment. Hence, heterogeneity of MDM2 expression between and within tumors may compromise therapeutic regimens. In conclusion, we distinguished P53-dependent and P53-independent circuits in MDM2 copy number amplified tumors whose interplay has implications for targeted therapy.

overall_design:
Samples from liposarcoma. ChIP-Seq and RNA-Seq data was extracted from Non Treated (NT) cells, or cells treated with HDM201.
NOTE FROM SUBMITTER: The tumor data is still pending IRB. The raw data will be submitted to the dbGAP

first: Samantha

last: Bevill

last: Casani-Galdon

first: Chadi

last: El Farran

first: Eli

middle: G

last: Cytrynbaum

first: Kevin

last: Macias

first: Sylvie

last: Oldeman

first: Kayla

last: Oliveira

first: Esmat

last: Hegazi

first: George

middle: D

last: Demetri

first: Sonia

last: Cohen

first: John

middle: T

last: Mullen

first: Nicolò

last: Riggi

first: Sarah

last: Johnstone

first: Bradley

last: Bernstein


title: SARS-CoV-2 infection and recovery in children: Distinct T cell responses in MIS-C compared to COVID-19

first: Donna

last: Farber

state: NY

address: 701 West 168th Street, HHSC 1313

institute: Columbia University

summary: While SARS-CoV-2 infection results in a mild disease course for most children, severe manifestations including respiratory failure and a Multisystem Inflammatory Syndrome in Children (MIS-C) can develop in rare cases. Here, we present a longitudinal immune profiling study of children who developed SARS-CoV-2-associated MIS-C during acute disease and following recovery, relative to children with more typical clinical manifestations of SARS-CoV-2 infection. We demonstrate distinct immune parameters associated with acute MIS-C including elevated levels of pro-inflammatory cytokines in circulation not observed in children with acute COVID-19 (non-MIS-C), aberrant expression of activation and tissue residency signatures by T cells, and expansion of T cells bearing specific TCR chains. Importantly, markers of tissue-derived T cells correlated with clinical measurements of cardiac damage suggesting key roles for T cells in the disease process. While these T cell alterations and the accompanying systemic inflammation resolved during recovery, children with prior MIS-C generated increased frequencies of functionally robust virus-specific T cells compared to children who recovered from mild disease, while both cohorts generated protective, long-lasting SARS-CoV-2-specific antibodies. Together our results reveal that the T cell response in acute MIS-C contributed to disease pathology, while also generating robust memory responses for future immune protection.

overall_design: RNA-seq profiling study of children who developed SARS-CoV-2-associated MIS-C during acute disease and following recovery, relative to children with more typical clinical manifestations of SARS-CoV-2 infection.

first: Ksenia

last: Rybkina

first: Joseph

middle: N

last: Bell

first: Teddy

last: Wohlbold

first: Marissa

last: Bradley

first: Marika

last: Scafuro

first: Wenzhao

last: Meng

first: Brett

middle: R

last: Anderson

first: Rachel

last: Weller

first: Joshua

last: Milner

first: Anne

last: Moscona

first: Matteo

last: Porotto

first: Eline

last: Luning Prak

first: Thomas

last: Connors


title: Programmed cell death-1 receptor mediated regulation of Tbet+NK1.1- Innate Lymphoid Cells Within the Tumor Microenvironment

city: Newcastle Upon Tyne

first: Jing Xuan

last: Lim

address: Framlington Place

department: Bioscience Institute

institute: Newcastle University

summary: Innate Lymphoid Cells (ILCs) play a key role in tissue mediated immunity and can be controlled by co-receptor signaling. Here we define a subset of ILCs that are Tbet+NK1.1- and are present within the tumor microenvironment (TME). We show programmed death-1 receptor (PD-1) expression on ILCs within TME is found in Tbet+NK1.1-ILCs. PD-1 significantly controlled the proliferation and function of Tbet+NK1.1-ILCs in multiple murine and human tumors. We found tumor derived lactate enhanced PD-1 expression on Tbet+NK1.1-ILCs within the TME, which resulted in dampened mTOR signaling along with increased fatty acid uptake. In line with these metabolic changes, PD-1 deficient Tbet+NK1.1-ILCs expressed significantly increased IFNg,  granzyme B and K. Furthermore, PD-1 deficient Tbet+NK1.1- ILCs contributed towards diminished tumor growth in an experimental murine model of melanoma. These data demonstrate that PD-1 can regulate anti-tumor responses of Tbet+NK1.1-ILCs within the tumor microenvironment.

overall_design: Tumour Infiltrating Lymphocytes were isolated using ficol density gradient centrifugation. Cells were then debulked with Thy1 microbeads and then stained with sample tags and Abseq antibodies along with Lineage cocktail antibodies and Thy1. Lin-Thy1+ cells were enriched through flow sorting and then loaded onto two cartridges and subjected to Rhapsody analysis.


title: Differential gene expression in WT and AMPK deficient CD4 T cells

city: Gyeongsan

first: Jae

middle: Hoon

last: Chang

state: Gyeongsangbuk

address: 280, Daehak-ro

department: Pharmacy

country: South Korea

institute: Yeungnam University

summary: The purpose of the study was to compare transcriptome profiling in WT and AMPK deficient CD4 T cells.

overall_design: CD4 T cells were isolated from the WT and AMPK knockout C57BL/6 mice. These isolated cells were done resting under RPMI-1640 for 1 hour and stimulated with anti-CD3/CD28 (2 ug/ml) for 24 hours. Thus prepared samples were collected for RNA-seq.

first: Mahesh

last: Pandit

first: Jae-Hoon


title: RNA-sequencing of the CD4 T cell culture under the various nutrients conditioned media.

summary: The purpose of the study are to compare and observe differential gene expressions in CD4 T cells when cultured under various nutrient conditions.

overall_design: CD4 T cells were isolated from the spleen and lymphnodes of C57BL/6 mice. Isolated murine CD4 T cell were stimulated with anti-CD3/CD28 antibody (2 µg/ml) under various conditioned media. Complete Media (RPMI-1640 supplemented with 10% FBS), DM (Dialyzed media) and DM + Met (Dialyzed media supplemented with 30 µM methionine). Followed by, in vitro cultured for 72 hours.


title: Epigenetic reactivation of tumor suppressor genes with CRISPRa technologies as precision therapy for hepatocellular carcinoma

city: Clayton

first: Joe

last: Cursons

state: VIC

address: 19 Innovation Walk

department: Biomedicine Discovery Institute

country: Australia

institute: Monash University

summary: Chromatin plasticity can be exploited therapeutically for the selective manipulation of the epigenetic state, to ultimately restore transcriptional aberrations driving tumorigenesis. Abnormal epigenetic silencing of tumor suppressors genes (TSGs) is one of the carcinogenesis features in hepatocellular carcinoma (HCC). We computationally identified a panel of 12 putative TSGs exhibiting an anti-correlation between transcript abundance and promoter DNA methylation, while harbouring no genetic alterations. Silencing of at least one of these TSGs was bioinformatically predicted for any given HCC sample case, and encompassing all the subtypes of HCC, making it possible the targeting of any patient in a personalized manner. To specifically reactivate one or more TSGs of this panel we produced CRISPR activators (CRISPRa) multiplexing systems tailored to representative HCC lines. Unlike epigenetic drugs, which lack locus-selectivity, CRISPRa systems enabled a potent and precise (“at will”) reactivation of up to 4 TSG simultaneously. We show that reactivation of HHIP, MT1M, PZP, and TTC36 TSGs in Hep3B cells led to a functional decrease in cell viability, cell proliferation, and cell migration. By combining different effector domains, we identified a toolbox of epigenetic effectors and gRNAs for the functional interrogation and targeting of putative TSGs silenced in liver cancer patients. We outlined a highly personalized precision oncology approach for the treatment of aggressive forms of HCC.

overall_design:
Infinium MethylationEPIC (850K) BeadChip data for Hep3B cells with a SpdCas9-VPR MS2-MCP-p65-HSF1 construct and no targeting gRNA (4 replicates), Hep3B cells with a SpdCas9-VPR MS2-MCP-p65-HSF1 construct and a gRNA targeting the HHIP promoter (4 replicates), Hep3B cells with a SpdCas9-TET1-CD MS2-MCP-p65-HSF1 construct and no targeting gRNA (4 replicates), and Hep3B cells with a SpdCas9-TET1-CD MS2-MCP-p65-HSF1 construct and a gRNA targeting the HHIP promoter (4 replicates).

Please note that for the SpdCas9-TET1-CD MS2-MCP-p65-HSF1 no gRNA samples, there are 3 biological replicates, one of which has two technical replicates. This is outlined in Supplementary Data S6 of the associated manuscript, but for clarity:
GSM6503183 - repL1 (Biol Rep 1)
GSM6503184 - repL2_1 (Biol Rep 2, Tech Rep 1)
GSM6503185 - repL3 (Biol Rep 3)
GSM6503186 - repL2_2 (Biol Rep 2, Tech Rep 2)

first: Sgro

last: Agustin

first: Pilar

last: Blancafort


title: Gene expression profile of Fcrls+/CD11b+/Ly6C- microglia from the brain

first: Zhuoran

last: Yin

summary: We used bulk RNA sequencing to investigate the molecular signature of microglia affected by deletion of miR-155 in microglia.

overall_design: Fcrls+/CD11b+/Ly6C- microglia were isolated by Fluorescence-activated cell sorting (FACS) and analyzed using Smartseq2 RNAseq.

first: Shawn

last: Herron

first: Sebastian

last: Silveira

first: Christian

last: Gauthier

first: Dania

last: Mallah

first: Kilian

last: Kleemann

first: Caterina

last: Cheng

first: Milica

last: Margeta

first: Kristen

middle: M

last: Pitts

first: Hannah

last: Shorey

first: Wesley

last: Brandao

first: Ana

last: Durao

first: Jean

last: Christophe Delpech

first: Charlotte

last: Madore

first: Mark

last: Jedrychowski

first: Amrendra

last: Ajay

first: Gopal

last: Murugaiyan

first: Samuel

last: Hersh

first: Seiko

last: Ikezu

first: Tsuneya

first: Oleg

last: Butovsky


title: Clinical relevance of BRCA1 promoter methylation testing in ovarian cancer patients.

city: Villejuif

first: Felix

last: Blanc-Durand

address: 114 rue Edouard Vaillant

department: Medical Oncology

country: France

institute: Institut Gustave Roussy

summary: Homologous-recombination deficiency (HRD) is closely related to PARPi benefit in ovarian cancer (OC). The capacity of BRCA1 promoter methylation to predict prognosis and HRD status remains unclear.  We aimed to correlate BRCA1 promoter methylation levels in patients with high-grade OC to HRD status and clinical behavior to assess its clinical relevance.

overall_design:
The detection of BRCA1 promoter methylation and quantification of methylation levels were performed by quantitative ddPCR methodology.
bisulfite-conversion + ddPCR

first: Etienne

last: Rouleau

first: Alexandra

last: Leary

last: Blanc Durand


title: Genome-wide mapping of H3K4me1 status in primary adipocytes isolated from wild-type (WT) and Lonp1-knockout (LONP1 AKO) mice

status: Public on Apr 08 2023

city: Nanjing

first: Zhenji

last: Gan

address: 12 Xuefu Road

department: Model Animal Research Center of Nanjing University

institute: Nanjing University

summary: Mitochondrial proteases are emerging as key regulators of mitochondrial plasticity acting both as protein quality surveillance and regulatory enzymes by performing highly regulated proteolytic reactions. It remains unclear, however, whether the regulated mitochondrial proteolysis is mechanistically linked to cell identity switch such as white-to-beige conversion of adipocytes. We found that disruption of LONP1-dependent proteolysis substantially impairs thermogenic molecular program and cellular respiration in in vitro differentiated primary adipocytes. qPCR analysis showed that expression levels of Ucp1 and other beige adipocyte thermogenic genes were remarkably downregulated in LONP1 AKO adipocytes. We took a deeper dive into the characterization of histone modifications in LONP1 AKO adipocytes using the global enhancer marker H3K4me1 ChIP-Seq. These unbiased ChIP seq data are strong evidence of that LONP1 loss results in decreased H3K4me1 on a broad array of thermogenic genes. These results collectively indicate that LONP1 is crucial to maintain proper epigenetic homeostasis and beige thermogenic gene expression.

overall_design: Using genome-wide approach, ChIPseq, to delineate active enhancer hallmark H3K4me1 on LONP1-dependent beige thermogenic genes

first: Zheng

last: Zhou


title: MAPK-induced miR-29 targets MAFG and suppresses melanoma progression

city: Tampa

first: Olga

last: Vera

state: FL

address: 12902 Magnolia Drive

department: Molecular Oncology

institute: Moffitt Cancer Center

summary: The tumor suppressive miR-29 family of microRNAs is encoded by two clusters, miR-29b1~a and miR-29b2~c, and is regulated by several oncogenic and tumor suppressive stimuli. Here we investigated whether oncogenic MAPK hyperactivation regulates miR-29 abundance and how this signaling axis impacts melanoma development. Using mouse embryonic fibroblasts and human melanocytes, we found that oncogenic MAPK signaling stimulates p53-independent and p53-dependent transcription of pri-miR-29b1~a and pri-miR-29b2~c, respectively. Expression analyses revealed that while pri-miR-29a~b1 remains elevated, pri-miR-29b2~c levels decrease during melanoma progression. Using a rapid mouse modeling platform, we showed that inactivation of miR-29 in vivo accelerates the development of frank melanomas and decreases overall survival. We identified MAFG as a relevant miR-29 target that has oncogenic potential in melanocytes and is required for growth of melanoma cells. Our findings suggest that MAPK-driven miR-29 induction constitutes a tumor suppressive barrier by targeting MAFG, which is overcome by attenuation of miR-29b2~c expression.

overall_design: The human immortalized melanocytes cell lines Hermes1, Hermes2, Hermes3A and Hermes4B were obtained from the Functional Genomics Cell Bank at St George’s, University of London, UK, and cultured in RPMI media supplemented with 10% FBS, 10ng/mL hSCF, 200nM TPA, 200pM Cholera Toxin and 10nM Endothelin-1 at 37˚C in a humidified atmosphere containing 10% CO2. SK-MEL28 cells were purchased from ATCC; WM164, WM35, WM793 and 1205Lu cells were a gift from M. Herlyn from the Wistar Collection of Melanoma cell lines. All melanoma cell lines were cultured in RPMI containing 5% FBS at 37˚C in a humidified atmosphere containing 5% CO2. Hermes1 cell line was transduced with either pLGH or BRAF-V600E expressing lentiviral constructs to mimic melanoma initiation (1). Prior to RNA isolation, all growth factors were withdrawn from media of melanocytes cell lines and melanoma cells were cultured in RPMI 10% FBS for 24 hours. RNAseq was performed to compare changes in gene expression during melanoma initiation (1) or between melanocytes and melanoma cells (2).

first: Ilah

last: Bok

first: Neel

last: Jasani

first: Koji

last: Nakamura

first: Xiaonan

last: Yu

first: Nicol

last: Mecozzi

first: Kenneth

middle: Y

last: Tsai

first: Florian

last: Karreth


title: Spic is required for the differentiation of metaphocyte progenitor cells

city: Hong Kong

first: Zilong

last: WEN

address: Rm6319, HKUST, Clear water bay, Kowloon

department: Life Science

country: Hong Kong

institute: Hong Kong University of Science and Technology

summary: Metaphocytes are tissue-resident myeloid-like cells of non-hematopoietic origin in zebrafish barrier tissues. How metaphocytes acquire myeloid cell properties from non-hematopoietic precursors during the development remains unknown. Here we show that metaphocytes are in situ generated from local progenitors guided by the ETS transcription factor Spic.  Tg(spic:EGFP) transgenic fish were found to label both metaphocytes and their progenitor cells. While spic is dispensable for the formation of the metaphocyte progenitors, it is required for the progenitors to differentiate into immature metaphocytes. To decipher the molecular basis underlying the developmental arrest of metaphocytes in spic mutants, we sorted the metaphocyte progenitor cells from the epidermis and gill of the mutants and siblings and performed bulk RNA-seq analysis. The data suggests that Spic orchestrates the development of metaphocytes by promoting the expression of myeloid-associated genes, either directly or indirectly through downstream transcription factors.

overall_design: Gene expression profiling analysis of bulk RNA-seq data for metaphocyte progenitor cells from the epidermis and gill of spic mutant fish and siblings

first: Changlong

last: Zhao

first: Yunbo

last: Li

first: Jinlin

last: Tang

first: Qiuxia

first: Xi

last: Lin

last: Wen


title: Identification of a protective microglial state mediated by miR-155 and interferon-γ signaling in a mouse model of Alzheimer's disease

first: Zhuoran

last: Yin

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

overall_design: Refer to individual Series


title: Analysis of human TAp63 regulated genes in CD4+ T cells

city: Chiba

first: Shigeru

last: Tanaka

address: 1-8-1 Inohana

department: Allergy and Clinical Immunology

country: Japan

institute: Chiba University

summary: By comparing the comprehensive gene expression profiles between human Th17 cells overexpressing TAp63 and those with TAp63 knockdown, FOXP3 was identified as one of the downregulated genes by TAp63

overall_design: Human Th17 cells were infected with retrovirus of LMP-non-silencing-IRES-Thy1.1 or LMP-hTAp63KD1-IRES-Thy1.1, and pMXs-IRES-NGFR or pMXs-hTAp63a-IRES-NGFR. Infected cells were sorted and subjected to RNA-sequence analysis.

first: Akira

last: Suto

first: Kensuke

last: Suga


title: Gene expression profile of 3D liver organoids composed of hepatocyte and Kupffer cells exposed to nanoparticles.

city: Los Angeles

first: Xia

last: Yang

state: CA

address: 610 Charles E. Young Drive East

institute: University of California, Los Angeles

summary: Single cell RNA sequencing of 3D liver spheroid exposed to vanadium pentoxide (V2O5), titanium dioxide (TiO2), or graphene oxide (GO) was used to elucidate the toxicological mechanisms of different nanoparticles.

overall_design: 3D liver organoids containing hepatocytes and Kupffer cells were cultured then exposed to 6.25 µg/mL of either vanadium pentoxide (V2O5), titanium dioxide (TiO2), or graphene oxide (GO) for 6 hrs. After 6hrs of treatment, single cell suspensions were isolated and ran using the 10x 3′ single-cell RNA-seq V3.1 protocol.

first: Graciel

last: Diamante


title: Eosinophils promote effector functions of lung ILC2s in allergic airway inflammation in mice

status: Public on Apr 07 2023

city: Scottsdale

first: Elizabeth

middle: A.

last: Jacobsen

state: AZ

address: 13400 E. Shea Blvd.

institute: Mayo Clinic

summary: We investigated reciprocal interactions between cocultures of isolated populations of eosinophils and ILC2s and found these cells promote changes in gene expression as compared to being cultured alone.

overall_design: Pure populations of eosinophils and ILC2s were obtained from C57BL/6J mice and cultured alone or cocultured for 24 hours, then flow cytometry sorted to separate populations, followed by RNA-seq and gene expression profiling


title: Pleiotrophic function of BAFF to promote a senescence-associated secretome and growth arrest

city: Baltimore

first: Supriyo

last: De

state: Maryland

address: 251 Bayview Blvd

department: Laboratory of Genetics and Genomics

institute: NIA-IRP, NIH

summary: Senescent cells release a variety of cytokines, proteases and growth factors collectively defined as the Senescent Secretory Associated Phenotype (SASP). Sustained SASP induces a pattern of chronic inflammation associated with aging and implicated in multiple age-related diseases. Here, we investigated the expression and function of the immunomodulatory cytokine BAFF (B-cell activating factor), a SASP factor, in multiple senescence models. First, we characterized BAFF production across different senescence models, including senescent human diploid fibroblasts (WI-38, IMR-90) and monocytic leukemia cells (THP-1), and tissues of mice induced to undergo senescence. We identified IRF1 (interferon response factor 1) as a transcription factor required for promoting BAFF mRNA transcription in senescence, and found that suppressing BAFF production decreased the senescent phenotype in both monocytes and fibroblasts. Importantly, the influence of BAFF on the senescence program was cell type-specific: in monocytes, BAFF promoted the early activation of NF-κB and general SASP secretion, while in fibroblasts, BAFF contributed to the production and function of TP53 (p53). Overall, BAFF silencing affected shared senescence-associated phenotypes including IL6 secretion, SA-beta-Gal staining, and γ-H2AX accumulation. We propose that BAFF is a novel biomarker of senescence and a potential target for senotherapy.

overall_design: Gene expression analysis of BAFF siRNA vs. Ctrl siRNA transfected cells in senescence.

first: Martina

last: Rossi

first: Carlos

last: Anerillas

first: Maria

last: Idda

first: Rachel

last: Munk

first: Chang

middle: H

last: Shin

first: Dimitrios

last: Tsitsipatis

first: Stefano

last: Donegà

first: Alison

middle: B

last: Herman

first: Jennifer

last: Martindale

first: Xiaoling

first: Yulan

last: Piao

first: Krystyna

last: Mazan-Mamczarz

first: Jinshui

last: Fan

first: Luigi

last: Ferrucci

first: Kotb

last: Abdelmohsen

first: Myriam

last: Gorospe


title: Inhibition of DYRK1B suppresses inflammation in allergic contact dermatitis model and Th1/Th17 immune response

city: Bangkoknoi

first: Thamrong

last: Wongchang

state: Bangkok

address: 2 Wanglang, Siriaj

department: Pharmacology

country: Thailand

summary: To investigate the inhibition effect of DYRK1B kinase function on human naïve CD4+ T cell differetionation which stimulated under Treg polarizing condition for 24 hours.

overall_design:
We then performed gene expression profiling analysis using data obtained from RNA-seq of 6 different sample.
Comparative gene expression profiling analysis of RNA-seq data for stimulated native CD4+ Tcells treated with a selective inhibitor and non-treated cells

first: Kitipong

last: Soontrapa

first: Dean

last: Thumkeo


title: Single-cell gene expression profiling of MYCN-amplified neuroblastoma cells following CRISPR activation of ferroptosis regulators

city: Heidelberg

first: Umut

last: Yildiz

address: Meyerhofstraße 1

institute: European Molecular Biology Laboratory

summary: Understanding the operational molecular, and metabolic networks that determine the balance between pro- and anti-ferroptotic regulatory pathways could unravel unique vulnerabilities to be exploited for cancer therapy. Using genome-wide and single-cell CRISPR activation screens, we identify the selenoprotein P (SELENOP) receptor, LRP8, as a key determinant protecting MYCN-amplified neuroblastoma cells from ferroptosis in vitro and in orthotopic neuroblastoma mouse models. Specifically, the exquisite dependency on LRP8-mediated selenocysteine import is caused by the failure of MYCN-amplified cells to efficiently utilize alternative forms of selenium/selenocysteine based uptake necessary for selenoprotein biosynthesis. Increased activity of one of such transporters, SLC7A11, in MYCN-amplified cells leads to cysteine overload, progressive mitochondrial decline and impaired proliferation. These data reveal in LRP8 a targetable, and specific vulnerability of MYCN-amplified neuroblastoma cells and disclose a yet-unaccounted mechanism for selective ferroptosis induction that has the potential to become an important therapeutic entry point for MYCN-amplified neuroblastoma.

overall_design: The CROP-seq experiment was performed in human SK-N-DZ cells stably expressing dCas9-VP64 and p65-HSF1. Cells were transduced with a library of CRISPR activation gRNAs targeting 36 hits derived from the primary ferroptosis resistance screens with two independent gRNAs. After puromycin selection and recovery, two lanes  of the 10X platform were used to partition individual cells into gel bead-in-emulsions (GEMs) and single-cell libraries were constructed.

first: Hamed

last: Alborzinia

first: Zhiyi

last: Chen

first: Andreas

last: Trumpp

first: José

middle: P

last: Friedmann Angeli


title: Peripheral Microglial Process TRAP shows evidence of new protein synthesis at perisynaptic and phagocytic structures

city: St Louis

first: Joseph

last: Dougherty

state: Missouri

address: 4566 Scott Ave, box 8232

department: Genetics and Psychiatry

institute: Washington University

summary: We utilize Peripheral Microglial Process TRAP to identify mRNA transcripts that are enriched within the peripheral processes of microglia

overall_design: Microglial TRAP and PMP-TRAP from p28 dorsal cortex and hippocampus of tamoxifen-treated and untreated CX3CR1-CreERT2 x RPL10a-GFP (fl/fl) mice

first: Michael

middle: J

last: Vasek

first: Stuart

last: Fass

first: Yating

last: Liu


title: Chronic hypoxia favours adoption to a castration-resistant cell state in prostate cancer

status: Public on Apr 06 2023

city: Toronto

first: Mathieu

last: Lupien

address: 101 College Street

institute: University Health Network

summary: Hypoxia is associated with poor prognosis in solid tumours and hypoxia has been associated with prostate cancer progression. The investigation of how chronic (CH) hypoxia affected prostate cancer show a loss in sensitivity to androgen deprivation and increased dependancy on glucose uptake in cells.

overall_design: Comparative gene expression profiling of LNCaPs to LNCaP-CH and LNCaP to LNCaP-V16A cell lines

first: Sarina

last: Cameron

first: James

last: Hawley

first: Ankita

last: Nand


title: Mechanistic connections among epithelial plasticity, skin barrier, inflammation, and whole-body metabolism [ChIP-seq]

city: Irvine

first: Morgan

last: Dragan

address: D288 Medical Sciences I

department: Biological Chemistry

institute: UC Irvine

summary: We found that epitheilial-specific Ovol1/2 iDKO mice develop spontaneous barrier defects and whole body physiological alterations. To investigate the mechanism behind these changes, we compared gene expression changes between homeostatic iDKO epidermis compared with control at "early" 25 days post injection (DPI) and at 52 DPI

overall_design: ChIP-seq with IgG and Ovol1.

first: Xing

last: Dai


title: Analysis of the Drosophila and Human DPR Elements Reveals a Distinct Human Variant Whose Specificity Can Be Enhanced by Machine Learning

city: La Jolla

first: Torrey

middle: E.

last: Rhyne

email: terhyne@ucsd.edu, jkadonaga@ucsd.edu

state: California

address: 9500 Gilman Dr.

department: Biological Sciences

institute: UCSD

summary: The RNA polymerase II core promoter is the site of convergence of the signals that lead to the initiation of transcription. Here, we perform a comparative analysis of the downstream core promoter region (DPR) in Drosophila and humans by using machine learning. These studies revealed a distinct human-specific version of the DPR and led to the use of the machine learning models for the identification of synthetic extreme DPR motifs with specificity for human transcription factors relative to Drosophila factors, and vice versa. More generally, machine learning models could be analogously used to design synthetic promoter elements with customized functional properties.

overall_design: HARPE and SVR analyses of the Drosophila downstream core promoter region (DPR). Its properties were compared to those of its human counterpart (Vo ngoc et al. 2020).

first: Long

last: Vo ngoc

middle: E

last: Kadonaga


title: Clonal hematopoiesis is associated with protection from Alzheimer's disease

city: Stanford

first: Julia

last: Belk

address: 450 Serra Mall

institute: Stanford University

summary: scATAC-seq datasets of brain nuclei from people with and without CHIP

first: Hind

last: Bouzid

first: Sidd

last: Jaiswal


title: Cell contractility drives mechanical memory of oral squamous cell carcinoma

first: Adam

middle: Jeffrey

last: Engler

address: 9500 Gilman Drive, MC 0412

department: Bioengineering

institute: UC San Diego

summary:
To study if non-invasive oral squamous cell carcinoma (OSCC) cells can acquire mechanical memory from stiff matrix and investigate the role of cell contractility in that process, Cal27 were conditioned (for 5 days) either by matrix stiffness or by direct modulation of cell contractility and then replated to a new niche lack of sources for the cells to learn.
Cal27 with different types of conditioning were categorized either "Learn" or "Don't Lean/Forget" based on the existence of mechanical memory.
Comparative gene expression profiling was done between "Learn" vs. "Don't Learn/Forget" Cal27 groups by RNA sequencing

overall_design:
Every group underwent 5 days of conditioning followed by a day of replating phase.
"Learn" group were prepared either by treating Cal27 with lisophosphatidic acid (LPA) or seeding cells on stiff (20 kPa polyacrylamide gel) during conditioning phase. "Don't Learn/Forget" group was prepared by seeding cells on soft gel without any treatment during conditioning phase or by inhibiting FAK signaling pathway during replating phase.
Transcriptomic difference between Cal27 groups with ("Learn") and without ("Don't Learn/Forget") memory was analyzed by identifying differentially expressed genes from comparing RNA sequencing data.

first: Paloma

last: de Campos

first: So

last: Moon

first: Bibiana

middle: F

last: Matte

first: Jesse

middle: K

last: Palcone

first: Virgilio

last: Zanella

first: Manoela

last: Martins

first: Marcelo

middle: L

last: Lamers


title: Gene expression profile at single cell level of CD11b+/CD45+ myeloid cells from the brain (scRNA-seq_female)

city: Boston

first: Zhuoran

last: Yin

address: 60 Fenwood Road

institute: Brigham and Women's Hospital

summary: We used single cell RNA sequencing (scRNA-seq) to analyze the subsets of myeloid cells affected by deletion of miR-155 in microglia.

overall_design: CD11b+/CD45+ myeloid cells were isolated by Fluorescence-activated cell sorting (FACS) and analyzed using scRNAseq.

first: Shawn

last: Herron

first: Sebastian

last: Silveira

first: Christian

last: Gauthier

first: Dania

last: Mallah

first: Kilian

last: Kleemann

first: Caterina

last: Cheng

first: Milica

middle: A

last: Margeta

first: Kristen

last: Pitts

first: Hannah

last: Shorey

first: Wesley

last: Brandao

first: Ana

last: Durao

first: Jean

last: Christophe Delpech

first: Charlotte

last: Madore

first: Mark

last: Jedrychowski

first: Amrendra

last: Ajay

first: Gopal

last: Murugaiyan

first: Samuel

middle: W

last: Hersh

first: Seiko

last: Ikezu

first: Tsuneya

first: Oleg

last: Butovsky


title: Examining the immunoepigenetic-gut microbiome axis in the context of self-esteem among Native Hawaiians and other Pacific Islanders

city: Honolulu

first: Rafael

middle: Aniu

last: Peres-David

state: Hawaii

address: 1800 East-West road

department: Anatomy

institute: University Of Hawaii

summary: Of our total study cohort (N=93),  survey scores qualified 71% of participants as having “normal” self-esteem (NSE; N=66) and those of the remaining 29% qualified as having “low” self-esteem (LSE; N=27). DNA methylation was evaluated using Infinium MethylationEPIC v2.0 BeadChip Kit  (Illumina)

overall_design: Bisulphite converted DNA from the 68 samples were hybridised to the Illumina Infinium MethylationEPIC v2.0 BeadChip Kit


title: Metaphocytes Are IL-22BP Producing Cells Regulated by ETS Transcription Factor Spic and Essential for Zebrafish Barrier Immunity

first: Zilong

last: WEN


title: The SMC like protein RecN is at the crossroads of Genotoxic Stress Responses in Escherchia coli

city: Paris

first: Olivier

last: Espeli

address: 11 place Marcelin Berthelot

department: CIRB

institute: College de France

summary: DNA damage repair (DDR) is an essential process for living organisms, it contributes to genome maintenance and evolution. DDR involves different pathways including Homologous recombination (HR), Nucleotide Excision Repair (NER) and Base excision repair (BER) for example.  The activity of each pathway is revealed in particular contexts but they also intervene concomitantly or subsequently to repair most DNA lesions. In the present study, we used two genotoxic antibiotics, mitomycin C (MMC) and Bleomycin (BLM), to decipher the interplays between these different pathways in E. coli. We demonstrated that only a small set of DDR proteins are common to the repair of the lesions induced by these two drugs. Among them, RecN, an SMC like protein, plays an important role by controlling sister chromatids dynamics and genome morphology at different steps of the repair processes. We demonstrated that RecN’s influence on sister chromatids dynamics is not equivalent during the processing of the lesions induced by the two drugs. We observed that RecN activity and stability requires a pre-processing of the MMC-induced lesions by the NER but not for BLM-induced lesions. In addition to its well-known importance to engage cells in the HR pathway to save cells presenting double strand breaks, our results show that RecN plays a major role in rescuing toxic intermediates generated by the BER pathway.

overall_design: Transposon Insertion Sequencing (TIS)

first: Adrien

last: Camus

last: Espéli


title: single-cell RNA sequencing analysis of metaphocytes and epidermal cells

summary: Metaphocytes are tissue-resident myeloid-like cells of non-hematopoietic origin in zebrafish barrier tissues. How metaphocytes acquire myeloid cell properties from non-hematopoietic precursors during the development remains unknown. Here we show that metaphocytes are in situ generated from local progenitors guided by the ETS transcription factor Spic. Tg(spic:EGFP) transgenic fish were found to label both metaphocytes and their progenitor cells. To reveal the genetic programs underlying the development of metaphocytes, we performed single-cell RNA sequencing analysis of spic:GFP+ cells in the  epidermis of adult Tg(spic:EGFP) fish. The result further supports that mature metaphocytes are generated from spic+ progenitors via an intermediate immature state and gradually obtain their myeloid characteristics during this process.

overall_design: scRNA-seq (10X genomics 3’ V3.1) analysis of FACS-sorted spic:EGFP+ and singlet cells  from the epidemris of 6-month-old Tg(spic:EGFP) fish

first: Changlong

last: Zhao

first: Yunbo

last: Li

first: Jinlin

last: Tang

first: Qiuxia

last: Zhou

first: Xi

last: Lin

last: Wen


title: Estrogen receptor alpha targeting in liver cancer and breast cancer

city: Perth

first: Zhaoyu

state: Western Australia

address: QE II, M block, 225C

department: School of Biomedical Sciences

institute: University of Western Australia

summary: This study is to identify estrogen receptor alpha targeting in liver cancer and breast cancer using RNA-Seq and ChIP-Seq and reveal the mechanisms underlying estrogen receptor alpha in the regulation of liver cancer and breast cancer.


title: A Tessellated Lymphoid Network Provides Whole-Body Antigen Surveillance in Zebrafish

city: Madison

first: Anna

last: Huttenlocher

state: WI

address: 1550 Linden Drive

department: Pediatrics

institute: University of Wisconsin School of Medicine and Public Health

summary: T cells survey the host for cognate antigen in mammals by trafficking to and searching within lymph nodes distributed throughout the body. Non-mammalian jawed vertebrates lack lymph nodes but still maintain highly diverse repertoires of T cells. How these T cells find antigen in hosts devoid of lymph nodes remains wholly unclear. In this study, we exploit the in vivo imaging amenability of the zebrafish to investigate T cell organization, trafficking, and antigen surveillance in an animal completely lacking lymph nodes. We find that T cells organize into a previously undescribed whole-body scale-associated pattern that we have named the tessellated lymphoid network (TLN). We find that the TLN harbors significantly more T cells than the gills, gut, kidney, and spleen, as well as populations of antigen-presenting cells, suggesting a central role in adaptive immune responses in fish. Within the TLN, T cells perform a highly directional ballistic streaming mode of motility, allowing them to traffic through the host in a coordinated ventral-to-dorsal loop. Local infection results in a dramatic shift in T cell motility, from rapid streaming to a slower random walk in regions proximal to the infection. Here, T cells sequentially interact with and ultimately form stable contacts with antigen-presenting cells. Finally, T cells within the TLN of infected fish show transcriptional changes consistent with TCR signaling and subsequent T cell activation. The TLN thus provides a mechanism for T cells to both traffic through the host and, in the context of an infection, effectively scan for and detect antigen, mirroring the function of lymph nodes in mammals.

overall_design: Adult zebrafish either without infection or infected by L. monocytogenes through puncture wounding were euthanized and descaled to collect associated cells at 4 days post-infection. Libraries were constructed from these cells using the Chromium Single Cell Gene Expression Solution 3’ v2 (10x Genomics) and sequenced using NovaSeq6000 (Illumina) with read lengths of 29-bp + 90-bp (Read1 + Read2).

first: Tanner

last: Robertson

first: Yiran

last: Hou

first: Simone

last: Shen

first: Julie

last: Rindy

first: John-Demian

last: Sauer

first: Huy

middle: Q

last: Dinh

first: Jonathan

last: Schrope


title: IL-6 prevents Th2 cell polarization by promoting SOCS3-dependent suppression of IL-2 signaling

city: Birmingham

first: Dave

middle: Durell

last: Hill

state: AL

address: 1720 Second Avenue South

department: Microbiology

institute: University of Alabama at Birmingham

summary: Defective interleukin-6 (IL-6) signaling has been associated with Th2 bias and elevated IgE. However, the underlying mechanism by which IL-6 may prevent the development of Th2-driven diseases remains unknown. Using a model of house-dust-mite (HDM)-induced Th2 differentiation and allergic airway inflammation, we show here that IL-6 signaling in allergen-specific T cells was required to prevent Th2 development and subsequent IgE response and allergic inflammation. Th2 cell lineage commitment required strong sustained IL-2 signaling. Importantly, we found that IL-6 turned off IL-2 signaling during early T cell activation and thus inhibited Th2 cell priming. Mechanistically, we found that IL-6-driven inhibition of IL-2 signaling in responding T cells was mediated by upregulation of suppressor of cytokine signaling 3 (SOCS3). Therapeutically, this mechanism can be mimicked by JAK1 inhibition. Collectively, our results identify an unrecognized mechanism that prevents the development of unwanted Th2 cell responses and associated diseases and outline potential preventive interventions.

overall_design: WT and Il6-/- mice were transferred with OTII TCR-transgenic CD4+ T cells and intranasally sensitized with 100 micrograms HDM extract (endotoxin content ~250 EU/mg) + 5 micrograms of LPS-free EndoFit ovalbumin. On day 3 and day 5, OTII cells were sorted from mediastinal lymph nodes and RNA-seq was performed (three replicates). The manuscript time references start at day 0 rather than day 1.

first: Holly

last: Bachus

first: Erin

last: McLaughlin

first: Cyrstal

last: Lewis

first: Amber

middle: M

last: Papillion

first: Etty

middle: N

last: Benveniste

first: D

middle: D

first: Alexander

last: Rosenberg

first: André

last: Ballesteros-Tato

first: Beatriz

last: León


title: Machine learning predictions of MHC-II specificities reveal alternative binding mode of class II epitopes

city: Lausanne

first: David

last: Gfeller

address: Rue du Bugnon 25A

department: Department of Oncology UNIL CHUV; Ludwig Institute for Cancer Research Lausanne

country: Switzerland

institute: University of Lausanne

summary: CD4 T cells from human donors were sorted with peptide-MHC multimers based on specific epitopes. For each donor the alpha and beta chains were sequenced separately

overall_design: TCR-seq of CD4+ T cells specific for various epitopes

first: Julien

last: Racle

last: Schmidt

first: Raphael

last: Genolet

first: Alexandre

last: Harari


title: Effect of lipopolysaccharide priming on the whole-genome expression of grapevine (Vitis vinifera cv Cabernet Sauvignon) during Pierce's disease

city: Davis

first: Mélanie

last: Massonnet

address: 2105 Robert Mondavi Institute-North Building

department: Viticulture and Enology

institute: University of California, Davis

summary: Transcriptome profiling by RNA-seq determined the genome-wide patterns of expression of grape genes in lipopolysaccharide-primed petioles during Pierce’s Disease.

overall_design: Potted Vitis vinifera ‘Cabernet sauvignon’ were mechanically inoculated with Xylella fastidiosa or 1X PBS (negative controls) 4 hours after priming with water or lipopolysaccharides from X. fastidiosa. Petioles were harvested at 4 h, 24 h, and 48 h post-inoculation (hpi) from the point of inoculation and 20 nodes above the point of inoculation.

first: Claudia

last: Castro

first: Nancy

last: Her

first: Biagio

last: diSalvo

first: Barbara

last: Jablonska

first: Daniel

middle: R

last: Jeske

first: Dario

last: Cantu

first: M

middle: C

last: Roper


title: Mechanistic connections among epithelial plasticity, skin barrier, inflammation, and whole-body metabolism [RNA-seq]

first: Morgan

last: Dragan

overall_design: Comparative gene expression profiling analysis of RNA-seq data for trypsin-isolated epidermis from control and iDKO mice .

first: Xing

last: Dai


title: ANGPTL3 deficiency alters the lipid profile and metabolism of cultured hepatocytes and human lipoproteins

city: Helsinki

first: Juuso

last: Taskinen

address: Tukholmankatu 8

country: Finland

institute: Minerva Foundation Institue for Medical Research

summary:
Even though the functions of ANGPTL3 in the circulation are relatively well characterized, many mechanistic questions regarding the molecular consequences of ANGPTL3 loss-of-function and protection against cardiovascular disease remain unanswered.
To help understant the molecular mechanisms exerted by ANGPTL3 deficiency, we studied the alterations of ANGPTL3 loss-of-function in hepatocytes and plasma lipid molecular species profiles.

overall_design: We characterized the impacts of ANGPTL3 depletion on the immortalized human hepatocyte (IHH) transcriptome, lipidome and human plasma lipoprotein lipidome. ANGPTL3 knock-down (KD) hepatocytes were created using an shRNA lentiviral vector. An approximately 90% KD efficiency was observed at shRNA and the KD effect was evident also at the protein level.

first: Hanna

last: Ruhanena

first: P

last: Haridas

first: Ilenia

last: Minicocci

middle: H

first: Francesco

last: Palmas

first: Alessia

last: di Costanzo

first: Laura

last: D'Erasmo

first: Jari

last: Metso

first: Jennimari

last: Partanen

first: Jesmond

last: Dalli

first: You

last: Zhou

first: Marcello

last: Arca

first: Matti

last: Jauhiainen

first: Reijo

last: Käkelä

first: Vesa

last: Olkkonen


title: Gene Expression Profile (GEP) of Cytokine-Induced Killer (CIK) cells

city: Modena

first: Rossella

last: Manfredini

address: Via Gottardi 100

department: Life Sciences

country: Italy

institute: Centre for Regenerative Medicine

summary: Cytokine Induced Killer (CIK) cells are heterogeneous ex vivo expanded T lymphocytes with mixed T-NK phenotype and endowed with a wide MHC-unrestricted antitumor activity against both solid and hematologic malignancies. CIK cells can be easily ex vivo expanded up to clinical relevant rates from circulating peripheral blood mononuclear cells (PBMC), according to a standard protocol involving timed stimulation with IFN- γ (day 0), Ab-antiCD3 OKT3 (day 1) and IL-2 (from day 1 to the end). Nowadays literature provides scanty information on cytokines, chemokines and growth factors secreted by CIK cells. A comprehensive molecular profiling of CIK cells is needed to complete the knowledge of their biologic and functional features. In this perspective we performed a gene expression profile analysis of basal PBMC and ex vivo expanded CIK cells generated from 3 patients with histologic confirmed Gastrointestinal Stromal Tumors (GIST). The principal component analysis (PCA) shows that the CIK cell samples clustered together and were clearly separated from PBMC ones. First, we checked the mRNA expression levels of secreted proteins and we found a significant upregulation of GM-CSF, IL-5, IL-13, and IFN-γ in CIK cells. Moreover, IL-1Ra, IL-1β, IL-6, IL-10, IL-15, IL-8, Eotaxin and MCP-1 are downregulated in CIK cells if compared to PBMC. Noteworthy, among differentially expressed genes (DEGs) we identified other secreted molecules that are upregulated in patient-derived CIK cells, which can contribute to their tumor killing activity: GZMA, GZMB, GZMK, PRF1, IL-32, and LTA. Furthermore, to better characterize the CIK cell phenotype, we studied the surface antigen expression. As expected, we found the downregulation of several myeloid differentiation markers (CD14, CD9, CD93, CSF2RA, CSF2RB, EPB41L3, Receptors for Fc fragments of immunoglobulins, ITGAX, MCEMP1, and TREM1) and B-cell antigens (CD19, CD24, CD79A, CXCR5, and MS4A1). Moreover, microarray data confirmed the upregulation in CIK cells of well-known surface antigens, like CD3D, CD3G, IL2Ra, IL2RG, CD226/DNAM1, ITGAL/LFA-1, KLRK1/NKG2D, NCR3/NKp30, and TRAIL/TNFSF10.  Next, in order to identify differentially expressed pathways during CIK cells maturation, we performed a functional analysis by using Ingenuity Pathway Analysis (IPA) software. Among inactivated functions in CIK cells, we found chemotaxis of myeloid cells, phagocytosis, migration of granulocytes and engulfment of leukocytes. Noteworthy, functional gene categories as proliferation of cells, cell death of lymphocytes, cell death of mononuclear leukocytes, apoptosis of B lymphocytes, and quantity of CD4+ T-lymphocytes resulted activated, in agreement with the selection and expansion of CIK cell precursors induced by in vitro treatment of PBMC. Finally, IPA analysis predicted as activated categories cytotoxicity of lymphocytes, cytotoxicity of cells, and cytotoxicity of natural killer cells. In particular, genes involved in cytotoxic mechanism of CIK cells like NCR3/NKp30, KLRK1/NKG2D, TRAIL/TNFSF10, CD226/DNAM1, CD244, CD69, CD96, GZMA, GZMB, PRF1 are differentially expressed. With this work we highlighted previously unknown secretory properties and provided for the first time a comprehensive molecular characterization of CIK cells, opening the possibility to operate on secretory level to improve CIK cell cytotoxic activity against tumors.

overall_design: Gene expression profile (GEP) was performed on total RNA derived from PBMCs and CIK cells derived from 3 patients with histologic confirmed Gastrointestinal Stromal Tumors (GIST).

first: Giulia

last: Mesiano

last: Montagner

first: Roberta

last: Zini

first: Nicoletta

last: Bianchi

first: Antonella

last: Chillemi

first: Massimo

last: Aglietta

first: Giovanni

last: Grignani

first: Ilaria

last: Lampronti

first: Fabio

last: Malavasi

first: Dario

last: Sangiolo

first: Roberto

last: Gambari

first: Davide

last: Ferrari


title: Single Cell RNA / Single Nucleus RNA Sequencing Data of Human Pancreaitc Islet

city: Duarte

first: Randy

middle: B

last: Kang

address: 1500 E Duarte Rd

department: Molecular and Cellular Endocrinology

country: USA

institute: City of Hope Beckman Research Center

summary: To investigate novel markers for snRNA seq and graft in-vivo snRNA seq.

overall_design: scRNA and integrate with snRNA, extract the DEG in each cluster, create reference and projection

first: Geming

last: Lu

first: Adolfo

last: Garcia-Ocana


title: PRC2.1 and PRC2.2 Specific Accessory Proteins Drive Recruitment of Different Forms of Canonical PRC1

city: Dublin

first: Cheng

last: Wang

address: College Green

department: Genetics

country: Ireland

institute: Smurfit Institute of Genetics, Trinity College Dublin

summary: Polycomb repressive complex 2 (PRC2) mediates H3K27me3 deposition, which is thought to recruit canonical PRC1 (cPRC1) via chromodomain-containing CBX proteins to promote stable repression of developmental genes. PRC2 forms two major subcomplexes, PRC2.1 and PRC2.2, but their specific roles remain unclear. Through genetic knockout and replacement of PRC2 subcomplex-specific subunits in naïve and primed pluripotent cells, we uncover distinct roles for PRC2.1 and PRC2.2 in mediating the recruitment of different forms of cPRC1. PRC2.1 catalyses the majority of H3K27me3 at Polycomb target genes and is sufficient to promote recruitment of CBX2/4-cPRC1, but not CBX7-cPRC1. Conversely, while PRC2.2 is poor at catalysing H3K27me3, we find that its accessory protein JARID2 is essential for recruitment of CBX7-cPRC1 and the consequent 3D chromatin interactions at Polycomb target genes. We therefore define distinct contributions of PRC2.1 and PRC2.2 specific accessory proteins to Polycomb mediated repression and uncover a new mechanism for cPRC1 recruitment.

overall_design: Refer to individual Series


title: PRC2.1 and PRC2.2 Specific Accessory Proteins Drive Recruitment of Different Forms of Canonical PRC1 [ChIP]

overall_design: Cells were collected and washed with PBS 2x, then suspended in 10 ml PBS. Next, cells were fixed with 1% formaldehyde (Sigma) with rotation for 10 minutes, followed by quenching with glycine. Fixed cells were washed with cold PBS (with proteinase inhibitors cocktails, PIC) then lysed in SDS lysis buffer (0.5% SDS, 100mM NaCl, 50mM Tris pH 8.0, 5mM EDTA pH 8.0) pulsed PIC. The lysate was pelleted by spinning at 1400 rpm for 5 minutes at room temperature. With discarding the supernatant, the chromatin pellet was suspended in 0.33% SDS incubation buffer (0.3% SDS, 1.6% Triton X-100, 100mM NaCl, 50mM Tris pH 8.0, 5mM EDTA pH 8.0) pulsed PIC at a concentration of ~30 million cells per ml, followed by Branson Sfx150 Sonifier sonication for total ON 4 minutes (1 second ON, 4 seconds OFF; 50% amplitude) to achieve enrichment of 200~500 bp DNA fragments. Sonicated chromatin was checked by agarose gel electrophoresis or TapeStation 2000 (Agilent) and quantitated with Qubit.    For regular ChIP, 10~20 μg chromatin and specific antibodies (e.g. SUZ12, JARID2, MTF2, FLAG, H3K27me3, and H2AK119ub) were incubated overnight with rotation at 4 °C. Following morning, 30 μl protein A/G magnetic beads were added to each ChIP sample, and incubated with rotation at 4 °C for 2 hours. After incubation, the beads were washed 5 times; once with low salt buffer (2 mM EDTA, 20 mM Tris, 1% Triton, 0.1% SDS and 150 mM NaCl), twice with high salt buffer (2 mM EDTA, 20 mM Tris, 1% Triton, 0.1% SDS and 500 mM NaCl) and twice with TE (1 mM EDTA and 10 mM Tris). After the last wash, beads were suspended in 200 μl elution buffer (1% SDS, 0.1 M NaHCO3) and incubated for 15 minutes with rotation at room temperature fallow incubation in the thermomixer for 10 minutes with gentle shaking (500 rpm) at 37 °C. The eluted ChIP supernatant was transferred to a new EP tube and incubated in a thermomixer overnight with 1000 rpm shaking at 65°C to remove the crosslinks. Then, RNase A was added and incubated for 1 hour at 37 °C with shaking; following  Proteinase K treatment for 2 hours at 55 °C with shaking. ChIP DNA was purified using the Qiagen MinElute PCR purification kit and measured with the Qubit High-Sensitive DNA assay kit. Quantitative PCR was performed to check the ChIP efficacy with suitable primers. The qPCR primer sequences were shown in SuppTable.     In addition, quantitative ChIP-Rx was performed using a modified published approach (Orlando et al., 2014, Healy et al., 2019). Briefly, 10% human NT2 chromatin or Drosophila  S2 chromatin was added to mouse chromatin lysate. Mixed chromatin was treated as single regular ChIP-seq experiment until the completion of DNA sequencing.

first: Eleanor

last: Glancy

first: Adrian

last: Bracken


title: Glis3 regulates the expression of thyroid hormone synthetic genes in coordination with other thyroid transcription factors

city: Research Triangle Park

first: Hong Soon

state: NC

address: 111 T.W. Alexander Dr.

department: IIDL

institute: NIH/NIEHS

summary: Comparison of cistromes from PAX8, NKX2.1, and FOXE1 ChIP-Seq analysis using mouse thyroid gland and rat thyrocyte PCCl3 cells revealed that there is a significant overlap between GLIS3 binding regions and those of PAX8, NKX2.1, and FOXE1 in genes associated with thyroid hormone biosynthesis.

sample_organism:
Mus musculus
Rattus norvegicus

platform_organism:
Mus musculus
Rattus norvegicus

overall_design: ChIP-seq for transcription factors PAX8, NKX2.1, GLIS3, and FOXE1 and histone modifications H3K27me3 and H3K4me3.

first: Anton

last: Jetten

first: Hong

middle: S

first: Sara

last: Grimm


title: The landscape of circular RNAs in neuroblastoma [IV]

city: Berlin

first: Steffen

last: Fuchs

address: Augustenburger Platz 1

department: Pediatric Oncology

country: Germany

institute: Charité - Universitätsmedizin Berlin

summary: Circular RNAs (circRNAs), a noncoding RNA class originating from alternative splicing, are highly abundant in neural tissues and were shown to regulate gene expression e.g. by interacting with microRNAs and RNA-binding proteins. Neuroblastoma is an embryonal neoplasia, which arises from undifferentiated neural crest cells. Here, we aimed to explore, whether circRNAs influence the pathogenesis of high-risk neuroblastoma. We performed whole-transcriptome sequencing of 104 primary neuroblastoma samples of all risk-groups and identified 5,203 unique circRNAs involving 2,302 genes. Candidate circRNA expression did not correlate with host gene expression, indicating independent regulatory mechanisms. circRNAs were significantly downregulated in the MYCN-amplified high-risk tumors. These findings were independently reproduced in a tetracycline-inducible MYCN-overexpression system based on a non MYCN-amplified neuroblastoma cell line, suggesting that MYCN drives this global circRNA repression. We identified the RNA helicase DHX9 as a mediator of this global suppressive effect of MYCN on circRNAs. Comparing our RNA sequencing data with other cancers and controls revealed a circRNA subset specifically upregulated in neuroblastoma that included a circRNA derived from the ARID1A tumor suppressor gene. Specific circARID1A knockdown resulted in reduced proliferation, cell numbers and viability, prompted apoptosis and induced a differentiated phenotype. Neither knockdown, nor overexpression of circARID1A influenced ARID1A mRNA and protein levels significantly. To dissect the potential mode of function, we performed a pulldown assay with subsequent mass spectrometry. We identified the RNA-binding protein KHSRP as an interaction partner that participates in the mechanism of action of circARID1A. In summary, this study highlights an important role of circRNAs in neuroblastoma biology and may establish this RNA class as a future therapeutic target and biomarker.

overall_design: Induction of MYCN expression in the neuroblastoma cell line SK-N-AS for 5d

first: Joerg

last: Fallmann

first: Johannes

last: Schulte


title: The landscape of circular RNAs in neuroblastoma II

overall_design: Knockdown of circARID1A by siRNA in the neuroblastoma cell line IMR-5 for 4d

first: Louisa-Marie

last: Kruetzfeldt

first: Clara

last: Danßmann


title: Cynomolgus monkey embryo model captures gastrulation and early pregnancy

city: Beijing

first: yuan

middle: qing

last: zhu

address: Zhongguancun north street

country: China

institute: Tsinghua University

summary: Here, we constructed monkey blastoids resembling blastocysts in morphology and transcriptomics using naïve ESCs and optimized protocol. The synthetic blastoids could develop to embryonic disk stage with the structure of yolk sac, chorionic cavity, amnion cavity, primitive streak, connecting stalk along the rostral–caudal axis by in-vitro prolonged culture (IVC). Primordial germ cells, gastrulating cells, visceral endoderm/yolk-sac endoderm, three germ layers and haemato-endothelial progenitors were identified in the monkey blastoid IVC embryo by single-cell transcriptomics or immunostaining. Besides, pregnancies with early gestation sacs were achieved by transferring monkey blastoids to surrogates. Our results revealed the in-vitro gastrulation and in-vivo early pregnancy of primate synthetic embryos, providing a powerful system to dissect primate embryonic development with less ethical concerns and restrict access.

overall_design: Single-cell profiling of monkey (cynomolgus macaque) blastoids (D7) and  further in-vitro cultured blastoids (D17).

first: Zhen

last: Liu

first: Jie

last: Li

first: Qingyuan

last: Zhu


title: The landscape of circular RNAs in neuroblastoma I

overall_design: Knockdown of DHX9 with 2 different siRNAs in the neuroblastoma cell line IMR-5

last: Krützfeldt


title: PRC2.1 and PRC2.2 Specific Accessory Proteins Drive Recruitment of Different Forms of Canonical PRC1 [4C-seq]

overall_design: Circularized chromosome conformation capture (4C) assays were performed using a modified published approach(van de Werken et al., 2012; Wang et al., 2019). Briefly, 10 million detached ESCs or EpiLCs were fixed with 1% final formaldehyde for 10 minutes, and quenched with final 0.125 M glycine. The cells were lysed in the 30 ml lysis buffer (50 mM Tris pH 7.5, 150 mM NaCl, 0.5% NP-40, 1% Triton, 5 mM EDTA, and  proteinase inhibitors: 2 μg/mL Aprotinin, 1 μg/mL Leupeptin, 1mM PMSF ) with rotation at 4 °C for 45  minutes. Isolated nuclei were then fully digested by restricted enzyme DpnII overnight digestion followed by overnight ligation with 50 U T4 DNA ligase at cool room temperature (18~20  °C) . Ligated circular DNA were purified after de-crosslinking incubation with proteinase K at 65 °C for overnight, followed by further incubation with Rnase A at 37  °C for 1 hour . The purified  DNA was further digested with restricted enzyme NlaIII and then ligated again and purified. The 4C libraries were amplified with the viewpoint-specific primers with inverse PCR using Roche Expand Long Template PCR System. For each viewpoint, at least 8 PCR reaction products were pooled to enhance the library complexity. The 4C PCR products were purified using Qiagen Quick PCR purification Kit. Tbx3 promoter viewpoint primers are AAGGGAAGAAGCTGCAGATC (Reading primer) and TGAAGGGAGCCCCACATG.


title: RNA-seq profiling of in vitro activated naïve B cells reveals differences in B-cell activation and mismatch repair machinery in common variable immune deficiency (CVID)

status: Public on Apr 05 2023

city: Vienna

first: Ola

last: Grimsholm

address: Wahringer Gurtel 18-20

department: Pathophysiology and Allergy Research

country: Austria

institute: Medical University of Vienna

summary:
To investigate the transcriptional dfferences during B-cell activation in healthy donors and patients with CVID we sorted naïve B cells from the two groups and activated with full BCR-engagement.
We then performed differential expression analysis on the seuqneces and we also investigated what pathway changes that we could detect between healthy donors and CVID patients.

overall_design: Comparative gene expression profiling analysis of RNA-seq data derived from in-vitro activated naïve B cells of health control and patients with common variable immune deficiency (CVID)

first: Shirin

middle: P

last: Akaber

first: Sabina

last: Barresi

first: Vanda

last: Friman


title: The Pumilio1 brain-region specific interactome reveals a direct connection with multiple RNA binding proteins, suggesting two disease mechanisms

city: New York

first: Vincenzo

last: Gennarino

address: 701 W 168th Street, Room 1402 HHSC

institute: Columbia University Medical Center

type: Non-coding RNA profiling by high throughput sequencing

summary: Different mutations in the RNA-binding protein Pumilio1 (PUM1) cause divergent phenotypes whose severity tracks with dosage: a mutation that reduces PUM1 levels by 25% causes late-onset ataxia, whereas haploinsufficiency causes developmental delay and seizures. Yet PUM1 targets are derepressed to equal degrees in both cases, and the more severe mutation does not hinder PUM1's RNA-binding ability. We therefore considered the possibility that the severe mutation might disrupt PUM1 interactions, and identified PUM1 interactors in the murine brain. We find that mild PUM1 loss derepresses PUM1-specific targets, but the severe mutation disrupts interactions with several RNA-binding proteins and the regulation of their targets. In patient-derived cell lines, restoring PUM1 levels restores these interactors and their targets to normal levels. Our results demonstrate that dosage sensitivity does not always signify a linear relationship with protein abundance but can involve distinct mechanisms. We propose that to understand the functions of RNA-binding proteins in a physiological context will require studying their interactions as well as their targets.

overall_design: Comparative analysis of microRNA sequencing on male and female cerebellum at 10 weeks of age in wt and Pum1-ko mice

first: Salvatore

last: Botta


title: Structural and functional properties of mSWI/SNF chromatin remodeling complexes revealed through single-cell perturbation and genomic profiling

city: Boston

first: Oana

last: Ursu

address: 415 Main Street

institute: Broad Institute

summary: Mammalian SWI/SNF (mSWI/SNF or BAF) chromatin remodeling complexes play critical roles in regulating DNA accessibility and gene expression and are comprised of three final-form assemblies, cBAF, PBAF, and ncBAF, which exhibit distinct modular organization, chromatin targeting, and roles in disease. However, the contributions of each subcomplex and their constituent subunits to gene expression remain incompletely defined. Here, we performed a large-scale CRISPR-Cas9 knockout screen targeting mSWI/SNF subunits individually and in selected combinations, followed by single-cell RNA-sequencing (Perturb-seq) and SHARE-Seq. We uncover complex- and module-specific contributions across cell regulatory pathways, define paralog subunit relationships, and identify shifted functions upon perturbations. Finally, we superimpose single-cell perturbation signatures over bulk primary human tumor gene expression profiles, defining unique subunit-specific signatures and identifying tumor features that mirror BAF loss-of-function. Taken together, these data highlight the utility of large-scale single-cell genomics to uncover gene expression and chromatin accessibility signatures in human disease states.

overall_design: 1. Perturb-seq experiments: To map the roles of different subunits of the mSWI/SNF complex, we performed Perturb-seq on MOLM13 cells, following either individual perturbation of subunits (“single” experiment) or combinatorial perturbation of selected subunit combinations (“combo” experiment). Cells were profiled with the 10X Chromium Single Cell 3’ RNA-seq v3 kit, with 6000 cells loaded per channel. The “single” and “combo” experiments were profiled with 15 and 3 channels, respectively, and were sequenced paired-end on an Illumina Hi-Seq instrument at a ratio of 3 channels of 10x to 1 lane of sequencing (5 for the “single” experiment, and 1 for the “combo” experiment), using paired end reads as follows.     For the “single” experiment, read 1 is 28 bases long (16 for cell barcode, 12 for UMI), read 2 is 91 bases long and sample index read is 8 bases long.     For the “combo” experiment, read 1 is 28 bases long (16 for cell barcode, 12 for UMI), read 2 is 91 bases long and sample index read is 8 bases long.     Perturbations were assigned to cells using dial-out PCR. Dial-out PCR products were sequenced on an Illumina Mi-seq, paired-end (for all experiments, read 1 is 28 bases and read 2 is 60 bases long), with read 1 having the same structure as in the high-throughput sequencing of the libraries described above. Each experiment required only one Mi-Seq run to process the dial-out libraries.     2. SHARE-seq experiment: To map the changes in accessibility and gene regulation controlled by mSWI/SNF complexes, we performed SHARE-seq on MOLM13 cells following CRISPR-mediated knockout of specific mSWI/SNF subunits, obtaining both RNA-seq and ATAC-seq readouts in the same single cell. RNA and ATAC libraries were sequenced using a 150 cycle high output Illumina kit on a NextSeq500 (Illumina) instrument. We performed SHARE-seq for 5 conditions: control guide, ARID1A, SMARCD2, SMARCA4 and BRD9, each with 2 replicate wells in an arrayed format. All cells from the SHARE-seq experiment were sequenced together in the same library, and can be demultiplexed by their well index (see Methods).

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