title: Tumor-associated macrophage functional heterogeneity is driven by niche diversity in breast cancer

city: San Francisco

first: Arjun

middle: Arkal

last: Rao

state: CA

address: 505 Parnassus Ave, S-447

department: CoLabs

institute: University of California, San Francisco

summary: The concept of macrophage niches has redefined the classification of macrophages, moving beyond ontogeny and function to encompass the mutually beneficial loop of signals in a given tissue environment. As such, tissue-resident macrophages adapt to local environmental signals within and between tissues to acquire specific functional adaptations. Neoplastic transition transforms the tissue environment, which then raises the question as to how existing macrophage subsets and their niche contribute to the tumor-associated macrophage (TAM) compartment. By combining single cell RNA sequencing and 2-photon imaging, we discovered that considerable TAM heterogeneity in mammary breast tumor is driven by niches that exist prior to tumor development, macrophage localization within the tumor and the stage of tumor malignancy. The differentiation of TAM subsets was associated with distinct signaling paths, homing and transcription factor signatures. We find similar functional heterogeneity in human breast TAMs. In overview, we show that specific niches within the tumor rather than defined activation states (e.g. the M1/M2 dichotomy) are the major drivers of TAM plasticity and heterogeneity. The distinctions created by this analysis show how treatments of different tumor indications should propose targeting specific TAMs at this niche/pathway level.

overall_design: 3 sorted tumor samples from a pool of 4 mice(spontaneous tumors; bulk myeloid, cd11b-hi-enriched, and cd11b-lo-enriched)  1 sorted tumor samples from a pool of 5 mice(orthotopically injected tumors; bulk myeloid)

first: Marie

last: Laviron

first: Maxime

last: Petit

first: Eléonore

last: Weber-Delacroix

first: Alexis

last: Combes

first: Sandrine

last: Barthélémy

first: Tristan

last: Courau

first: David

last: Hume

first: Christophe

last: Combadière

first: Matthew

middle: F

last: Krummel

first: Alexandre

last: Boissonnas


title: Single-cell ATAC-sequencing analysis of mouse totipotent potential stem cells, extended pluripotent stem cells and embryonic stem cells

status: Public on May 04 2022

city: Beijing

first: Yiming

last: Sun

state: Beijing

address: NO.5, Yiheyuan Road, Haidian District

department: School of Life Sciences

country: China

institute: Peking University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: It is challenging to derive totipotent stem cells in vitro that functionally and molecularly resemble cells from totipotent embryos. Here, we report that a chemical cocktail enables the derivation of totipotent-like stem cells, designated as totipotent potential stem (TPS) cells, from 2-cell mouse embryos and extended pluripotent stem cells that can be stably maintained long-term in vitro. TPS cells shared features with 2-cell mouse embryos in terms of totipotency markers, transcriptome, chromatin accessibility and DNA methylation patterns. In vivo chimeric assays show that these cells have embryonic and extraembryonic developmental potentials at the single cell level. Moreover, TPS cells can be induced into blastocyst-like structures resembling preimplantation mouse blastocysts. Mechanistically, inhibition of HDAC1/2 and DOT1L activity and activation of RARγ signaling are important for inducing and maintaining totipotent features of TPS cells. Our study opens up a path toward fully capturing totipotent stem cells in vitro.

overall_design: Total of 4 samples were analyzed, which include mouse totipotent potential stem (TPS) cells, extended pluripotent stem (EPS) cells and embryonic stem (ES) cells. For mouse TPS cells, 2 cell lines were analyzed, which included 1 cell line converted from mouse EPS cells and 1 cell line derived from 2-cell embryos. For mouse EPS cells and ES cells, 1 cell line was analyzed respectively.

first: Yaxing

last: Xu

first: Jingru

last: Zhao

first: Yixuan

last: Ren

first: Xuyang

last: Wang

first: Yulin

last: Lyu

first: Cheng

last: Li

first: Jun

first: Hongkui

last: Deng


title: Mutant p53 drives an immune cold tumor immune microenvironment in oral squamous cell carcinoma

city: Houston

first: Roberto

last: Rangel

state: Texas

address: 1515 Holcomb Blvd

department: Head and Neck Surgery

country: USA

institute: The University of Texas MD Anderson Cancer Center

type: Expression profiling by high throughput sequencing

summary: The critical role of the tumor immune microenvironment (TIME) in determining response to immune checkpoint inhibitor (ICI) therapy underscores the importance of understanding cancer cell–intrinsic mechanisms driving immune-excluded (“cold”) TIMEs. One such cold tumor is oral cavity squamous cell carcinoma (OSCC), a tobacco-associated cancer with mutations in the TP53 gene which responds poorly to ICI therapy. Because altered TP53 function promotes tumor progression and plays a potential role in TIME modulation, here we developed a syngeneic OSCC models with defined Trp53 (p53) mutations and characterized their TIMEs and degree of ICI responsiveness. We observed that a carcinogen-induced p53 mutation promoted a cold TIME enriched with immunosuppressive M2 macrophages highly resistant to ICI therapy. p53-mutated cold tumors failed to respond to combination ICI treatment; however, the combination of a programmed cell death protein 1 (PD-1) inhibitor and stimulator of interferon genes (STING) agonist restored responsiveness. These syngeneic OSCC models can be used to gain insights into tumor cell–intrinsic drivers of immune resistance and to develop effective immunotherapeutic approaches for OSCC and other ICI-resistant solid tumors.

overall_design: We generated gene expression profiles of stable p53-knockdown (KD) and non-targeting control-shRNA ROC1 tumor cells to determine the role of mutant p53 T122N in the expression of inflammatory cytokines and chemokines involved in oral cancer. From each group we run triplicate samples and genes differentially expressed were evaluated.

first: Jeffrey

middle: N

last: Myers


title: In vivo dissection of a clustered-CTCF domain boundary reveals developmental principles of regulatory insulation [Capture-C]

status: Public on May 05 2022

city: Berlin

first: Rafael

middle: Domínguez

last: Acemel

state: Berlin

address: Hannoversche Str. 28

department: Berlin Institute of Medical Systems Biology (BIMSB)

country: Germany

institute: Max Delbrück Center for Molecular Medicine (MDC)

summary: Vertebrate genomes organize into topologically associating domains (TADs), delimited by boundaries that insulate regulatory elements from non-target genes. However, how boundary function is established is not well understood. Here, we combine genome-wide analyses and transgenic mouse assays to dissect the regulatory logic of clustered-CTCF boundaries in vivo, interrogating their function at multiple levels: chromatin interactions, transcription and phenotypes. Individual CTCF binding sites (CBS) deletions revealed that the characteristics of specific sites can outweigh other factors like CBS number and orientation. Combined deletions demonstrated that CBS cooperate redundantly and provide boundary robustness. We show that divergent CBS signatures are not strictly required for effective insulation and that chromatin loops can be formed by non-canonically oriented sites, which suggests a loop interference mechanism. Further, we observe that insulation strength constitutes an effective modulator of gene expression and phenotypes. Our results highlight the modular nature of boundaries and their control over developmental processes.

overall_design: Capture Hi-C experiments at the Epha4 locus in developing distal limbs of several homozygous mutants and WT mice at E11.5

first: Chiara

last: Anania

middle: D

first: Johanna

last: Jedamzick

first: Adriano

last: Bolondi

first: Giulia

last: Cova

first: Norbert

last: Brieske

first: Ralf

last: Kühn

first: Lars

last: Wittler

first: Francisca

middle: M

last: Real

first: Darío

middle: G

last: Lupiáñez


title: Single-cell RNA sequencing analysis of mouse totipotent potential stem cells, extended pluripotent stem cells, embryonic stem cells, E17.5 TPS-derived chimeric placental cells, TPS-derived teratomas and TPS-derived blastoids

first: Yiming

last: Sun

overall_design: Total of 11 samples were analyzed, which include mouse totipotent potential stem (TPS) cells, extended pluripotent stem (EPS) cells, embryonic stem (ES) cells, E17.5 TPS-derived chimeric placental cells, TPS-derived teratomas and TPS-derived blastoids. For mouse TPS cells, 2 cell lines were analyzed, which included 1 cell line converted from mouse EPS cells and 1 cell line derived from 2-cell embryos. For mouse EPS cells, 2 cell lines were analyzed. For mouse ES cells, 2 cell lines were analyzed. For E17.5 TPS-derived chimeric placental cells, 2 samples were analyzed. For TPS-derived teratomas, 2 samples were analyzed. For TPS-derived blastoids, 1 sample was analyzed.

first: Yaxing

last: Xu

first: Jingru

last: Zhao

first: Yixuan

last: Ren

first: Xuyang


title: DNA methylation signatures in airway cells from adult children of asthmatic mothers reflect subtypes of severe asthma (RNA sequencing)

city: Chicago

first: Carole

last: Ober

state: IL

address: 920 E 58TH ST

department: Human Genetics

institute: The University of Chicago

summary: Maternal asthma (MA) is among the most consistent risk factors for asthma in her children. Possible mechanisms for this observation are epigenetic modifications in utero that have lasting effects on developmental programs in children of mothers with asthma. To test this hypothesis, we performed differential DNA methylation analyses of 398,186 individual CpG sites in primary bronchial epithelial cells (BECs) from 42 non-asthma controls and 88 asthma cases, including 56 without MA (NMA) and 32 with MA. We used weighted gene co-expression network analysis (WGCNA) of 69 and 554 differentially methylated CpGs (DMCs) that were specific to NMA or MA cases, respectively, compared to controls. WGCNA grouped 66 NMA-DMCs and 203 MA-DMCs into two and five co-methylation modules, respectively. The eigenvector of one MA-associated module (turquoise) was uniquely correlated with 85 genes expressed in BECs and enriched for 36 pathways, 16 of which discriminated between NMA and MA using machine learning. Genes in all 16 pathways were decreased in MA compared to NMA cases (P=7.1x10-3), a finding that replicated in nasal epithelial cells from an independent cohort (P=0.02). Functional interpretation of these pathways suggested impaired T cell signaling and responses to viral and bacterial pathogens. The MA-associated turquoise module eigenvector was additionally correlated with clinical features of severe asthma and reflective of type 2 (T2)-low asthma (i.e., low total serum IgE, fractional exhaled nitric oxide, and eosinophilia). Overall, these data suggest that maternal asthma alters diverse epigenetically-mediated pathways that lead to distinct subtypes of severe asthma in adults including hard-to-treat T2-low asthma.

overall_design: There were 118 samples that passed RNA-seq quality control and had paired DNA methylation data.

first: Kevin

last: Magnaye

first: William

last: Wentworth-Sheilds


title: Global DNA methylation analysis of mouse totipotent potential stem cells, extended pluripotent stem cells and mouse embryonic stem cells

type: Methylation profiling by high throughput sequencing

overall_design: Total of 10 samples were analyzed, which include mouse totipotent potential stem (TPS) cells, extended pluripotent stem (EPS) cells and embryonic stem (ES) cells. For mouse TPS cells, 2 cell lines were analyzed, which included 1 cell line converted from mouse EPS cells and 1 cell line derived from 2-cell embryos. For mouse EPS cells, 2 cell lines were analyzed. For mouse ES cells, 1 cell line was analyzed. For each sample, two technical replicates were included.

last: Wang

first: Yulin

last: Lyu

first: Cheng

last: Li

first: Jun

first: Hongkui

last: Deng


title: FTO mediates LINE1 m6A demethylation and chromatin regulation in mESCs and mouse development

first: Xiaolong

last: Cui

address: 680 N Lake Shore Drive

department: Department of Preventive Medicine

institute: Northwestern University

summary: N6-methyladenosine (m6A) is the most abundant internal modification on mammalian messenger RNA (mRNA). It is installed by a writer complex and can be reversed by erasers such as the fat mass and obesity-associated protein (FTO). Despite extensive research, the primary physiological substrates of FTO in mammalian tissues and development remain elusive. Here, we show that FTO mediates m6A demethylation of long-interspersed element-1 (LINE1) RNA in mouse embryonic stem cells (mESCs), regulating LINE1 RNA abundance and the local chromatin state, which in turn modulates transcription of LINE1-containing genes. FTO-mediated LINE1 RNA m6A demethylation also plays regulatory roles in shaping chromatin state and gene expression during mouse oocyte and embryonic development. Our results suggest broad effects of LINE1 RNA m6A demethylation by FTO in mammals.

overall_design: ATAC-Seq was performed on control and shFTO MEL624 cells, respectively. Additional ChIP-Seq for H3K27Ac was also included.

first: Jiangbo

last: Wei

first: Chuan

last: He


title: Global transcriptional, ATAC-sequencing, and methylation analyses of mouse totipotent potential stem cells and extended pluripotent stem cells

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

overall_design: Refer to individual Series


title: Immune transcriptome and secretome differ between CD71+ Erythroid Cells from adult bone marrow and fetal liver parenchyma

city: Novosibirsk

first: Roman

middle: Yurievich

last: Perik-Zavodskii

state: Novosibirsk region

address: Yadrincevskaya 14

country: Russia

institute: "Research Institute of Fundamental and Clinical Immunology" (RIFCI)

summary: In this work, we compared the expression profiles of cytokine genes between fetal liver CECs and adult bone marrow CECs.

overall_design: We profiled total RNA from the CD71+ erythroid cells (CECs) from either fetal human liver or bone marrow on NanoString Sprint Platform using Immunology V2 panel.


title: CPEB4-mediated translational regulation of LPS response in Bone Marrow Derived Macrophages (BMDMs)

city: Barcelona

first: Adria

last: Caballe

address: C/ Baldiri Reixac 10

department: biostatistics

country: Spain

institute: IRB

summary: Here we show the transcriptomic profile of the LPS response in BMDMs wild type and KO for the RNA-binding protein CPEB4.

overall_design: RNAseq in Wild type and CPEB4-KO BMDMs stimulated with LPS for 0, 1, 3, 6 and 9 h.

first: Clara

last: Suñer

first: Raul

last: Mendez


title: Proteasome-dependent truncation of the negative heterochromatin regulator Epe1 mediates antifungal resistance

city: Edinburgh

first: Robin

last: Allshire

state: Scotland

address: Max Born Crescent

department: Institute of Cell Biology

country: United Kingdom

institute: University of Edinburgh

summary: Epe1 histone demethylase restricts H3K9-methylation-dependent heterochromatin, preventing it from spreading over, and silencing, gene-containing regions in fission yeast. External stress induces an adaptive response allowing heterochromatin island formation that confers resistance on surviving wild-type lineages. Here we investigate the mechanism by which Epe1 is regulated in response to stress. Exposure to caffeine or antifungals results in Epe1 ubiquitylation and proteasome-dependent removal of the N-terminal 150 residues from Epe1, generating truncated tEpe1 which accumulates in the cytoplasm. Constitutive tEpe1 expression increases H3K9 methylation over several chromosomal regions, reducing expression of underlying genes and enhancing resistance. Reciprocally, constitutive non-cleavable Epe1 expression decreases resistance. tEpe1-mediated resistance requires a functional JmjC demethylase domain. Moreover, caffeine-induced Epe1-to-tEpe1 cleavage is dependent on an intact cell-integrity MAP kinase stress signalling pathway, mutations in which alter resistance. Thus, environmental changes provoke a mechanism that curtails the function of this key epigenetic modifier, allowing heterochromatin to reprogram gene expression, thereby bestowing resistance to some cells within a population. H3K9me-heterochromatin components are conserved in human and crop plant fungal pathogens for which a limited number of antifungals exist. Our findings reveal how transient heterochromatin-dependent antifungal resistant epimutations develop and thus inform on how they might be countered.

overall_design: Comparison of H3K9me2 methylation and gene expression in three Spombe geneotypes: epe1D, Epe1-GFP, Epe1DNP150-GFP

first: Imtiyaz

last: Yaseen

first: Sharon

middle: A

last: White

first: Sito

last: Torres-Garcia

first: Christos

last: Spanos

first: Marcel

last: Lafos

first: Elisabeth

last: Gaberdiel

first: Rebecca

last: Yeboah

first: Meriem

last: El Karoui

first: Juri

last: Rappsilber

first: Alison

middle: L

last: Pidoux

middle: C


title: RPAP2 regulates a transcription initiation checkpoint by prohibiting assembly of preinitiation complex [TT-seq]

city: Shanghai

first: Aixia

last: Song

address: Dong'an Road

institute: Fudan University

summary: RNA polymerase II (Pol II)-mediated transcription in metazoan requires precise regulation. RNA polymerase II-associated protein 2 (RPAP2) was previously identified to transport Pol II from cytoplasm to nucleus and dephosphorylates Pol II C-terminal domain (CTD). We found that RPAP2 binds hypo/hyper-phosphorylated Pol II with undetectable phosphatase activity. Structure of RPAP2-Pol II shows mutually exclusive assembly of RPAP2-Pol II and pre-initiation complex (PIC) due to three steric clashes. RPAP2 prevents/disrupts Pol II-TFIIF interaction and impairs in vitro transcription initiation, suggesting a function in prohibiting PIC assembly. Loss of RPAP2 in cells leads to global accumulation of TFIIF and Pol II at promoters, indicating critical role of RPAP2 in inhibiting PIC assembly independent of its putative phosphatase activity. Our study indicates that RPAP2 functions as a gatekeeper to prohibit PIC assembly and transcription initiation and suggests a novel transcription checkpoint.

overall_design: We performed TT-seq to investigate the role of RPAP2 in regulating transcription. In this study, samples for each condition were collected in biological duplicates. Cells were treated with dTAG13 for 3, 6 or 12 hours, the corresponding control cells were treated for the same duration with vehicle only (DMSO) at the same vol/vol dilution.

first: Zhenning

last: Wang

first: Fei

middle: X

last: Chen


title: ScRNA-seq Analysis of Whole Skin Cells from Irradiated and Naïve Mice

city: Jerusalem

first: Jonathan

middle: H.

last: Axelrod

state: n

address: Ein Kerem

department: Goldyne Savad Institute of Gene Therapy

country: Israel

institute: Hadassah Hebrew University Hospital

summary: We report the transcriptomic analysis of single cells derived from irradiated and naive murine whole skin. The aim of the study was to elucidate the molecular and cellular drivers of irradiation-induced alopecia and dermatitis (IRIAD), two of the most visually recognized complications of radiotherapy treatments in human cancer patients.

overall_design: Single-cell RNA-seq analysis was performed on pooled, viable cells isolated from whole skin samples derived from either irradiated (15 Gy) female (C57BL/6) mice 14 days post-treatment (n=4), or from age-matched naïve (C57BL/6) controls (n=4).

middle: H

last: Axelrd

first: Mor

last: Paldor

first: Orr

last: Levkovitch-Siany

first: Yuval

last: Nevo


title: Human Peripheral Blood mononuclear cells from postmenopausal women: grapefruit juice vs. flavanone-free placebo

status: Public on May 03 2022

city: Kannapolis

first: Dragan

last: Milenkovic

state: NC

address: 600 Laureate Way

department: Department of Food Bioprocessing & Nutrition Sciences

institute: North Carolina State University

summary: Transcriptional profiling of Peripheral Blood mononuclear cells (PBMCs) in apparently healthy postmenopausal women consuming flavanones in grapefruit juice (340 ml/day providing around 210 mg of naringenin glycosides) or flavanone-free placebo drink for 6 months in a cross-over design with a 2-month washout period between consumption of two studied drinks. Goal was to determine the effects of chronic consumption of flavanones in garpefruit juice on global gene expression profiles in humans

overall_design: Two-condition experiment-Cross-over study with two treatments: flavanone-rich grapefruit juice vs flavanone-free placebo drink. Biological replicates: 9 grapefruit juice replicates, 9 placebo drink replicates

first: Christine

last: Morand


title: Single-cell dissection of Comma-1D cultures reveals cell populations with unique differentiation and transformation properties

city: Memphis

first: Jiyang

last: Yu

state: TN

address: 262 Danny Thomas Pl

department: Department of Computational Biology

institute: St Jude Children's Research Hospital


title: RPAP2 regulates a transcription initiation checkpoint by prohibiting assembly of preinitiation complex [ChIP-seq]

overall_design: We performed ChIP-seq, ChIP-Rx or TT-seq to investigate the role of RPAP2 in regulating transcription. In this study, samples for each condition were collected in biological duplicates. Cells were treated with dTAG13 for 3, 6 or 12 hours, the corresponding control cells were treated for the same duration with vehicle only (DMSO) at the same vol/vol dilution.


title: Integrative Genomic Analysis of Medulloblastoma Identiﬁes a Molecular Subgroup That Drives Poor Clinical Outcome

city: San Diego

first: Jill

middle: P

last: Mesirov

address: 9500 Gilman Drive

department: Medicine

institute: University of California, San Diego

summary:
Medulloblastomas are heterogeneous tumors that collectively represent the most common malignant brain tumor in children. To understand the molecular characteristics underlying their heterogeneity and to identify whether such characteristics represent risk factors for patients with this disease, we performed an integrated genomic analysis of a large series of primary tumors.
Identified are six molecular subgroups of medulloblastoma, each with a unique combination of numerical and structural chromosomal aberrations that globally influence mRNA and miRNA expression. We reveal the relative contribution of each subgroup to clinical outcome as a whole and show that a previously unidentified molecular subgroup, characterized genetically by c-MYC copy number gains and transcriptionally by enrichment of photoreceptor pathways and increased miR-183-96-182 expression, is associated with significantly lower rates of event-free and overall survivals. Our results detail the complex genomic heterogeneity of medulloblastomas and identify a previously unrecognized molecular subgroup with poor clinical outcome for which more effective therapeutic strategies should be developed.

overall_design: We profiled the mRNA transcriptome of 194 medulloblastomas and performed high-density single nucleotide polymorphism array and miRNA analysis on 115 and 98 of these, respectively. Non-negative matrix factorization-based clustering of mRNA expression data was used to identify molecular subgroups of medulloblastoma; DNA copy number, miRNA profiles, and clinical outcomes were analyzed for each. We additionally validated our findings in three previously published independent medulloblastoma data sets.

first: Yoon-Jae

last: Cho

first: Aviad

last: Tsherniak

first: Pablo

last: Tamayo

first: Sandro

last: Santagata

first: Azra

last: Ligon

first: Heidi

last: Creulich

first: Rameen

last: Berhoukim

first: Vladimir

last: Amani

first: Liliana

last: Goumnerova

first: Charles

last: Eberhart

first: Ching

last: Lau

first: James

last: Olson

first: Richard

middle: J

last: Gilbertson

first: Amar

last: Gajjar

first: Olivier

last: Delattre

last: Kool

first: Keith

first: Matthew

last: Meyerson

first: Scott

last: Pomeroy


title: Human branching cholangiocyte organoids recapitulate embryogenic bile duct development and provide an unique model to study biliary diseases. RNA-Seq

city: Guangzhou

first: Haoyu

last: Wu

address: Kai Yuan Avenue 190

institute: GIBH

summary: Although providing promising and unique tools for studying cholangiocytes, current tissue-derived cholangiocyte-organoid systems do not recapitulate the complex architecture of the intrahepatic bile ducts in vitro. Here, we report a new method for creating branching cholangiocyte organoids (BRCO) from human adult tissue to study the intrahepatic biliary tree and diseases. BRCOs self-organize into large complex tubular structures, while closely resembling primary cholangiocytes on a transcriptomic and functional level. They are capable of mimicking branching bile duct development as well as being used for studying diseases in which the biliary tree does not develop properly (Alagille Syndrome). Furthermore, we deliver evidence that our culture method allows for formation of complex cholangiocyte cancer (cholangiocarcinoma, CCA) organoids. These branching CCA organoids resemble the primary tumor more closely compared to previously published protocols as well as showing unique tumor-specific drug responses. In conclusion, our culture method allows for creation of novel (malignant) cholangiocyte-organoids to study the intrahepatic bile ducts.

overall_design: bulk RNAsequencing analysis of paired BRCCAO and CCAO (n=3) samples and tumor tissue samples (n=2)

first: Floris

last: Roos

first: Gilles

last: van Tienderen

first: Monique

last: Verstegen

first: Luc

last: van der Laan


title: Global transcriptional analysis of mouse totipotent potential stem cells and extended pluripotent stem cells

first: Yiming

last: Sun

summary: It is challenging to derive totipotent stem cells in vitro that functionally and molecularly resemble cells from totipotent embryos. Here, we report that a chemical cocktail enables the derivation of totipotent-like stem cells, designated as totipotent potential stem (TPS) cells, from 2-cell mouse embryos and extended pluripotent stem cells that can be stably maintained long-term in vitro. TPS cells shared transcriptomic features with 2-cell mouse embryos and express totipotent markers. In vivo chimeric assays show that these cells have embryonic and extraembryonic developmental potentials at the single cell level. Moreover, we show that TPS cells can be induced into blastocyst-like structures resembling preimplantation mouse blastocysts. Our study demonstrates the feasibility of capturing and maintaining totipotency in vitro.

overall_design: Total of 12 samples were analyzed, which include mouse totipotent potential stem cells and extended pluripotent stem cells. For mouse totipotent potential stem cells, 3 cell lines were analyzed, which include 2 technicial replicates for each cell line. For mouse extended pluripotent stem cells, 3 cell lines were analyzed, which include 2 technicial replicates for each cell line.

first: Yaxing

last: Xu

first: Jingru

last: Zhao

first: Yixuan

last: Ren

first: Xuyang

last: Wang

first: Jun

first: Hongkui

last: Deng


title: Human branching cholangiocyte organoids recapitulate embryogenic bile duct development and provide an unique model to study biliary diseases. scRNA-Seq

overall_design: single cell transcriptomic profiles of ICO and BRCO samples


title: Single-cell dissection of Comma-1D cultures reveals cell populations with unique differentiation and transformation properties [scRNA-Seq]

first: Jiyang

last: Yu

summary: The mammary epithelial tree is a heterogeneous structure that contains two main populations, luminal and basal, with distinct molecular and functional characteristics. The investigation of how this heterogeneity is developed and how it influences breast tumorigenesis has been hampered by the need to perform these studies using animal models due to the lack of relevant, tractable, and fully characterized in vitro models. Comma-1D is an immortalized mouse mammary epithelial cell line that has unique morphogenetic properties particularly its ability to generate cells of luminal and basal lineages. Here, we perform single-cell RNA-seq studies to dissect its heterogeneous nature. Our analysis reveals that Comma-1D cultures consist of two main populations with luminal and basal features and a smaller population with mixed lineage and bipotent characteristics. Additionally, we demonstrate that multiple transcription factors associated with the differentiation of the mammary epithelium in vivo also modulate this process in Comma-1D cultures. Finally, we have used the Comma-1D model to study how the cellular subtype of origin influences the response to cancer-promoting mutations. We found that only cells with luminal molecular features were able to acquire transformed characteristics after an oncogenic HER2 mutant was introduced in their genomes. Overall, the studies present here characterize at a single-cell level the heterogeneity of the Comma-1D cell line and illustrate how the unique properties of Comma-1D cells can be used as an experimental model to study both the differentiation and the transformation processes in vitro.

overall_design: We performed scRNA-seq on unsorted and sorted Comma1D cell populations

first: Rachel

last: Werner

first: Koon-Kiu

last: Yan

first: Jose

last: Silva


title: FTO mediates LINE1 m6A demethylation and chromatin regulation in mESCs and mouse tissues

first: Chuan

last: He

address: 929 E 57th Street

department: Chemistry

institute: University of Chicago

summary: N6-methyladenosine (m6A) is the most abundant internal modification in mammalian messenger RNA (mRNA). It is installed by writer proteins and can be reversed by erasers. FTO was the first RNA demethylase shown to catalyze oxidative demethylation of m6A in RNA. Despite extensive studies, the main physiological substrates of FTO and the related functional pathways remain elusive in many systems, in particular during early mammalian development. Here, we show that FTO mediates the m6A demethylation of chromosome-associated repeat RNAs in mouse embryonic stem cells (mESCs), especially the long-interspersed element-1 family (LINE1) RNA, thereby affecting their abundances to regulate chromatin state.

overall_design:
ChIP profiles of histone marks and transcription factors in WT and Fto knockout mESCs were generated by deep sequencing, in duplicate using Illumina NovaSeq 6000. Nuclear RNA lifetime and nascent RNA synthesis of WT and Fto knockout mESCs were generated by deep sequencing, in triplicate, using Illumina NovaSeq 6000 machine.Embryos at GV, MII and morula stages were transferred to 500 ?l Trizol Reagent and then mixed well with 100 ?l chloroform added. The mixture was transferred Qiagen MaXtract High Density tubes (129046). After centrifuge, upper and aqueous phase was removed. RNA was precipitated by adding equal volume isopropanol and washing by 75% ethanol. Then purified RNA were analyzed by RT-qPCR or subjected to library generation using SMARTer Stranded Total RNASeq Kit (Takara Bio) following manufacturer?s protocol. Libraries were sequenced on an Illumina NovaSeq 6000 platform in paired-end read mode with 150 bp per read.
ATAC-Seq was performed on wild-type and FTO knockout mESCs.

first: Xianbin

first: Jiangbo

last: Wei


title: Spontaneous and ART-induced large offspring syndrome: similarities and differences in DNA methylome

city: Columbia

first: Rocío

middle: Melissa

last: Rivera

state: MO

address: 920 East Campus Drive

department: Division of Animal Sciences

institute: University of Missouri, Columbia

summary: Large/abnormal offspring syndrome (LOS/AOS) is a congenital overgrowth syndrome reported in ruminants produced by assisted reproduction (ART-LOS) which exhibit global disruption of the epigenome and transcriptome. LOS/AOS shares phenotypes and epigenotypes with the human congenital overgrowth condition Beckwith-Wiedemann syndrome. We have reported that LOS occurs spontaneously (SLOS), however, to date, no study has been conducted to determine if SLOS has the same methylome epimutations as ART-LOS. In this study, we performed whole genome bisulfite sequencing to examine global DNA methylation in SLOS and ART-LOS tissues.  We observed unique patterns of global distribution of differentially methylated regions (DMRs) over different genomic contexts, such as promoters, CpG islands and surrounding regions, and repetitive sequences from different treatment groups. In addition, we identified hundreds of LOS-vulnerable DMRs across different cattle breeds when analyzing four idependent LOS experiments together. Specifically, there are 25 highly vulnerable DMRs that could potentially serve as molecular markers for the diagnosis of LOS, including at the promoters of DMRT2 and TBX18, at the imprinted gene bodies of IGF2R, PRDM8, and BLCAP/NNAT, and at multiple CpG islands. We also observed tissue-specific DNA methylation patterns between muscle and blood, and conservation of ART-induced DNA methylation changes between muscle and blood. We conclude that as in ART-LOS, alterations of the epigenome are involved in the etiology of SLOS.  In addition, SLOS and ART-LOS share some similarities in methylome epimutations.

overall_design: Whole genome bisulfite sequencing (WGBS) for tissues (mainly blood and muscle) of bovine calves from control and treatment groups. Animals from the US include three from the control group, eight from the SLOS group, and three which are relatives to one SLOS animal. Animals from Spain (ES) include three from the control group, four from the ART group, three from the reproductive fluid (RF) group, and two from the RF_necropsy group.

first: Yahan

last: Li

first: Jordana

last: Sena Lopes

first: Pilar

last: Coy Fuster


title: RPAP2 regulates a transcription initiation checkpoint by prohibiting assembly of preinitiation complex [ChIP-Rx]

first: Aixia

last: Song

first: Zhenning

first: Fei

last: Chen


title: Single-cell dissection of Comma-1D cultures reveals cell populations with unique differentiation and transformation properties [bulk RNA-Seq]


title: Circulating microRNA signatures associated with disease severity and outcome in COVID-19 patients

city: Padova

first: Giulio

last: Ceolotto

address: Via Orus 2B

country: Italy

institute: University of Padova

type: Non-coding RNA profiling by high throughput sequencing

summary: In this study, we sought to identify circulating microRNA (miRNA) signatures associated with COVID-19 severity and outcome through small RNA-sequencing of serum samples from 89 COVID-19 patients and 45 healthy controls. As results, a set of miRNAs associated with lung disease, vascular damage and inflammation were upregulated in serum of COVID-19 patients vs controls, while miRNAs that inhibit pro-inflammatory cytokines and chemokines, angiogenesis and stress response were downregulated. In addition, patients with severe COVID-19 vs mild or moderate disease had a circulating miRNA signature associated with sepsis, hearth failure, tissue fibrosis, inflammation, and impairment of type I IFN and antiviral responses. A subset of the differentially expressed miRNAs predicted ICU admission, sequelae and mortality in COVID-19 patients. Investigation of the differentially expressed circulating miRNAs in relevant human cell types in vitro showed that some of these miRNAs were modulated directly by SARS-CoV-2 infection or indirectly by type I IFN stimulation.

overall_design: The study population included 89 COVID-19 patients, who were admitted at the IRCCS Sacro Cuore Don Calabria hospital, Negrar, Verona, Italy, in the period between May 2020 and December 2020. Inclusion criteria for the study were age ≥18 years and diagnosis of SARS-CoV-2 infection confirmed by molecular testing on nasopharyngeal swabs. As exclusion criteria, pregnant women were not enrolled. Disease severity was scored into mild, moderate, and severe at the time of hospital admission according to World Health Organization COVID-19 disease severity classification criteria 67. Peripheral blood samples were collected at the time of hospital admission, and before starting medications. Sera were separated from whole blood by centrifugation for 15 min at 3,000 rpm at 4 °C and stored at -80 °C until processing. Serum samples from 45 healthy volunteers collected before September 2019 and stored at -80 °C were used as negative control group.

first: Alessandra

last: Giannella


title: C5aR1 antagonism suppresses microglial polarization and mitigates disease progression in a mouse model of Alzheimer’s disease

status: Public on May 02 2022

city: Irvine

first: Ali

last: Mortazavi

address: 2300E Biological Sciences III

institute: UC Irvine

summary: Despite the discovery of PMX205 more than 20 years ago and its reported beneficial effects in Alzheimer’s disease, the specific mechanism that might be driving the improvement in cognition are still not know. Here, we used the Tg2576 mouse model of Alzheimer’s disease and treated them with PMX205 at the onset of the amyloid pathology to further determine the effects of this C5aR1 antagonist on microglial cells. The results presented in this study demonstrated a neuroprotective effect of PMX205, which rescues the excessive synaptic pruning and synapse loss associated with Alzheimer’s disease. This finding seems to be linked to the reduction of a unique microglial subpopulation associated to synaptic pruning in the PMX205 treated mice. Interestingly, we also show here that blocking C5a-C5aR1 signaling in the Tg2576 mouse model of AD results in the increase of the DAM2 microglial subpopulation, suggesting that PMX205 might be inducing a disease mitigating phenotype on microglial cells. Our data further supports the use of C5aR1 antagonists as potential therapeutic targets to treat or slow the progression of Alzheimer’s disease.

overall_design:
Single-cell RNA-seq libraries of 2 distinct mice genotypes and 2 distinct treatments
Mice were perfused with PBS and cortex was dissected. scRNA was extracted from cortex. Approximately 15,000 microglia and astrocytes were collected for scRNA-seq. Briefly, cellular viability was assessed and cells were subsequently washed with PBS prior to library preparation. scRNA-seq protocol was performed as previously described Illumina 2016).

first: Carvalho

last: Klebea

first: Gomes-Arboledas

last: Angela

first: Mortazavi

last: Ali;

first: Tenner

last: Andrea


title: Interplay between gene expression, histone modifications, and DNA methylation mediated by miR778 during beet cyst nematode parasitism of Arabidopsis [ChIP-seq]

city: Knoxville

first: Morgan

last: Bennett

email: mmbennet@vols.utk.edu, thewezi@utk.edu

address: 2505 E J Chapman Drive

department: Plant Sciences

institute: University of Tennessee

summary:
Purpose: The aim of this study is to investigate the role of H3K9 methylation in the development of syncytia induced by the beet cyst nematode (BCN, Heterodera schachtii). For this, ChIP-seq analysis was conducted in root tissues of Col-0 wild-type versus single suvh5 and suvh6 mutant plants and under BCN-infected and noninfected conditions.
Methods: Methods: Col-0 wild-type and suvh5 and suvh6 mutant plants were grown and 10-day-old plants were inoculated with 100 second-stage juvenile (J2) H. schachtii at 10 days old. Five days post inoculation, root tissues were collected from infected and noninfected plants in triplicate. ChIP DNA was precipitated using the H3k9me2 antibody (Abcam) and prepared into libraries. Then, high throughput ChIP DNA sequencing was performed using the Illumina NovoSeq 6000. Raw reads were trimmed of adapter sequences and overrepresented reads. Then, high quality, paired-end reads were aligned to the Arabidopsis thaliana reference genome (TAIR10.28) using Bowtie2. Finally, histone peaks were called using MACS2 for the identification of genes associated with differentially dimethylated H3K9 in wild-type versus mutant roots under BCN-infected and noninfected conditions.
Results and Conclusions: Under noninfected conditions, 511 and 143 H3K9me2 peaks were enriched in Col-0 compared to suvh5 and suvh6, respectively. The majority of these H3K9me2 peaks are located within TEs, which is consistent with previous findings that H3K9me2 corresponds to the repression of detrimental TEs. Our results suggest, however, that H3K9me2 adopts a new role in response to BCN. BCN-infected Col-0 had 791 H3K9me2 peaks compared to noninfected Col-0, 743 of which are located in the promoter or body of protein-coding genes. These peaks are associated with 1180 protein-coding genes. Similarly, suvh5 and suvh6 single mutants had 353 and 3106 H3K9me2 peaks compared to BCN-infected Col-0, respectively. The majority of these peaks are located in protein-coding genes and are associated with 558 and 4594 protein-coding genes, respectively. Interestingly, about 65% of peaks enriched in the BCN-infected wild type overlapped with peaks depleted in the suvh6 mutant, indicating that SUVH6 contributes to the majority of H3K9me2 during BCN parasitism.

overall_design: Genome-wide profiling of H3K9me2 mediated by SUVH5 and SUVH6 in Arabidopsisin response to H. schachtii infection

first: Tarek

last: Hewezi


title: Developmental disruption to the cortical transcriptome and synaptosome in a model of SETD1A loss-of-function

city: Exeter

first: Nicholas

middle: E

last: Clifton

address: RILD Building

institute: University of Exeter

summary: We employed a multi-omics approach to study the effects of heterozygous Setd1a LoF on gene expression and synaptic composition in mouse cortex across five developmental timepoints from embryonic day 14 to postnatal day 70. Using RNA sequencing, we observed that Setd1a LoF resulted in the consistent downregulation of genes enriched for mitochondrial pathways. This effect extended to the synaptosome, in which we found age-specific disruption to both mitochondrial and synaptic proteins.

overall_design: Quantification of RNA and protein alterations in mouse frontal cortex after heterozygous Setd1a knockout, at multiple stages of development


title: Human branching cholangiocyte organoids recapitulate embryogenic bile duct development and provide an unique model to study biliary diseases.

first: Haoyu

last: Wu

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

overall_design: Refer to individual Series


title: Interplay between gene expression, histone modifications, and DNA methylation mediated by miR778 during beet cyst nematode parasitism of Arabidopsis [RNA-seq]

summary:
Purpose: The aim of this study is to investigate the role of H3K9 methylation in the development of syncytia induced by the beet cyst nematode (BCN, Heterodera schachtii). For this, differential gene expression analysis was performed in root tissues of Col-0 wild-type and suvh5 and suvh6 single mutants under BCN-infected and noninfected conditions.
Methods: Col-0 wild-type and suvh5 and suvh6 mutant plants were grown and 10-day-old plants were inoculated with 100 second-stage juvenile (J2) H. schachtii at 10 days old. Five days post inoculation, root tissues were collected from infected and noninfected plants in triplicate. Total RNA was extracted, mRNA libraries were prepared, and then high throughput RNA sequencing was performed using the Illumina NovoSeq 6000. High quality, paired-end reads were then aligned to the Arabidopsis thaliana reference genome (TAIR10.28) using STAR and uniquely mapped reads were counted using HTSeq. Finally, differentially expressed genes (DEGs) between the mutant lines and wild-type plants under non-infected and infected conditions were identified using DESeq package.
Results and conclusions: At least 20 million reads were uniquely mapped to the Arabidopsis reference genome with an average length of about 300 bp for paired-end reads. A total of 1057 and 3390 DEGs were identified for suvh5 and suvh6 versus Col-0, respectively, under noninfected conditions. Upon infection of BCN, Col-0 had 4566 DEGs compared to the noninfected wild type. BCN-infected single mutants suvh5 and suvh6 had 2487 and 846 DEGs compared to BCN-infected Col-0, respectively. Of these DEGs, 1277 and 592 or about 51% and 70% are also differentially expressed in BCN-infected Col compared to noninfected Col-0, respectively, and 324 of the 1277 and 592 DEGs are common to BCN-infected suvh5 and suvh6 compared to the control. DEGs common to BCN-infected Col-0 and/or suvh5, and suvh6 are involved in stress and defense responses, primary and secondary metabolism, signal transduction, cell wall organization and other cellular processes. This suggests that SUVH5 and SUVH6 play a major role in transcriptome reprogramming to facilitate cellular, molecular, and metabolic processes involved in the compatible plant-nematode interaction.

overall_design: Transcriptional profiles of H. schachtii infected and noninfected suvh5 and suvh6 mutants compared to Col-0 wild type


title: Cas9-guided haplotyping of three truncation variants in autosomal recessive disease

city: Sapporo

first: Yoshikazu

last: Furuta

state: Hokkaido

address: North 20 West 10, Kita-ku

department: International Institute for Zoonosis Control

country: Japan

institute: Hokkaido University

summary: An autosomal recessive disease is caused by biallelic loss-of-function mutations. However, when more than two disease-causing variants are found in a patient’s gene, it has been challenging to determine which two of the variants are responsible for the disease phenotype. To decipher the pathogenic variants by precise haplotyping, we applied nanopore Cas9-targeted sequencing (nCATS) to three truncation COL7A1 variants detected in a patient with recessive dystrophic epidermolysis bullosa (EB). The distance between the most 5’ and 3’ variants was around 19 kb at the level of genomic DNA. nCATS successfully delineated that the most 5’ and 3’ variants were located in one allele while the variant in between was in the other allele. Intriguingly, the proband’s mother, who was phenotypically intact, was heterozygous for the allele that harbored the two truncation variants, which could otherwise be misinterpreted as those of typical recessive dystrophic EB. Our study illuminates nCATS as a useful tool to determine haplotypes of complicated genetic cases. Haplotyping of multiple variants in a gene can tell which variant should be therapeutically targeted when nucleotide-specific gene therapy is applied.

overall_design: Genomic region of COL7A1 which is about 8 kb long was enriched by PCR or nCATS, then applied for MinION sequencing.

first: Ken

last: Natsuga

first: Shota

last: Takashima

first: Takuma

last: Nohara

first: Hsin-Yu

last: Huang

first: Satoru

last: Shinkuma

first: Hideki

last: Nakamura

first: Yousuke

last: Katsuda

first: Hideaki

last: Higashi

first: Chao-Kai

last: Hsu

first: Satoshi

last: Fukushima

first: Hideyuki

last: Ujiie


title: RNA-seq analyses of human ameloblastoma and odontogenic keratocyst

city: Seoul

first: Han-Sung

last: Jung

address: 50, Yonsei-ro, Seodaemun-gu

department: Dental School

country: South Korea

institute: Yonsei University

summary: Ameloblastoma (AM) is a benign but locally invasive tumor with high recurrence rates. Invasive behavior of the odontogenic tumor results in destruction of the adjacent jawbone and formation of daughter cysts, hindering the complete elimination of cancer cells during surgery. To understand the underlying mechanism of AM invasion, we compared the transcriptome of AM with that of an odontogenic keratocyst (OKC), a dental epithelium-originated cyst with non-invasive characteristics.

overall_design: RNA was extracted from three ameloblastoma and three odontogenic keratocyst tissues collected from individual patients and subjected into RNA-seq anlaysis.

first: Hyun-Yi

last: Kim

first: Shujin

last: Li

first: Young-Soo


title: Comparative Transcriptomics Reveal Determinants of Amphotericin B Resistance in Candida auris

city: NEWARK

first: Neeraj

last: Chauhan

state: New Jersey

address: 225 Warren Street

department: Department of Microbiology, Biochemistry & Molecular Genetics

institute: NJMS-PHRI

summary: Candida auris is an emerging multidrug-resistant human fungal pathogen often refractory to treatment by all classes of antifungal drugs. Amphotericin B (AmB) is a fungicidal drug that, despite its toxic side effects, remains a drug of choice for the treatment of drug-resistant fungal infections, including those caused by C. auris. However, the molecular mechanisms underlying AmB resistance are poorly understood. In this study, we present data that suggests membrane lipid alterations and chromatin modifications are critical processes that contribute to or cause adaptive AmB resistance in clinical C. auris isolates. To determine the plausible cause of increased AmB resistance, we performed RNA-seq of AmB-resistant and sensitive C. auris isolates. Remarkably, AmB-resistant strains show a pronounced enrichment of genes involved in lipid and ergosterol biosynthesis, adhesion, drug transport as well as chromatin remodeling. The transcriptomics data confirm increased adhesion and reduced lipid membrane permeability of AmB-resistant strains compared to the sensitive isolates. The AmB-resistant strains also display hyper-resistance to cell wall perturbing agents, including congo red, calcofluor white and caffeine. Additionally, we noticed an increased phosphorylation of Mkc1 cell integrity MAP kinase upon AmB treatment. Collectively, these data identify differences in the transcriptional landscapes of AmB-resistant vs AmB-senstive isolates, and provide a framework for the mechanistic understanding of AmB resistance in C. auris.

overall_design: We performed RNA-seq analysis of logarithmically growing  Amphotericin B resitant (AmBR) in comparison to Amphotericin B susceptible (AmBS)  Candida auris isolates  in the Yeast Extract Peptone Dextrose broth medium (YPD; 1% yeast extract, 2% peptone, and 2% dextrose) at 30°C. We sequenced four different isolates AmBR1, AmBR2, AmBR3 and AmBS in biological triplicates.

first: Raju

last: Shivarathri

first: Manju

first: Sabrina

last: Jenull


title: RNA-seq analysis of peripheral nerve with conditional knockout of Bap1 deubiquitinase.

city: Madison

first: John

last: Svaren

state: WI

address: 1500 Highland Ave.

department: Comparative Biosciences

institute: University of Wisconsin

summary: The goal of the experiment was to determine the role of H2A deubiquitinase in the nerve injury response in peripheral nerve. A Schwann cell specific knockout of the H2A deubiquitinase (Bap1) was generated to compare with wild type mice.

overall_design: For each condition, six independent biological replicates were prepared.

first: Phu

last: Duong


title: Interplay between gene expression, histone modifications, and DNA methylation mediated by miR778 during beet cyst nematode parasitism of Arabidopsis

first: Morgan

last: Bennett


title: Gene expression profile at single cell level of a mesothelioma patient specimen treated with ADU-S100

city: Boston

first: Michael

last: Tolstorukov

state: MA

address: 360 Longwood Avenue

department: Informatics

institute: Dana Farber Cancer Institute

summary: STING agonists are in clinical development, yet their mechanisms of action remain unclear. We used single cell RNA sequencing (scRNA-seq) to analyze response to ADU-S100 in a mesothelioma patient specimen ex vivo.

overall_design: Tumor pieces from a patient with mesothelioma were treated with ADU-S100 and analyzed using scRNAseq.

first: Erik

last: Knelson

first: Matthew

last: Booker

first: David

last: Barbie


title: Single cell analysis of RORgt-expressing immune cells in the large intestine of healthy mice

city: New York

first: Gregory

last: Putzel

state: NY

address: 430 East 29th Street

department: Microbiology

institute: NYU Langone Health

summary: RORgt is a lineage-specifying transcription factor expressed among immune cells that are enriched in the gastrointestinal tract and critically orchestrate immunity, inflammation, and tissue homeostasis. However, the spectrum of cellular heterogeneity among these cell types, the mechanisms controlling their tissue protective versus inflammatory roles, and their functional redundancy, are not fully understood. To address these fundamental gaps in knowledge, we performed single-cell RNA sequencing on RORgt-expressing immune cells of the large intestine of healthy mice and identified 10 distinct clusters of cells based on expression of signature genes.

overall_design: RORgt+ immune cells were sorted from the large intestine lamina propria of RORgt-EGFP mice and mixed at a 1:1 ratio of TCRb+ cells to TCRb- cells, to enrich the non-T cell population. Cells were pooled from two mice.

first: Wenqing

last: Zhou

last: Sonnenberg


title: BMP gradient along the intestinal villus axis controls zonated enterocyte and goblet cell states

city: Utrecht

first: Jens

last: Puschhof

address: Uppsalalaan 8

country: Netherlands

institute: Hubrecht Institute

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Single cell and bulk RNA sequencing was performed from human intestinal organoids differentiated for 5 days in the presence or absence of BMP-2 and BMP-4 ligands. For in vivo validation, intestines of wildtype and BMPR1a knockout mice were subjected to bulk RNA sequencing.

first: Joep

last: Beumer

first: Fjodor

middle: Y

last: Yengej

first: Lianzheng

last: Zhao

first: Adriana

last: Silgado

first: Marloes

last: Blotenburg

first: Harry

last: Begthel

first: Charelle

last: Boot

first: Alexander

last: van Oudenaarden

first: Ye-Guang

last: Chen

first: Hans

last: Clevers


title: mRNA and microRNA Transcriptional Profiling of Gestation Day 21 Female Rat Liver and Placenta following Gestation Day 6 - 20 Ketoconazole Exposure

city: Indianapolis

first: Kamin

last: Johnson

state: IN

address: 9330 Zionsville Rd

institute: Corteva Agriscience

summary: We report the RNAseq-based mRNA and microRNA transcriptome profiles of gestation day 21 female rat liver and placenta following oral ketoconazole exposure from gestation day 6 to 20.  A treatment-related change in mRNA and microRNA levels was observed in both liver and placenta.

overall_design: Gestation day 21 female rat liver and placenta mRNA and microRNA RNAseq profiles following exposure to ketoconazole from gestation days 6 - 20.

first: Shreedharan

last: Sriram


title: Popliteal adipocytes from mice infected with L. monocytogenes

city: Baltimore

first: Immunometabolism

last: Department

state: Maryland

address: 1650 Orleans Street

department: Immunometabolism

institute: Johns Hopkins University

summary: A wild type strain of Listeria monocytogenes (L. mono) was used for infections. Mice were infected subcutaneously on the footpad with a sublethal dose of 1 × 10^6 colony-forming units (CFU). Mice were left for indicated times before being sacrificed. Control cells were from either PBS injected mouse or from the uninjected leg of infected animal.

overall_design: Adipocytes were isolated via centrifugation. Two independent biological replicates per condition were used.

first: George

last: Caputa

first: Mai

last: Azuma

last: Sanin

first: Edward

last: Pearce

first: Erika


title: DNA methylation signatures in airway cells from adult children of asthmatic mothers reflect subtypes of severe asthma (DNA methylation)

city: Chicago

first: Carole

last: Ober

address: 920 E 58TH ST

department: Human Genetics

institute: The University of Chicago

summary: Maternal asthma (MA) is among the most consistent risk factors for asthma in her children. Possible mechanisms for this observation are epigenetic modifications in utero that have lasting effects on developmental programs in children of mothers with asthma. To test this hypothesis, we performed differential DNA methylation analyses of 398,186 individual CpG sites in primary bronchial epithelial cells (BECs) from 42 non-asthma controls and 88 asthma cases, including 56 without MA (NMA) and 32 with MA. We used weighted gene co-expression network analysis (WGCNA) of 69 and 554 differentially methylated CpGs (DMCs) that were specific to NMA or MA cases, respectively, compared to controls. WGCNA grouped 66 NMA-DMCs and 203 MA-DMCs into two and five co-methylation modules, respectively. The eigenvector of one MA-associated module (turquoise) was uniquely correlated with 85 genes expressed in BECs and enriched for 36 pathways, 16 of which discriminated between NMA and MA using machine learning. Genes in all 16 pathways were decreased in MA compared to NMA cases (P=7.1x10-3), a finding that replicated in nasal epithelial cells from an independent cohort (P=0.02). Functional interpretation of these pathways suggested impaired T cell signaling and responses to viral and bacterial pathogens. The MA-associated turquoise module eigenvector was additionally correlated with clinical features of severe asthma and reflective of type 2 (T2)-low asthma (i.e., low total serum IgE, fractional exhaled nitric oxide, and eosinophilia). Overall, these data suggest that maternal asthma alters diverse epigenetically-mediated pathways that lead to distinct subtypes of severe asthma in adults including hard-to-treat T2-low asthma.

overall_design: There were 142 samples that passed DNA methylation quality control and 130 samples with available variables that were analyzed in the manuscript.

first: Kevin

last: Magnaye

first: William

last: Wentworth-Sheilds


title: TCF-1 mediates 3D genome reconfiguration during T cell development (HiChIP)

city: Philadelphia

first: Wenliang

last: Wang

state: PENNSYLVANIA

address: 421 Curie Boulevard

institute: University of Pennsylvania

summary: Lineage-determine transcription factors (LDTFs) can determine gene expression programs of a specific cell type, by rewiring the linear regulatory network. However, it’s not clear whether and how LDTFs can alter the 3D genome organization during development. Here, we investigated the effect of T cell factor 1 (TCF-1) on the reconfiguration of 3D genome during T cell development. Topologically associated domains (TADs) that have higher density of TCF-1 gain more intra-TAD interactions during T cell development. TCF-1 colocalized with CCCTC-binding factor (CTCF) substantially reduced the insulation and increased the local interactions, which additionally have stronger interactions with T cell development genes. We further found that a considerable number of boundaries are lost during T cell development, and that TCF-1 is enriched at the lost boundaries, including the ones near key T cell development genes. Overexpression of TCF-1 in fibroblast demonstrates that both high density of TCF-1 and TCF-1 colocalized with CTCF can optimally increase the local interactions. Correspondingly, knocking out of TCF-1 in DN3 cell line leads to decreased intra-TAD interactions and gain of insulation at some of its binding sites. Finally, we found that TCF-1 can recruit and maintain cohesin at its binding sites, which probably cause reconfiguration of the 3D genome. Taken together, our study shows that TCF-1 can change the 3D genome at both TAD level and finer scale potentially through the recruitment of cohesin.

overall_design: Study the 3D genome reorganization mediated by TCF-1

first: Aditi

last: Chandra

first: Naomi

last: Goldman

first: Golnaz

last: Vahedi


title: Transcriptome comparison between Zbtb46-deficient and -sufficient group 3 innate lymphoid cells

first: Gregory

last: Putzel

summary: We unexpectedly identify one unique subset of group 3 innate lymphoid cells (ILC3s) that co-expresses Zbtb46, a transcription factor that specifies conventional dendritic cells (cDCs). Zbtb46 is robustly expressed by CCR6+ lymphoid tissue inducer (LTi)-like ILC3s that are developmentally and phenotypically distinct from cDCs. To determine the function of Zbtb46 in ILC3s, we performed RNA sequencing on CCR6+ ILC3s sorted from the large intestine of Zbtb46 f/f and Rorc Cre Zbtb46 f/f mice at steady state.

overall_design: CCR6+ ILC3s (CD45-dim CD90-hi CD3- CD5- NK1.1- TCR gamma delta- CD11b-CD11c-B220-CD127+CCR6+) were sorted from the large intestine of Zbtb46f/f and RorcCreZbtb46f/f mice at steady state. Cells were sorted from 4 individual mice as replicates for each group.

first: Wenqing

last: Zhou

last: Sonnenberg


title: A plant tethering system for the functional study of protein-RNA interactions in vivo

city: St. Louis

first: R. Keith

last: Slotkin

state: Missouri

address: 975 North Warson Road

institute: Donald Danforth Plant Science Center

summary: The sorting of RNA transcripts dictates their ultimate post-transcriptional fates, such as translation, decay or degradation by RNA interference (RNAi). This sorting of RNAs into distinct fates is mediated by their interaction with RNA-binding proteins. While hundreds of RNA binding proteins have been identified, which act to sort RNAs into different pathways is largely unknown. Particularly in plants, this is due to the lack of reliable protein-RNA artificial tethering tools necessary to determine the mechanism of protein action on an RNA in vivo. Here we generated a protein-RNA tethering system which functions on an endogenous Arabidopsis RNA that is easily tracked by the quantitative flowering time phenotype. Unlike other protein-RNA tethering systems that have been attempted in plants, our system circumvents the inadvertent triggering of RNAi. We successfully in vivo tethered a protein epitope, deadenylase protein and translation factor to the RNA, which function to tag, decay and boost protein production, respectively. We demonstrated that our tethering system 1) is sufficient to engineer the downstream fate of an RNA, 2) enables the determination of any protein’s function upon recruitment to an RNA, and 3) can be used to discover new interactions with RNA-binding proteins.

overall_design: Examination of small RNA accumulation from the SOC1 mRNA and other regions in Arabidopsis thaliana lines with constructs that tether specific proteins to SOC1 and controls.

first: Diego

last: Cuerda-Gil

first: Kaushik

last: Panda

first: R

middle: K


title: TCF-1 mediates 3D genome reconfiguration during T cell development (ChIP-Seq)


title: TCF-1 mediates 3D genome reconfiguration during T cell development (RNA-Seq)


title: Cyclic hypoxia induces transcriptomic changes in mast cells leading to a hyperresponsive phenotype after FcεRI crosslinking

status: Public on May 01 2022

city: Mexico city

first: Monica

last: Lamas

state: Distrito Federal

address: Calzada de los Tenorios 235

department: Pharmacobiology

country: Mexico

institute: CINVESTAV

summary:
Mast cells (MCs) play an important role in tumor development, executing pro or antitumoral functions depending on tumor type and local conditions present in the tumor microenvironment such as cyclic hypoxia (cyH), which is a distinguishing feature of almost all solid tumors.
In this study, we analyzed transcriptional changes of murine bone marrow-derived mast cells (BMMCs) exposed to an in vitro protocol of cyH, consisting of interleaved cycles of hypoxia and re-oxygenation at 1% and 21% of oxygen, respectively. Utilizing a mouse M22K microarray (Microarray Unit, Cellular Physiology Institute, UNAM, Mexico City), we identified 2512 genes in MCs subjected to cyH whose expression displayed changes compared with normoxic cells. Furthermore, functional enrichment analysis revealed that cyH-related transcriptomic alterations were for genes associated with oxidative phosphorylation and the FcεRI signaling pathway. Interestingly, after IgE/ antigen (Ag) challenge, FcεRI-dependent degranulation, calcium mobilization, and PLC-γ activity were enhanced in BMMCs exposed to cyH compared to those cells maintained in normoxic conditions. In addition, the expression of calcium signaling-related cytokines such as TNF-α, IL-4, and IL-2 was increased in BMMCs exposed to cyH after IgE/Ag challenge. Taken together, these findings indicate that cyH induces transcriptomic modifications in MCs that are translated into a phenotypic change characterized by hyperresponsiveness to IgE/Ag challenge.

overall_design: A two-color microarrays experiment was performed. We separately pooled equal amounts of high-quality RNA from three different cell cultures of BMMCs, which were exposed to normoxia (21%oxygen for 2.5 hours) or cyclic hypoxia (1 hour of hypoxia at 1% oxygen and 30 minutes at 21% oxygen, followed by a second hour of hypoxia at 1% oxygen.). Then, cDNA synthesis was performed and ach sample was labeled as follows: Alexa555 (Cy3) labeling was used for normoxia while Alexa647 (Cy5) labeling was used for cyclic hypoxia. Equal quantities of both samples were hybridized in the same microarray.

first: D

last: Segura-Villalobos

first: M

first: C

last: González-Espinosa


title: RNA-seq and m6A-seq were used to analysis RNA m6A methylation between hypertrophic and normal mouse hearts

city: zhengzhou

first: Weidong

last: Li

address: kexuedadao zhengzhou

institute: Zhengzhou University

summary: In this experiment, we constructed a mouse myocardial hypertrophy model through transverse aortic constriction (TAC) to further explore the relationship between m6A modification and myocardial hypertrophy, MeRIP-seq and RNA-seq were performed to identify genes with differences m6A modification or expression in transcriptome profile.

overall_design: Comparative gene expression profiling analysis of RNA-seq data and examing the m6A modification between hypertrophic and normal mouse hearts(sham and TAC).

first: Hongkun

last: Fan


title: MOZ and Menin-MLL Complexes are Complementary Regulators of Chromatin Association and Transcriptional Output in Gastrointestinal Stromal Tumor [ChIP-seq and CUT&Tag]

city: Worcester

first: Matthew

last: Hemming

address: 364 Plantation Street

institute: UMass Medical School

summary: Gastrointestinal stromal tumor (GIST) is commonly characterized by activating mutations in the receptor tyrosine kinase KIT. Tyrosine kinase inhibitors are the only approved therapy for GIST, and other complementary treatment strategies are urgently needed. As GIST both lacks oncogene amplification and relies upon an established network of transcription factors, we hypothesized that unique chromatin modifying enzymes are essential in orchestrating the GIST epigenome. Here, we identified through genome-wide CRISPR screening that MOZ and Menin-MLL chromatin regulatory complexes are cooperative and unique dependencies in GIST. These complexes were enriched at GIST-relevant genes and regulated their transcription. Inhibition of MOZ and Menin-MLL complexes decreased GIST cell proliferation by disrupting interactions with transcriptional regulators, such as DOT1L. Menin inhibition caused significant reductions in tumor burden in xenograft models, with superior effects observed in combination with imatinib. These results define unique chromatin regulatory dependencies in GIST and identify therapeutic strategies for clinical application.

overall_design: ChIP-seq and CUT&Tag of GIST cell line GIST-T1


title: Simultaneously profiling of mRNA and miRNA in epididymitis

city: Hong Kong

first: Jianlin

address: L 7-08, Department of Obstetrics and Gynecology  Laboratory block, 21th Sassoon Road, Hong Kong

country: Hong Kong

institute: The University of Hong Kong

summary: Chronic epididymitis (CE) refers to a long-lasting inflammatory condition of the epididymis, which is considered the most common site of intrascrotal inflammation and an important aetiological factor of male infertility. Recent studies demonstrate that small RNAs secreted from epididymal epithelium modulate embryo development and offspring phenotypes via sperm transmission, and the resulting modifications may lead to transgenerational inheritance. However, to date, the genome-wide analysis of small RNA together with the transcriptomic expression profiles of human epididymis and CE is still lacking. In this study, we collect inflamated epdidymis from patients as well as control sample. By simultaneous mRNA and  small RNA suquencing, we identified segment-specific inflammation signitures including leukocyte chemotaxis, ion channel and transporter-related processes. We also compare the miRNA, tsRNA, and other small RNA distribution in CE. Together, Interative analysis of miRNA and mRNA identified strong candidate epigenitic cycle in CE.

overall_design: Two group of samples, with 5 biological replicates within each group and total of 6 samples were analyzed

first: Jialei

last: Gong

first: Dandan

last: Cao

first: Yonggang

last: Duan


title: A circulating risk score, based on combination of exo-miR130a-3p and fibrinopeptide A, as predictive biomarker of relapse in early stage non-small cell lung cancer patients

city: Genova

first: simona

last: coco

address: L.go R. Benzi, 10

institute: IRCCS Ospedale Policlinico San Martino

type:
Non-coding RNA profiling by array
Other

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

overall_design: Refer to individual Series


title: A circulating risk score, based on combination of exo-miR130a-3p and fibrinopeptide A, as predictive biomarker of relapse in early stage non-small cell lung cancer patients [miRNA]

summary: To date, the 5-year overall survival of early stage non-small cell lung cancer (NSCLC) is still unsatisfactory at 59.8%. Therefore, reliable prognostic factors are needed. Growing evidence shows that cancer progression may depend on a double interconnection between cancer cells and the surrounding tumor microenvironment, and circulating molecules may represent promising markers of cancer recurrence. Here, we performed in-depth analysis of circolome-derived markers, including Exo-miRnome and peptidome in 67 radically resected NSCLCs, to identify a prognostic score. The miRnome profile selected exo-miR130a-3p as the most overexpressed in patients with relapse. Peptidome analysis identified 4 fibrinopeptide A (fpA), which were depleted in progressing patients. Notably, the combination of exo-miR130a-3p and the greatest fpA (2-16) built a score where high-risk patients had 24 months of disease-free survival. Moreover, in vitro transfections showed that higher levels of exo-miR-130a-3p lead to a deregulation of some pathways involved in metastasis, in angiogenesis, such as in coagulation. In conclusion, the identified risk score integrating circulating markers can help clinicians predict early-stage NSCLC patients who are more likely to relapse after initial surgery.

overall_design: A total of 67 resected NSCLC samples were analyzed to define a miRNA signature capable to predict patients who experience progression of the disease (PD).

first: Simona

last: Coco

first: Luca

last: Longo

first: Giovanni

last: Rossi

first: Giovanna

last: Chiorino

first: Francesca

last: Guana

first: Paola

last: Ostano


title: Gene expression (GeneChip) data from parental (wild-type) and CRMP2A KO (CRISPR) A549 cells.

city: Edmonton

first: Evangelos

middle: D.

last: Michelakis

state: Alberta

country: Canada

institute: University of Alberta

summary:
We sought to assess how permanent loss of the microtubule-associated protein CRMP2A could affect global gene expression in the A549 lung cancer cells.
We used microarrays to identify differentially regulated genes when CRMP2A is permanently removed from A549 cells using CRISPR/Cas9.

overall_design: Parental (wild-type) and CRMP2A KO (CRISPR) cells were cultured under identical conditions for RNA extraction and hybridization on Affymetrix microarrays. We sought to compare the gene expression profiles between the two cell lines and correlate it with other in vitro and in vivo findings.

first: Aristeidis

last: Boukouris


title: RNA-Seq Analysis of Wild Type and Elovl2C234W Mouse Retinal Transcriptomes

city: Irvine

first: Dorota

last: Skowronska-Krawczyk

state: CA

address: 837 Health Sciences Rd

department: Physiology & Biophysics

institute: University of California, Irvine

summary: Purpose:  The lipid elongation enzyme ELOVL2 is a molecular regulator of retina aging in both human and mice. The point mutation Elovl2C234W in mice disrupts Elovl2-specific enzymatic activity and results in loss of ELOVL2-specific function. The goal of this study is to detect the roles of gene Elovl2 in transcription regulation in mouse retina by high-throughput transcriptome data analysis in the gene loss-of-function model.

overall_design: Fresh bulk retina tissue of both Wild Type and Elovl2C234W mice in triplicate were homogenized thoroughly in TRIzol Reagent (15596026, Invitrogen) followed by RNA isolation. The poly-adenylated (poly-A) transcripts from the isolated total RNA sample were enriched and performed strand specific mRNA-Seq TruSeq libraries preparation followed by Next Generation Sequencing. mRNA-Seq raw reads data was mapped to mm10 assembly mouse genome by homer (v4.11) aligner tool STAR. The uniquely mapped tags in exons on minus strand (one isoform per locus) were quantified by analyzeRepeats.pl. Raw reads count was normalized to experiment totals and performed differential expression by DESeq v1.24.0 program.

first: Qianlan

last: Xu


title: KrasG12D/+ co-mutation alters the transcriptional landscape of lung adenocarcinoma

city: Parkville

first: Waruni

last: Abeysekera

state: Victoria

address: 1G Royal Parade

department: Bioinformatics

country: Australia

institute: Walter and Eliza Hall Institute of Medical Research

summary: The tumor microenvironment (TME) contains a rich source of nutrients that sustain cell growth and ultimately facilitate tumor progression. The availability of glucose and glutamine in the TME are essential for the development and activation of effector T cells that exert anti-tumor function. Recently, inhibition of glutaminase, the enzyme that hydrolyzes glutamine to glutamate, has garnered interest as an approach to both decrease tumor metabolism and growth, while increasing available glutamine in the TME for effector T cells. Checkpoint blockade immunotherapy unleashes anti-tumor effector capabilities of T cells, and although there are good responses in many solid tumors, a significant proportion of patients respond poorly. In lung adenocarcinoma, response to immunotherapy is reported to be impaired in KRAS-mutant patients harboring concurrent KEAP1 and STK11/Lkb1 mutations. To investigate the metabolism and immune microenvironment of KRAS-mutant lung adenocarcinoma, we generated a series of murine models that reflect the KEAP1 and STK11/Lkb1 mutational landscape seen in patients. Here we show increased glutamate abundance in the Lkb1-deficient TME associated with CD8 T cell activation in response to anti-PD1 checkpoint immunotherapy. Combination treatment with the glutaminase inhibitor CB-839 inhibited clonal expansion and cytotoxic activation of tumor-infiltrating CD8 T cells. Thus, CD8 T cells exposed to checkpoint immunotherapy have a dependence on glutamine and reliance on glutaminase activity and are negatively impacted by the glutaminase inhibitor in this highly activated state. Therefore, we discern that the combination of immunotherapy and glutaminase inhibition is not efficacious for CD8 T cell activation in the tumor microenvironment.

overall_design: Mice of defined genotypes (n=5 mice per genotype) were administered intranasal Ad5-CMV-Cre to recombine genetic alleles. When moribund with breathing difficulties, tumor nodules were removed (2 independent nodules per mouse) and flash frozen in liquid nitrogen. Uninfected littermate control lungs were used as the "normal lung tissue". Lungs and tumor nodules were crushed in liquid nitrogen and RNA extracted using a combination of the QIAshredder and RNeasy kit.

first: Jackson

last: McDonald

first: Sarah

last: Best

first: Kate

last: Sutherland


title: A deleterious impact on the C. elegans fertility over generation after a chronic exposure to ionizing irradiation: A multigenerational transcriptomic evidence

city: Saint Paul lez Durance

first: olivier

last: ARMANT

address: Cadarache BP3

department: SERIS

country: France

institute: IRSN

summary: Parental exposure to environmental stress factors during gestation as well as embryonal developmental  may lead to deleterious consequences onto the offspring. In the present study, we assessed the adverse effects and mode of toxic action of chronic exposure to ionizing radiation over 4 generations in Caenorhabditis elegans (C. elegans). Our data show a decrease of 23% in the number of offspring on the first generation F0 and a decrease of more than 40% in subsequent generations F1, F2 and F3 from parental exposure.Transcriptomic analysis  revealed modulation of a common set of genes functions betweenthrough generations  that included transcription factors, as well as genes involved in stress response, unfolded protein response, lipid metabolism and reproduction. Furthermore, a drastic increase in the number of differentially expressed genes involved in defense response was measured in the last two generations, suggesting a cumulative stress effect of ionizing radiation exposure.  Integration of these multi-scale data demonstrates that chronic gamma exposure can impact the germline and induce an increase in defense mechanisms, including apoptosis, antioxidant enzymes, and a modulation of lipid metabolism  to improve resistance to stress. Harmful effects  intensify over generations, after F0 at the phenotypic level F0 and F1 at the transcriptomic level , emphasizing the importance of studying effects of chronic exposure to pollutants over several generations.

overall_design: RNAseq analysis of C elegans exposed chronically to 50 mGy/h of gamma ionizing radiation over 4 generations (exposed), or exposed over 1 generation and then grown under control conditions for a maximum of 3 generations (recovery), plus non exposed controls (control).  All experiments made in  3 biological replicates.

first: Remi

last: Guedon

first: Catherine

last: Lecomte

first: Anders

last: Bred

first: Olivier

last: Armant


title: Multi-Dimensional Profiling Reveals a Molecularly Distinct Injury-Responsive Population of Regulatory T Cells

city: Boston

first: James

last: Lederer

address: 20 Shattuck Street

institute: Brigham and Women's Hospital


title: A bacterial macrolide acts as signalling molecule and rewires the fungal metabolism in the Streptomyces – Aspergillus interaction

city: Tulln

first: Harald

last: Berger

state: NOE

address: Konrad-Lorenzstraße 43

department: DAGZ

country: Austria

institute: Universität f. Bodenkultur Wien

summary: Aspergillus nidulans competes in its natural environment with other microorganisms. It has been known that the bacterium Streptomyces rapamycinicus induces the production of Orsellinic acid in A. nidulans and we found that the major trigger of this induction is Polaramycin B produced by S. rapamycinicus. Here we show the transcriptome of A. nidulans following treatment with 0.5µg/ml Polaramycin B after 30 minutes and 3 hours.

overall_design: An overnight culture of A. nidulans was treated for 0 hours, 0.5 hours and 3 hours with Polaramycin B (0.5µg/ml ); each experiment is replicated

first: Markus

last: Bacher

first: Roman

last: Labuda

first: Isabel

last: Eppel

first: Florentina

last: Bayer

first: Michael

last: Sulyok

first: Erika

last: Gasparotto

first: Maria

last: Doppler

first: Hannes

last: Gratzl

first: Alisha

last: Buzzi

first: Joseph

last: Strauss


title: Uncoupling of IL-6 signaling and LC3-associated phagocytosis (LAP) drives immunoparalysis during sepsis

city: Groningen

first: Kieu

last: Le

state: Groningen

address: Antonious Deusinglaan 1

department: Genetics

institute: University Medical Centre Groningen

summary: Immune deactivation of phagocytes is a central event in pathogenesis of sepsis that remains molecularly unexplored. In particular, how cytokine deregulation induced by sepsis compromises the microbicidal activity of phagocytes and results in secondary infections by opportunistic bacterial and fungal pathogens is incompletely understood. Herein, we identify a master regulatory role of cytokine signaling on LC3-associated phagocytosis (LAP), and reveal that uncoupling of these two processes during sepsis induces immunoparalysis in monocytes/macrophages.

overall_design: Patients admitted to the ICU from April 2014 to December 2017 with clinical diagnosis of septic shock and APACHE II score >=8. Patients are sub-catergorized into three groups depending on the intact of LAP activities. RNA were isolated from blood samples for RNAseq profile.

first: Tonia

last: Akoumianaki

first: Katerina

last: Vaporidi

first: Eleni

last: Diamantaki

first: Frederic

last: Pene

last: Baeu

first: Mark

last: Gresnigt

first: Marina

last: Gkountzinopulou

first: Jame

last: El-Benna

first: George

last: Samonis

first: Vinod

last: Kumar

first: Dimitrios

last: Georgopoulos

first: Fank

last: van de Veerdonk

first: Mihai

last: Netea

first: Jean-Paul

last: Latge

first: Georgios

last: Chamilos


title: Multi-Dimensional Profiling Reveals a Molecularly Distinct Injury-Responsive Population of Regulatory T Cells [Bulk RNA-seq]

first: James

last: Lederer

summary: CD4+ regulatory T cells (Tregs) activate and expand in response to many different types of injuries, suggesting that they play a critical role in controlling the host response to tissue damage. Here we use multi-dimensional profiling techniques to comprehensively characterize injury responsive Tregs in mice.  CD44high Tregs expand in response to injury and are more suppressive than CD44low Tregs. T cell receptor (TCR) analysis reveal that CD44high Tregs undergo TCRαβ clonal expansion and increased CDR3 diversity in response to injury. Bulk and single-cell RNA sequencing with paired TCR clonotype analysis demonstrate that important Treg-associated genes were significantly differentially expressed in both CD44high and CD44low Tregs with expanded TCR clonotypes. CyTOF analysis verified protein expression of these genes on CD44high Tregs, with increases in Helios, Galectin-3 and PYCARD expression following injury. Collectively, these data provide new insights into CD44high Tregs, an important subset in the response to injury to tissue damage.

overall_design: Lymph node CD4+CD44highFoxP3-GFP+ and CD4+CD44lowFoxP3-GFP+ T cells or FoxP3-GFP+ T cells were purified by FACS sorting at 7 days after injury from injured or uninjured Foxp3DTR mice

first: Fei

last: Guo


title: Immune and Repair Responses in Joint Tissues and Lymph Nodes after Knee Arthroplasty Surgery in Mice

city: New York

first: David

last: Oliver

state: New York

address: 535 E 70th St

department: Research

institute: Hospital for Special Surgery

summary: The aim of this study was to better understand the resolution of inflammation and fibrosis in the soft tissue of the knee joint and the draining iliac lymph nodes (LNs) after surgery in a mouse tibial implant model. To do this, we collected the joint tissues from operated mice at 5 different timepoints (3, 7, 14, 21, and 35 days), plus joint tissue from the contralateral, non-operated limb at 35 days as a negative control. We also collected iliac LNs at days 7 and 14 post implantation surgery from the operated and contralateral sides; as controls we used LNs from non-operated mice. Transcriptomic analysis of the joint soft tissues revealed molecular pathways active during postoperative tissue repair.

overall_design: Transcriptomic analysis to understand molecular pathways active during postoperative tissue repair and in draining lymph nodes.

first: Lionel

last: Ivashkiv

first: Upneet

last: Sokhi


title: Transcriptome diversity in human medullary thymic epithelial cells

status: Public on Apr 30 2022

city: Cold Spring Harbor

first: Hannah

middle: Verena

last: Meyer

address: 1 Bungtown Road

institute: Cold Spring Harbor Laboratory


title: miRNA sequencing using Jurkat and JinB8 T cell line (CD47-deficient) and their Extracellular vesicles (EVs)

city: Bethesda

first: Sukhbir

last: Kaur

address: 10 Center Drive

department: NCI

institute: NIH

summary: Extracellular vesicles (EVs) mediate cell-cell communication including the intercellular transfer of miRNAs, which alter gene expression in target cells. Previously, we have shown that CD47 is also present on EVs and alters their effects on target cells, suggesting that specific surface markers define functionally distinct EVs. This hypothesis will be addressed by comparing total cellular versus EVs miRNA contents between Jurkat and JinB8 (CD47 deficient) T cells. The Jurkat T cell line (E6.1) was purchased from ATCC. The cells were maintained using RPMI 1640 containing 10% FBS, glutamine, penicillin, and streptomycin (Gibco). The Jurkat and JinB8 T cells used for experiments were maintained less than 6 weeks in culture. The cultured Jurkat were washed with IXPBs followed by a second wash with HITES medium (DMEM/F12 supplemented with, bovine serum albumin, hydrocortisone, insulin, transferrin, and trace elements). EVs were isolated from Jurkat T cells were cultured overnight using HITES medium. The conditioned media from Jurkat T cells were collected, and cell debris were removed using centrifugation at 300 g for 5 minutes and 2500 g for 10 minutes respectively. Centrifuged media were concentrated using Ultra-4 centrifugal filters to about 1 ml volume and further centrifuged at 10,000 g for 10 minutes. EVs were isolated using an Exo-Quick kit (SBI). miRNaseazy (Qiagen) kits were used to extract total RNA from Jurkat and JinB8 T cells and their EVs according to the manufacturer's instructions.

overall_design: Jurkat and JinB8 cells were cultured using serum-free HITES media and EVs were extracted. RNA was extracted using miRNA easy kit from Qiagen

middle: D

last: Roberts


title: Steroid nuclear receptor coactivator 2 controls immune tolerance by promoting induced Treg differentiation via upregulating Nr4a2

city: Duarte

first: ZUOMING

last: SUN

address: 1500 E Duarte Rd, Duarte, CA 91010

institute: City of Hope Medical Center

summary: Purpose:Steroid nuclear receptor coactivator 2 (SRC2) is a member of family of transcription co-activators. While SRC1 inhibits the differentiation of regulatory T cells (Tregs) critical for establishing immune tolerance, we show here that SRC2 stimulates Treg differentiation. SRC2 is dispensable for the development of thymic Tregs, whereas naïve CD4+ T cells from mice deficient of SRC2 specific in Tregs (SRC2fl/fl/Foxp3YFP-Cre) display defective Treg differentiation. Furthermore, the aged SRC2fl/fl/Foxp3YFP-Cre mice spontaneously develop autoimmune phenotypes including enlarged spleen and lung inflammation infiltrated with IFNγ-producing CD4+ T cells. SRC2fl/fl/Foxp3YFP-Cre mice also develop severer experimental autoimmune encephalomyelitis (EAE) due to reduced Tregs. Mechanically, SRC2 recruited by NFAT1 binds to the promoter and activates the expression of Nr4a2 which then stimulates Foxp3 expression to promote Treg differentiation. Members of SRC family co-activators thus play distinct roles in Treg differentiation and are potential drug targets for controlling immune tolerance

overall_design: RNA-seq analysis was performed to detect the transcriptome of Foxp3YFP-Cre and SRC2fl/fl/Foxp3YFP-Cre CD4+ cells 36 hours post Treg polarization.

first: Zuoming

last: Sun

first: Wencan

last: Zhang


title: Transcriptome diversity in human medullary thymic epithelial cells - 5'Cap sequencing

summary: The induction of central T-cell tolerance in the thymus depends on the  presentation of peripheral self-epitopes by medullary thymic epithelial cells (mTECs), enabled by a process known as promiscuous gene expression (pGE). Transcriptome diversity generated during pGE has many contributors, including non-canonical transcription initiation, alternative splicing and expression of endogenous retroelements (EREs). However, their significance and regulation are poorly understood in the healthy human thymus. Here, we mapped the expression of genome-wide transcripts in immature and mature human mTECs using high-throughput 5'Cap and RNA sequencing. Overall, 96% of protein coding genes were represented across five human mTEC samples, with mature mTECs showing increased rates of global transcript mis-initiation. Both mTEC populations have increased splicing entropy, which appears to be driven by expression of peripheral splicing factors. Furthermore, EREs enriched in long terminal repeat retrotransposons are up-regulated during mTEC maturation and enriched in genomic proximity to differentially expressed genes. We provide an interactive interface to explore the transcriptome diversity we uncovered at http://transcriptomediversity.cshl.edu/. Our findings represent an important first step towards the generation of a comprehensive map of transcriptome diversity in the healthy human thymus. Ultimately, a complete map of thymic expression diversity will allow for the identification of epitopes that contribute to the pathogenesis of auto-immunity and that drive immune recognition of tumor antigens.

overall_design: Genome-wide transcription start site analysis using 5'Cap sequencing data for human mature (MHCII high) and immature (MHCII) medullary thymic epithelial cells

middle: V

first: Jason

last: Carter


title: Expression data from mammary epithelia and mammary fibroblasts from mice with and without epithelial KrasG12D and with and without stromal p53 deletion

city: Charleston

first: Michael

last: Ostrowski

state: SC

address: 86 Jonathon Lucas Street

department: Biochemistry & Molecular Biology

institute: Medical University of South Carolina

summary:
p53 is a known tumor suppressor. We have found that stromal p53 regulates breast cancer development, the immune landscape, and survival in an oncogene specific manner when comparing the epithelial oncogenes ErbB2 and KrasG12D
In this dataset, we have evaluated mammary epithelia from mice with and without epithelial KrasG12D and with and without stromal p53 deletion. We have also evaluated mammary fibroblasts from mice with and without epithelial KrasG12D and with and without stromal p53 deletion.

overall_design: 32 total samples were analyzed. 16 epithelial cell isolates and 16 fibroblast isolates (4 biological replicates for each condition - with or without epithelial KrasG12D and with or without fibroblast p53 deletion using FspCre and p53fl/fl alleles)

first: Blake

last: Hildreth III


title: Tumor rejection in Cblb-/- mice depends on IL-9 and Th9 cells

city: Innsbruck

first: Dominik

last: Wolf

address: Anichstrasse 35

department: Dpt. Hematology and Oncology

institute: Medical University Innsbruck

summary: Casitas B lymphoma-b (Cbl-b) is a central negative regulator of cytotoxic T and NK cells, explaining its intracellular checkpoint function in cancer. Increasing evidence highlights the importance of CD4+ T helper cell populations (e.g., IL-9 producing T cells: Th9 cells) for anti-cancer immunity. We here provide experimental evidence that Cbl-b acts as rheostat favoring regulatory T cells (Treg) at the expense of Th9 cell differentiation. Adoptively transferred tumor-model antigen specific Cblb-/- Th9 cells exert superior anti-tumor activity leading to improved melanoma control in vivo. Accordingly, blocking IL-9 in melanoma cell-exposed Cblb-/- mice reverses their tumor rejection phenotype. Single cell RNA sequencing of in vitro differentiated Th9 cells from naïve T cells isolated from WT and Cblb-/- animals revealed the transcriptomic basis for increased Th9 cells differentiation. In summary, we here first establish IL-9 and Th9 cells as key anti-tumor executers in Cblb-/- animals. This knowledge may be helpful for future improvement of adoptive T cell therapies in cancer.

overall_design: in vitro differentiated Th9 cells from wild type (n=3) and Cblb-/- (n=3) mice

first: Oliver

last: Schanz

first: Isabelle

last: Cornez

first: Sowmya

last: Parampalli Yajnanarayana

first: Friederike

middle: S

last: David

first: Sebastian

last: Peer

first: Thomas

last: Gruber

first: Peter

middle: M

last: Krawitz

last: Brossart

first: Gottfried

last: Baier


title: Rice root response atlas

city: Riverside

first: Mauricio

middle: A

last: Reynoso

address: 4126 Genomics Bldg

department: Botany and Plant Sciences

institute: University of California Riverside

summary: Rice has evolved regulatory programs and specialized cell types that allow the plant to withstand different environments. To understand how rice root systems cope with water stresses, we profiled translatomes (ribosome-associated mRNAs) and accessible chromatin of developmentally-defined root cell populations from well-watered and drained control (aerobic control), water deficit, waterlogged, fully submerged plants and recovery conditions.  Whereas, the waterlogging responses are limited to specific root domains, water deficit and submergence signatures are extensive, and mostly reversible after 1 day of  recovery, relative to control roots. Root systems were also evaluated in rice cultivated in a paddy field. Specific responses include a halt in the cell-cycle and DNA synthesis-related genes translation in meristematic tissue under submergence and exo/endodermis suberin-related pathways bolstering under water deficit. Chromatin accessibility and translatome data integration was used to generate inferred regulatory networks that are dynamically regulated by changing water availability. The data collection is further enriched by translatome and chromatin accessibility data for the root systems of plate-grown seedlings (7 day old) and those cultivated in a paddy field (49 day old). An atlas of eight cell population translatomes for field-grown plants exhibited robust cell type expression. Collectively, these data for specific cell populations at multiple developmental ages and in multiple environments including growth two limiting water stresses will serve as a community resource.

overall_design: Evaluation of root domains TRAP seq and ATAC-seq profiles in rice roots in different environments and water stresses. Greenhouse pot-based translating ribosome affinity purification and chromatin experiments were conducted in five independent replicates of each line, with T1 or T2 seed stocks and whole root tissue was quickly dissected and frozen in liquid nitrogen. All tissue collection and TRAP and INTACT-ATAC library processing was performed with three biological replicates. Complete root systems were harvested. Nine conditions were assessed. Plate: plants grown from seed on petri dishes with 1X MS, 1 % [w/v] Sucrose, pH 5.8. Plate-based TRAP experiments were conducted with three or four independent replicates. Greenhouse grown plants were cultivated in GreenGrades Profile with seven treatments. (1) Control: controlled well-watered and drained irrigation growth. (2) Water Deficit: 5-6 days of no watering, with a fall to 72% (v/v) relative water content. (3) Waterlogging: 5 days of standing water covering root systems. (4) Submergence: 5 days of complete plant submergence. (5) Water Deficit Recovery: water deficit followed by a 1 day irrigation. (6) Waterlogging Recovery: pots allowed to fully drain for 1 day. (7) Submergence Recovery: plants returned to the bench and pots allowed to drain for 1 day. Field: Seedlings grown in the greenhouse transplanted to a paddy field at age 14 days; tissue collected after 35 days. This was performed with six randomized independent replicates of each line. For greenhouse and field sampling, 3 bioreplicates of up to 2 RNA pools (total mRNA, TRAP-RNA for different root domains) and ATAC-seq.

first: Germain

last: Pauluzzi

first: Julia

last: Bailey-Serres


title: ERAP2 is a novel target involved in autophagy and activation of pancreatic stellate cells via unfolded protein response signaling pathway

city: Fukuoka

first: Weiyu

last: Guan

state: Fukuoka

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

department: Department of Surgery and Oncology

institute: Kyushu university, Graduate school of medical science

summary: Pancreatic ductal adenocarcinoma (PDAC) is characterized by excessive desmoplasia and autophagy-dependent tumorigenic growth. Pancreatic stellate cells (PSCs) as a predominant stromal cell type play a critical role in PDAC biology. Autophagy facilitates PSC activation. However, the mechanism remains unknown. To investigate the mechanism of autophagy in PSC activation, gene expression profiles between patient-derived PSCs from pancreatic cancer and chronic pancreatitis were compared using a gene expression microarray. Here, we found that endoplasmic reticulum aminopeptidase 2 (ERAP2), which resides in the endoplasmic reticulum (ER) membrane, was highly expressed in both cancer-associated PSCs and pancreatic cancer cells (PCCs). We found that high stromal ERAP2 expression is associated with a poor prognosis of PDAC patients. Knockdown of ERAP2 inhibited autophagy of PSCs and PCCs. In PSCs, inhibition of autophagy by ERAP2 knockdown led to inactivation of PSCs and attenuated tumor-stromal interactions. This process was mediated by ER stress and consequent IRE1α and PERK unfolded protein response (UPR) signaling pathways. In orthotopic models, ERAP2 knockdown in PSCs inhibited growth and fibrosis of xenografted tumor compared with coimplantation of PSCs without ERAP2 knockdown, and gemcitabine treatment further inhibited tumor growth. Our findings demonstrate a novel mechanism of PSCs activation regulated by autophagy. ERAP2 as a promising therapeutic target may provide a novel strategy for the treatment of PDAC.

overall_design: Human primary cultured PSCs were established from fresh surgical specimens of patients with pancreatic cancer or chronic pancreatitis using the outgrowth method.

first: Kohei

last: Nakata