title: Identification of binding sites of the Six1 transcription factor in mouse primary myoblasts and myotubes

city: Ottawa

first: Alexandre

last: Blais

address: 451 Smyth Road

institute: University of Ottawa

summary: Binding sites of the Six1 transcription factor were identified in primary myoblasts and myotubes of mice using ChIP-sequencing

overall_design: Four samples were obtained: anti-Six1 ChIP-seq in primary mouse myoblasts; anti-Six1 ChIP-seq in primary mouse myotubes; normal rabbit IgG ChIP-seq; input chromatin. No biological replicates were obtained

first: Imane

last: Chakroun

first: Yubing

last: Liu


title: Age-related changes in blood and spleen-derived immune cells in long and short lived mice strains [RNA-seq]

status: Public on Dec 01 2021

city: Farmington

first: Duygu

last: Ucar

state: Connecticut

address: 10 Discovery Drive

country: USA

institute: The Jackson Laboratory

type: Expression profiling by high throughput sequencing

summary: Immune system aging contributes significantly to declining health with age. It is unknown to what extent human and mice immune systems go through similar age-related changes and whether there are conserved biomarkers of aging. We characterized age-related changes in the immune system of long (C57BL/6J) and short-lived (NZO/HILtJ) strains and compared them with blood-driven human aging signatures. Peripheral blood lymphocytes (PBL), spleen cells and spleen-driven naive and memory CD8+ T cells were profiled using flow cytometry, RNA-seq and ATAC-seq from young (3 months) and old (18 months) mice. Pro-inflammatory myeloid genes were activated with age across strains and tissues, echoing human 'inflammaging' signatures. ATAC-seq footprinting analyses uncovered increased binding of pro-inflammatory transcription factors with age (e.g., AP1 complex, NFKB signaling pathway). Machine learning models identified inflammation, cytotoxic, and naive T cell derived genes to be strong signatures of immunosenescence. These data are publicly shared as a resource for immune aging (https://immune-aging.jax.org/mice).

overall_design: Spleen, PBL and spleen sorted CD8+ T cell profiles of B6 and NZO mice at 3,12 and 18 months

first: Onur

middle: E

last: Karakaslar

first: Emily

last: Lachatara

first: Siddhartha

last: Sharma


title: Age-related changes in blood and spleen-derived immune cells in long and short lived mice strains [ATAC-seq]

type: Genome binding/occupancy profiling by high throughput sequencing


title: Chromatin accessibility sequencing (ATAC-seq) after adipogenesis induction of 3T3-L1 cells

status: Public on Nov 19 2021

first: Michael

middle: Joseph

last: Guertin

state: CT

address: 400 Farmington Ave

department: Center for Cell Analysis and Modeling

institute: University of Connecticut

summary: These experiments identify the early and terminal changes in chromatin accessibility that results from treating cells with 0.25 U/mL Insulin, 250 nM dexamethasone, and 0.5 mM 3-isobutyl-1-methylxanthine

overall_design: Cells were treated with this differentiation cocktail for 20 min, 40 min, 60 min, 2 hr, 3 hr, 4hr, and 6 days in triplicate

first: M

middle: J


title: Myeloid cell subsets that express latency-associated peptide promote cancer growth by modulating T cells

status: Public on Nov 24 2021

city: Cambridge

first: Dunaduan

last: Ma

state: Ma

address: 77 Massachusetts Ave

department: Koch Institute

institute: MIT

summary: Myeloid suppressor cells promote tumor growth by a variety of mechanisms which are not fully characterized. We identified myeloid cells (MCs) expressing the latency-associated peptide (LAP) of TGF-b on their surface, LAPHi MCs, that stimulate Foxp3+ Tregs while inhibiting effector T cell proliferation and function. Blocking TGF-b inhibits the tolerogenic ability of LAPHi MCs. Furthermore, adoptive transfer of LAPHi MCs promote Treg accumulation and tumor growth in vivo. Conversely, anti-LAP antibody, which reduces LAPHi MCs, slows cancer progression. Single-cell RNA-Seq analysis on tumor-derived immune cells revealed LAPHi dominated cell subsets with distinct immunosuppressive signatures, including those with high levels of MHCII and PD-L1 genes. Analogous to mice, LAP is expressed on myeloid suppressor cells in humans, and these cells are increased in glioma patients. Thus, our results identify a previously unknown function by which LAPHi MCs promote tumor growth and offer therapeutic intervention to target these cells in cancer.

overall_design: single-cell RNA-seq experiment characterizing myeloid cells expressing surface LAP in the context of cancer. Experiment was performed with four biological replicates.

first: Galina

last: Gabriely

first: Howard

middle: L

last: Weiner


title: CUX1 deficiency potentiates RAS signaling to drive malignancy

status: Public on Nov 16 2021

city: Chicago

first: Megan

middle: E.

last: McNerney

state: Illinois

address: 900 East 57th Street, KCBD 5128

department: Pathology

institute: The University of Chicago

summary: -7/del(7q) is genetic event prevalent in high-risk myeloid neoplasms. For reasons that are unclear, gain-of-function mutations in the RAS pathway frequently co-occur with monosomy 7.  Here we identify a genetic interaction between RAS and the 7q-encoded transcription factor, CUX1.  Concomitant mutations in RAS genes and CUX1 are wide-spread across tumor types, suggesting cooperativity in tumorigenesis.   To test this, we generated mice with oncogenic NrasG12D and Cux1 knockdown. Double mutant mice developed myeloid malignancies with higher penetrance and faster onset than either single allele alone, with leukemic transformation in one third of cases. Oncogenic RAS imparts increased self-renewal on CUX1-deficient hematopoietic stem and progenitor cells (HSPCs).  Reciprocally, CUX1 knockdown amplifies RAS signaling through decreased transcriptional expression of negative regulators of RAS and PI3K signaling.  Accordingly, NrasG12D;Cux1-knockdown HSPCs have heighted growth factor-sensitivity and downstream RAS pathway activation.  Double mutant HSPCs were responsive to PIK3 or MEK inhibition, suggesting that these may be promising therapeutic targets in patients with 7/del(7q) malignancies.  Our results implicate the loss of CUX1 as the underlying explanation for the association of -7/del(7q) with oncogenic RAS.  Furthermore, we report the unexpected convergence of an oncogene and tumor suppressor gene on the same pathway.

overall_design: RNA-seq of HSPCs from Cux1-low and Cux1-low;NrasG12D mice


title: Hierarchical chromatin regulation during blood formation uncovered by single-cell sortChIC

status: Public on Nov 30 2021

city: Utrecht

first: Jake

last: Yeung

address: Uppsalalaan 8

country: Netherlands

institute: Hubrecht Institute

summary: Post-translational histone modifications modulate chromatin packing to regulate gene expression. How chromatin states, both at euchromatic and at heterochromatic regions, underlie cell fate decisions in single cells is relatively unexplored. We develop sort assisted single-cell chromatin immunocleavage (sortChIC) and apply it to map active (H3K4me1 and H3K4me3) and repressive (H3K27me3 and H3K9me3) histone modifications in hematopoietic stem and progenitor cells (HSPCs), and mature blood cells in the mouse bone marrow. During differentiation, HSPCs acquire distinct active chromatin states that depend on the specific cell fate, mediated by cell type-specifying transcription factors. By contrast, most regions that gain or lose repressive marks during differentiation do so independent of the specific cell fate. Joint profiling of active H3K4me1 and repressive H3K9me3 in single cells demonstrates that cell types within the myeloid lineage have distinct active chromatin regulation but share a similar myeloid-specific heterochromatin-repressed state. Our results suggest a hierarchical chromatin regulation program during hematopoiesis, by which heterochromatin dynamics define differentiation trajectories and lineages, while euchromatin dynamics establish cell types within lineages.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: single-cell sortChIC data on K562 and mouse bone marrow cells

first: Peter

last: Zeller

first: Helena

last: Viñas Gaza

first: Buys

last: de Barbanson

first: Vivek

last: Bhardwaj

first: Maria

last: Florescu

first: Alexander

last: van Oudenaarden


title: Genome-wide binding (ChIP-Seq) of histone-like protein (HstA) of Haloferax volcanii

status: Public on Oct 29 2021

city: Durham

first: Amy

last: Schmid

state: NC

address: 3242 French Family Science Center

institute: Duke University

summary: Diverse studies including protemoics, genome-wide binding, and transcriptional profiling of the model halophile Halobacterium salinarum suggest that its putative histone protein acts not as a chromatin protein but a direct and indirect transcriptional regulator. Here, we characterise the putative histone (HstA) of another model halophile (Haloferax volcanii) with ChIP-Seq to understand its genome-wide binding, and compare it with binding patterns seen from histones, nucleoid-associated proteins, and transcription factors of Halobacterium salinarum, other archaea, and eukaryotes. Analysis of this data by visual inspection, start site occupancy profiles, DNA motif searching, and dinucleotide periodicity suggests that the binding mode of halophilic histones shares features with TFs, NAPs, and more typical archaeal/eukaryotic histones.

overall_design: Examination of HstA binding in optimal conditions (rich media, 42⁰C, mid-exponential growth phase) using C-terminal HA-tagged HstA expressed in-trans under its native promoter.

first: Saaz

last: Sakrikar


title: Quantitative transcriptomic analysis of Wild Type and DfosDN macrophages of Drosophila melanogaster embryos

status: Public on Nov 10 2021

city: Klosterneuburg

first: Daria

last: Siekhaus

address: Am Campus 1

department: Bertalanffy Building

country: Austria

institute: IST Austria

summary: RNA seq data of macrophages FACS sorted from wild type Drosophila melanogaster embryos and those expressing DfosDN in macrophages

overall_design: mRNA profiling and differential expression analysis of FACS-sorted Wild-Type and DfosDN macrophages from Drosophila melanogaster embryos from stage 11

first: Vera

last: Belyaeva


title: Regulation of alternative splicing by C9ORF78, a novel BRR2 interactor

status: Public on Dec 31 2021

city: Berlin

first: Marco

last: Preussner

address: Takustrasse 6

country: Germany

institute: FU berlin

summary: The RNA helicase BRR2 (SNRNP200) is one of the key remodeling factors of the spliceosome. Here we show its direct interaction with C9ORF78, a poorly characterized protein predicted to be largely intrinsically disordered. We present cryo-EM structures showing how C9ORF78 and the spliceosomal B-complex protein FBP21 wrap around the C-terminal helicase cassette of BRR2 and that binding of the two proteins is mutually exclusive. C9ORF78 associates with the spliceosome, as we confirm via proteomics and RNA UV-crosslinking. An siRNA mediated C9ORF78 knockdown reveals changes in alternative splicing of specific target pre-mRNAs, which in part depend on its interaction with BRR2. In particular, C9ORF78 regulates a substantial number of alternative 3’ splice sites, which might be facilitated through an additional interaction with human PRP22 (DHX8).

overall_design: HEK293T cells were transfected with Rotifect either with a CTRL siRNA or an siRNA targeting C9orf78; 72h after transfection RNA was extracted and analyzed by RNA-Seq.

first: Florian

last: Heyd

first: Alexandra

last: Bergfort

first: Markus

middle: C

last: Wahl


title: Immune-complex activated gasdermin D facilitates mitochondrial DNA membrane translocation and extracellular release in systemic lupus erythematous [RNA-Seq]

status: Public on Nov 08 2021

city: Shanghai

first: Zhuning

last: Wang

address: 280 Chong Qing Nan Road

country: China

institute: Shanghai Jiao Tong University School of Medicine

summary: The purpose of this study was to study the roles of GSDMD in systemic lupus erythematous. Here, we used RNA sequencing from wild type, Pristane induced wild type and Pristane induced GSDMD knockout mice to detect changes in transcriptomes of kidneys.

overall_design: RNA sequencing libraries were generated for kidneys of wild type, Pristane induced mice and GSDMD knockout mice.

first: Naijun

last: Miao

first: Jing


title: Self-organized stem cell-derived human lung buds with proximo-distal patterning and novel targets of SARS-CoV-2

status: Public on Dec 21 2021

city: New York

first: Edwin

middle: Antonio

last: Rosado-Olivieri

state: NY

address: 1230 York Ave

institute: The Rockefeller University

summary: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the global COVID-19 pandemic and the lack of therapeutics hinders pandemic control. Although lung disease is the primary clinical outcome in COVID-19 patients, how SARS-CoV-2 induces tissue pathology in the lung remains elusive. Here we describe a high-throughput-based platform to generate tens of thousands of self-organizing, nearly identical, and genetically matched human lung buds derived from human pluripotent stem cells (hPSCs) cultured on confined geometries on micropattern chips. Strikingly, in vitro-derived human lung buds resemble fetal human lung tissue and display in vivo-like proximo-distal coordination of alveolar and airway tissue differentiation whose 3D epithelial self-organization is directed by the levels of KGF. Single-cell transcriptomics unveiled the cell identities and ontogeny of airway and alveolar tissue and the specification of WNThi cycling alveolar stem cells from alveolar progenitors. These synthetic human lung buds are susceptible to SARS-CoV-2 infection and can be used to track cell type-dependent susceptibilities to infection, intercellular transmission and cytopathology in airway and alveolar tissue in individual synthetic lung buds.  We detected an increased susceptibility to infection in alveolar cells and identified cycling alveolar stem cells as targets of SARS-CoV-2. We used this platform to test neutralizing antibodies isolated from convalescent plasma that efficiently blocked SARS-CoV-2 infection and intercellular transmission. Our platform offers unlimited, rapid and scalable access to disease-relevant lung tissue that recapitulate human lung development and can be used to track SARS-CoV-2 infection and identify pre-clinical candidate therapeutics for COVID-19.

overall_design: Examination of cell type composition and heterogeneity in synthetic human lungs.

middle: A

first: Brandon

last: Razooky

first: Hans

middle: H

last: Hoffmann

first: Riccardo

last: De Santis

first: Charles

middle: M

last: Rice

first: Ali

last: Brivanlou


title: Identify Flag-bound transcripts via RIP-seq

status: Public on Oct 28 2021

city: Monrovia

first: Jianjun

last: Chen

state: CA

department: Systems Biology

institute: Beckman Research Institute of City of Hope

summary: To identify the directly bound transcripts of Flag antibody (the backgroud control for our METTL16 RIP-seq), RNA immunoprecipitation sequencing (RIP-seq) was conducted HEK293T. Briefly, HEK293T cells were infected with pmiRNA1-empty vector. Only the GFP-positive cells were used for study and expanded in DMEM medium.

overall_design: The GFP-positive HEK293T cells were expanded in 150mm dish and collected at 70-80% confluency. The were cross-linked by 0.75 % formaldehyde (F8775, Sigma-Aldrich) with gentle rotation at room temperature for 10 min and the reaction was quenched by 125 nM glycine (final concentration) with shaking at room temperature for 5 min. Then cells were collected, lysed, and subjected to RIP with Flag antibody. Two biological replicates were included to confirm the data quality.

first: Rui

last: Su

first: Lei

last: Dong


title: Determine the methyltransferase activities of METTL3, METTL14, and METTL16 via MeRIP-seq (m6A-seq)

summary: All the three enzymes, METTL3, METTL14, and METTL16, belong to methyltransferase like (METTL) family and possess the ability to deposit N6-methyladenosine (m6A) in mRNA. Via immunofluorescence staining and wertern blotting, we discovered either METTL3 or METTL14 mainly localize in nuclear, but METTL16 localizes in both cytosol and nuclei. To compare the m6A methyltransferase activities of the three m6A 'writers' and better understand their differences, MeRIP-seq (m6A-seq) was conducted with poly(A) RNAs isolaed from HEK293T cells with METTL3, 14 and 16 knockout.

overall_design: The HEK293T-Cas9 single clone cells were infected with sgMETTL3-3, sgMETTL14-3, sgMETTL16-3, and sgNS lentivirus to induce knockout of the m6A writers. The positive-infected cells were selected with 10ug/ml blasticidin (for Cas9) and 100ug/ml hygromycin (for sgRNAs) for 10 days. For each group, we included 2 biological replicates. The the cells were collected and total RNA were isolated from the whole cells via Qiazol. The poly(A) RNAs were further enriched from the total RNA and then subjected to m6A IP.


title: GABAergic neuronal IL-4R mediates T cell effect on memory [Il4rkoT]

status: Public on Oct 12 2021

city: Saint Louis

first: Jonathan

last: Kipnis

state: MO

address: 4515 McKinley Ave

department: Pathology and Immunology

institute: Washington University in Saint Louis

summary: The immune and nervous systems are highly interactive, however, little is known about how immune cells and their derived molecules impact brain function. Here we show that T cells play a critical role in retrieval of contextual fear conditioning (CFC) memory. Single-nuclei sequencing (snRNAseq) of engram neurons and non-engram neurons from the dentate gyrus one day after CFC learning revealed new evidence about differential expression of immune-related and synaptic plasticity-related genes, in SCID vs. wild-type mice. These findings present a comprehensive picture on how T cells and their derived cytokines could regulate memory retrieval and provide new insights into the nature of neuroimmune interactions at the transcriptional level in neurons.  Overall, restoration of T cells or IL-4R signaling might lead to selective rescue of memory retrieval, and proffer a valuable strategy for treating memory loss.

overall_design: This experiment aims to compare the transcriptional profile of nuclei from 2 genotypes (SCID+T cell(WT)/SCID+T cell(IL4KO)) and 2 activation states (engram/nonengram). All nuclei were collected from the dentate gyrus of mice after fear conditioning. All mice were housed under standard conditions and injected with labelled viruses to detect the activation state.

first: Jasmin

last: Herz

first: Zhongxiao

last: Fu

first: Taitea

last: Dykstra


title: GABAergic neuronal IL-4R mediates T cell effect on memory [Il4raFlox]

overall_design: This experiment aims to compare the transcriptional profile of nuclei from 2 genotypes (Gad2cre+::IL-4Rfl/fl/Gad2cre-::IL-4Rfl/fl)  and 2 activation states (engram/nonengram). All nuclei were collected from the dentate gyrus of mice after fear conditioning. All mice were housed under standard conditions and injected with labelled viruses to detect the activation state.


title: Determining the methyltransferase activities of METTL3, METTL14, and METTL16

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

overall_design: Refer to individual Series


title: Transcriptomic analysis of arcA difficient Acinetobacter baumannii

status: Public on Dec 11 2021

city: Tampa

first: Lindsey

middle: N

last: Shaw

state: FL

address: 4202 E Fowler Avenue

department: CMMB

institute: University of South Florida

summary: A. baumannii response to arcA disruption

overall_design: A. baumannii RNA was extracted to compare gobal gene expression of AB5075 wildtype to AB5075 arcA transposon mutant strain in the presence of sub-inhibitory concentrations of CCCP. Each sample was generated by pooling three independent biological replicate RNA preps.

first: Brooke

middle: R

last: Tomlinson


title: Determine METTL16-mediated spatiotemoral installation of m6A via MeRIP-seq (m6A-seq) with nascent RNA and nuclear poly(A) RNA

summary: METTL16 belong to methyltransferase like (METTL) family and possesses the ability to deposit N6-methyladenosine (m6A) in mRNA. We have conducted m6A-seq with poly(A) RNAs isolated from the whole HEK293T cells upon METTL16 knockdown to identify the transcripts with hypo m6A peaks after METTL16 knockdown, and also conducted RIP-seq with HEK293T cells to determine the METTL16-bound transcripts. However, we found that the transcripts with hypo m6A peaks upon METTL16 knockdown only account for a small proportion of METTL16-bound transcripts. We suppose one of the possibility reasons might be due to  spatiotemporal installation of m6A. To support our hypothesis, we performed additional m6A seq with nascent RNA and nuclear poly(A) RNA isolated from HEK293T cells with METTL16 knockdown.

overall_design: The HEK293T cells were infected with shMETTL16-1, shMETTL16-2, and shNS lentivirus to induce knockdown of METTL16. The positive-infected cells were selected with 2ug/ml puromycin for 1-2 weeks. For purification of nascent RNA, all the cells were pulsed with 4-thiouridine (4sU) for 1 hours before collection. The nuclear pellets were collected with buffer RLN and then nuclear RNAs were extracted. The nascent RNAs were enriched and subjected to m6A IP. Poly(A) RNAs were enriched from nuclear RNAs and then subjected to m6A IP. Two different shRNAs against METTL16 were included to ensure the repeatibility and avoid potential off-target effects.


title: VprBP directs epigenetic gene silencing through H2A phosphorylation in colon cancer

status: Public on Oct 06 2021

city: Los Angeles

first: Nikhil

middle: Baban

last: Ghate

address: 1450 Biggy st, Norris Research Tower

department: Biochemistry and Molecular Medicine

institute: University of Southern California

summary: Histone modification is aberrantly regulated in cancer and generates an unbalanced state of gene transcription. VprBP, a recently identified kinase, phosphorylates histone H2A on threonine 120 (T120) and is involved in oncogenic transcriptional dysregulation; however, its specific role in colon cancer is undefined. Here we show that VprBP is overexpressed in colon cancer and directly contributes to epigenetic gene silencing and cancer pathogenesis. Mechanistically, the observed function of VprBP is mediated through H2AT120 phosphorylation (H2AT120p)-driven transcriptional repression of growth regulatory genes, resulting in a significantly higher proliferative capacity of colon cancer cells. Our preclinical studies using organoid and xenograft models demonstrate that treatment with VprBP inhibitor B32B3 impairs colonic tumor growth by blocking H2AT120p and reactivating transcriptional program resembling that of normal cells. Collectively, our work describes VprBP as a master kinase contributing to the development and progression of colon cancer, making it a new molecular target for novel therapeutic strategies.

overall_design: mRNA-seq of scrambled (n=3) and VprBP-knocked down (n=3) in colon cancer cell line (SW620)

first: Suhn

middle: K

last: Rhie

first: Woojin

last: An


title: Temporal single cell tracing reveals clonal revival and expansion of precursor exhausted T cells during anti-PD-1 therapy in lung cancer

city: Beijing

first: Zemin

last: Zhang

address: No. 5, Summer Palace Road, Haidian District, Beijing

department: BIOPIC

institute: Peking University

summary: Immune-checkpoint therapies have shown unprecedented clinical success in the treatment of non-small-cell lung cancer, but the underlying mechanisms of anti-PD-1–induced tumour rejection remain incompletely understood. Here, we performed temporal single-cell RNA and paired T cell receptor sequencing on 47 clinical tumour biopsies from 36 non-small-cell lung cancer patients before and during combination therapies of PD-1 blockade with chemotherapy. We found that the treatment in responsive tumours preferentially increased the precursors of exhausted T cells (Tex), characterized by the low expression of co-inhibitory molecules and high expression of GZMK. By contrast, non-responsive tumours failed to accumulate Tex precursors (Texp), suggestive of the critical role of such cells in PD-1-based immunotherapies. Although these post-treatment Texp cells shared clonotypes extensively with terminal Tex subset prior to treatment, our data suggested that they were not derived from the reinvigoration of terminal Tex cells; instead, such Texp cells were likely accumulated by (i) the expansion of pre-existing Texp cells, and (ii) the replenishment with peripheral T cells, a phenomenon we named clonal revival. Furthermore, post-treatment Texp cells expressed high level of CXCL13, which was exclusively expressed by terminal Tex subset in treatment-naïve tumours, implying that the emergence of CXCL13-expressing Texp cells resulted from the blockade of the transition to terminal Tex cells by PD-1 antibodies. Our study provides new insights into mechanisms underlying PD-1-based therapies, implicating both clonal revival and expansion of Texp cells as steps to enhance the clinical response of lung cancer patients.

overall_design: Single-cell RNA-seq and single-cell TCR-seq of pre- and post-treatment tumour biospies (n=47) from 36 non-small-cell lung cancer patients, spanning 33 treatment-naïve, 9 site-matched post-treatment responsive (from 8 tumours), and 5 post-treatment non-responsive tumour specimens following combination therapies of PD-1 blockade with chemotherapy.

first: Baolin

last: Liu


title: Modeling Malignant Progression in Glioma

status: Public on Sep 30 2021

city: Houston

first: Akdes

last: Serin Harmanci

state: TX

address: One Baylor Plaza

institute: Baylor College of Medicine

summary: Gliomas are the most common primary brain tumor in humans. Low-grade gliomas (WHO grade II) invariably progress to high-grade gliomas (WHO grade III or IV). Although malignant progression may take many years, the survival rate after transformation to a high-grade glioma is poor, often only 12-15 months. In this data set, we have identified low-grade gliomas that have progressed to high-grade gliomas or high-grade gliomas that have progressed from low-grade gliomas. Some cases are matched pairs (meaning we have both the original low-grade tumor and the subsequent high-grade tumor). The samples deposited have been analyzed with bulk-RNA sequencing. They are also de-identified but are clinically annotated. When available, genetic information including IDH mutation status, 1p/19q deletion and histological subtype are also included.

overall_design: mRNA profiles of low- and high-grade human glioma samples

first: Ganesh

last: Rao


title: Cancer phylogenetics using single-cell RNA-seq data

status: Public on Sep 26 2021

city: Dunedin

first: Jiří

last: Moravec

address: 133 Union Street East

department: Computer Science department

country: New Zealand

institute: University of Otago

summary: To test if scRNA-seq contains sufficient phylogenetic information to reconstruct a population history of cancer, immunosuppressed NU/J mice were injected with human cancer cells (MDA-MB-231-LM2). The tumors that develop are derived from the same population and thus share a common ancestor, but evolved independently in each mouse and should form separate clades on reconstructed phylogenetic trees when analysed together. We explore and compare results of phylogenetic analyses based on both expression levels and SNVs called from our scRNA-seq data. Both techniques are shown to be useful for reconstructing phylogenetic relationships between cells, refecting the clonal composition of a tumor. Without an explicit error model, standardized expression values appears to be more powerful and informative than the SNV values at a lower computational cost, due to being a by-product of standard expression analysis. Our results suggest that scRNA-seq can be a competitive alternative or useful addition to conventional scDNA-seq phylogenetic reconstruction. Our results open up a new direction of somatic phylogenetics based on scRNA-seq data. Further research is required to refne and improve these approaches to capture the full picture of somatic evolutionary dynamics in cancer.

overall_design: Phylogenetic analysis of 5 tumour samples from 3 individual mice seeded with human cancer cells derived from a single cell line.

first: Rob

last: Lanfear

first: David

last: Spector

first: Sarah

middle: D

last: Diermeier

first: Alex

last: Gavryushkin


title: The CTPase activity of ParB acts as a timing mechanism to control the dynamics and function of prokaryotic DNA partition complexes

status: Public on Sep 24 2021

city: Geneva 4

first: Patrick

middle: H.

last: Viollier

address: Rue Michel Servet 1

department: Dept.  Microbiology and Molecular Medicine

country: Switzerland

institute: University of Geneva, Faculty of Medicine / CMU

summary: DNA partitioning CTPases of the ParB family mediate the segregation of bacterial chromosomes and low-copy number plasmids. They act as DNA-sliding clamps that are loaded at parS motifs in the centro-meric region of target DNA molecules and then spread laterally to form large nucleoprotein complexes that serve as docking points for the DNA segregation machinery. Here, we identify conformational changes that underlie the CTP- and parS-dependent closure of ParB clamps. Moreover, we solve crystal structures of ParB in the pre- and post-hydrolysis state and provide insight into the catalytic mechanism underlying nucleotide hydrolysis. The characterization of CTPase-deficient ParB variants reveals that CTP hydrolysis serves as a timing mechanism to control the sliding time of ParB. Hyperstable clamps are trapped on the DNA, leading to excessing spreading and severe chromosome segregation defects in vivo. These findings clarify the role of the ParB CTPase cycle in partition complex dynamics and function and thus complete our understanding of this prototypic CTP-dependent molecular switch.

overall_design: Examination of ParB whole genome binding/occupancy (ChIP-Seq) in the model organism Myxococcus xanthus

first: Manuel

last: Osorio-Valeriano

first: Florian

last: Altegoer

first: Chandan

last: Kumar Das

first: Wieland

last: Steinchen

first: Gaël

last: Panis

first: Giacomo

last: Giacomelli

first: Helge

last: Feddersen

first: Lara

last: Connolley

first: Laura

last: Corrales-Guerrero

first: Pietro

last: Giammarinaro

first: Juri

last: Hanßmann

first: Marc

last: Bramkamp

first: Seán

last: Murray

first: Lars

last: Schäfer

first: Gert

last: Bange

first: Martin

last: Thanbichler


title: Mitochondrial pyruvate metabolism and glutaminolysis toggle steady-state and emergency myelopoiesis

status: Public on Sep 27 2021

city: Tucson

first: Deepta

last: Bhattacharya

state: Arizona

address: 1656 E. Mabel Street, MRB 217

department: Department of Immunobiology

institute: University of Arizona

summary: RNA-sequencing was performed to look for transcriptional differences between wild-type and Mpc2-deficient granulocyte-macrophage progenitors. Very few genes were dysregulated in knockout cells relative to wild-type. Gls Mpc2 double knockouts were additionally analyzed to examine extent of Gls and Mpc2 deletion relative to single knockouts. Paper abstract: Hematopoietic homeostasis is maintained by stem and progenitor cells in part by extrinsic feedback cues triggered by mature cell loss. We demonstrate a different mechanism by which hematopoietic progenitors intrinsically anticipate and prevent the loss of mature progeny through metabolic switches. We examined hematopoiesis in mice conditionally deficient in long-chain fatty acid oxidation (carnitine palmitoyltransferase 2, Cpt2), glutaminolysis (glutaminase, Gls), or mitochondrial pyruvate import (mitochondrial pyruvate carrier 2, Mpc2). While genetic ablation of Cpt2 or Gls minimally impacted most blood lineages, deletion of Mpc2 led to a sharp decline in mature myeloid cells. However, MPC2-deficient myeloid cells rapidly recovered due to a transient increase in myeloid progenitor proliferation. Competitive bone marrow chimera and stable isotope tracing experiments demonstrated that this proliferative burst was intrinsic to MPC2-deficient progenitors and accompanied by a metabolic switch to glutaminolysis. Thus, hematopoietic progenitors intrinsically adjust to metabolic perturbations independently of feedback from downstream mature cells to maintain homeostasis.

overall_design: RNA-sequencing of wild-type, Mpc2 knockout, Gls knockout, and Gls Mpc2 double knockout granulocyte-macrophage progenitors

first: Hannah

last: Pizzato


title: Transcriptome profiles of Col-0 and scl30a-1 germinating seeds

status: Public on Sep 19 2021

city: Cerdanyola del Vallès

first: Guiomar

last: Martín

state: Barcelona

address: Vall Moronta

country: Spain

institute: Centre for Research in Agricultural Genomics

summary: RNA-seq experiment of Col-0 and the scl30a-1 mutant seeds grown for 18 h in continuous white light.

overall_design: 50 mg of seeds of WT (Col-0) and scl30a-1 were stratified for 3 days at 4ºC and sown on MS medium. Seeds were collected at 18 h after stratification and RNA-seq libraries were prepared. Triplicate biological samples were analyzed.

first: Tom

last: Laloum

first: Paula

last: Duque


title: Evaluation of changes in microRNA expression in naïve and influenza-virus specific CD8+ T cells

status: Public on Sep 02 2021

city: La Jolla

first: Jennifer

last: Hope

address: 10901 N Torrey Pines Road

institute: Sanford Burnham Prebys Medical Discovery Institute

summary: MicroRNA microarray expression dataset evaluating relative changes in microRNA expression levels between naïve and effector OT-I CD8+ T cells during influenza virus infection in mice.

overall_design: Naïve OT-I CD8+ T cells were FACS sorted (based on CD44- and CD62L+ expression) from the spleens on uninfected female OT-I mice. Naïve cells were kept for analysis of microRNA expression in naïve OT-I CD8+ T cells. Additionally, 1x10e4 cells were adoptively transferred into uninfected female C57BL/6 mice (via tail vein injection) followed by intranasal infection with WSN-OVA influenza virus. At day 9, donor OT-I cells were FACS sorted from the lungs of influenza virus-infected mice for analysis of microRNA expression in effector OT-I CD8+ T cells. Total RNA was isolated from sorted cells and miRNA gene expression was assessed using an Affymetrix murine miRNA 4.0 gene chip. CEL files were normalized and analyzed using Affymetrix Expression Console, and miRNA annotation is from miRBASE version 21.

first: P

last: Katsikis

first: J


title: Elucidation of tumor-stromal heterogeneity and the ligand-receptor interactome by single cell transcriptomics in real-world pancreatic cancer biopsies

status: Public on Aug 25 2021

first: Anirban

last: Maitra

address: Smilow 910, 522 1st Ave

department: Perlmutter Cancer Center

institute: New York University Grossman School of Medicine

summary: Through single-cell RNA sequencing (scRNA-seq), we identify substantial  transcriptomic evolution of PDOs propagated from the parental tumor, which may alter  predicted drug sensitivity. In addition, we performed an integrative analyses of PDAC biopsies and provide an in-depth characterization of the heterogeneity within the tumor  microenvironment, including cancer-associated fibroblast (CAF) subclasses, and predicts for a multitude of ligand-receptor interactions, revealing potential targets for  immunotherapy approaches.

overall_design: Single cell RNA sequencing analysis of 9 biopsies, 8 PBMCs.

first: Jaewon

last: Lee

first: Vincent

last: Bernard

first: Alexnader

last: Semaan

first: Maria

last: Monberg

first: Jonathan

last: Huang

first: Bret

last: Stephens

first: Daniel

last: Lin

first: Kimal

middle: I

last: Rajapakshe

first: Brian

last: Weston

first: Manoop

middle: S

last: Bhutani

first: Cara

last: Haymaker

first: Chantale

last: Bernatchez

first: Cullen

last: Taniguchi

first: Paola

last: Guerrero


title: Transcriptional regulation of cell fate decisions in early B cell activation.

status: Public on Aug 30 2021

city: Vienna

last: Fischer

address: Campus-Vienna-Biocenter 1

institute: Research Institute of Molecular Pathology

summary: The cell fate decisions between plasmablasts (PBs), germinal center B cells (GCBCs) and non-GC-derived early memory B cells (eMBCs) during early B cell activation determine the outcome of the immune responses to pathogens and vaccines. To characterize these poorly understood lineage choices, we dissected the early B cell responses to T-dependent antigen in mice by single-cell RNA-sequencing. Early after immunization, a homogenous population of activated precursors (APs) gave rise to a transient wave of PBs, followed a day later by the emergence of the first GCBCs, with the transcriptomes of both rapidly diverging from that of APs. The majority of APs started to withdraw from the cell cycle very early on and gave rise to eMBCs, a developmental transition that involved limited transcriptional changes. These events were controlled by antigen availability that rapidly declined soon after immunization, and provision of antigen excess interfered with the cell cycle exit of APs and induced a new wave of PBs. Fate mapping experiments revealed a prominent contribution of eMBCs to the overall MBC pool. Generation of the earliest GCBCs was tightly controlled by the transcriptional repressor Bhlhe40 in the absence of which this population was increased in numbers. Bhlhe40 also restrained in a cell intrinsic fashion the response of T follicular helper (TFH) cells, a specialized T helper cell subset required to mount the GC response. In B cells, Bhlhe40 executed its function in the first days after immunization by selectively restricting the generation of the earliest GCBCs but not of eMBCs or PBs. Conditional Bhlhe40 inactivation confirmed cell-autonomous functions of Bhlhe40 in both GCBCs and TFH cells, while the GC phenotype was further enhanced upon loss of Bhlhe40 in both cell types. This negative regulation of the GC reaction by Bhlhe40 was of crucial importance, as Bhlhe40-deficient mice with progressing age succumbed to a B cell lymphoma characterized by the accumulation of monoclonal GCBCs-like cells and polyclonal TFH cells in various tissues.

overall_design: 23 bulk RNA-seq samples, 3 single cell RNA-seq, 10 bulk RNA-seq, 2 ChIP-seq

first: René

last: Rauschmeier

first: Annika

last: Reinhardt

first: Charlotte

last: Gustafsson

first: Vassilis

last: Glaros

first: Artem

middle: V

last: Artemov

first: Reshma

last: Taneja

first: Igor

last: Adameyko

first: Robert

last: Månsson

first: Meinrad

last: Busslinger

first: Taras

last: Kreslavsky

first: Sebastian

last: Ols

first: Aikaterini

last: Emmanouilidi

first: Yuanyuan

last: You

first: Claudio

last: Mirabello

first: Åsa K.

last: Björklund

first: Laurent

last: Perez

first: Neil P.

last: King

first: Davide

last: Angeletti

first: Karin

last: Loré


title: Transcriptomic analyses of lung adenocarcinoma cells and genome-wide maps of chromatin state in lung adenocarcinoma cells

status: Public on Aug 21 2021

city: Daegu

first: Hyo-Gyoung

last: Kang

address: Hoguk-ro, Buk-gu

department: Cell and Matrix Research Institute, School of Medicine

country: South Korea

institute: Kyungpook National University


title: Genome-wide maps of chromatin state in lung adenocarcinoma cells

summary: We investigated the association between genetic variants in the histone modification regions and the prognosis of lung adenocarcinoma after curative surgery. Potentially functional SNPs were selected using integrated analysis of ChIP-seq and RNA-seq. The SNPs were analyzed in a discovery set (n=166) and a validation set (n=238). The associations of the SNPs with overall survival (OS) and disease-free survival (DFS) were analyzed. This study showed that genetic variants in the histone modification regions could predict the prognosis of lung adenocarcinoma after surgery.

overall_design: Examination of 2 different histone modifications in lung adenocarcinoma cells.

first: Hyo

middle: G

first: Jae

middle: Y

last: Park

first: Shin

last: Lee


title: Systematic calibration of epitranscriptomic atlas using a synthetic modification-free RNA library

status: Public on Aug 18 2021

city: Guangzhou

first: Zhang

last: Zhang

state: Guangdong

address: Xingangxi Roda 135#

institute: Sun Yat-sen University

type: Methylation profiling by high throughput sequencing

summary: Recent years has witnessed rapid progress of the field epitranscriptomics. Functional interpretation of epitranscriptome relies on mapping technologies which determine the localization and stoichiometry of various RNA modifications. However, contradictory results are derived from different studies, questioning the biological impacts of certain RNA modifications. Here, we develop an approach for the generation of synthetic RNA library resembling the endogenous transcriptome but lacking modifications. Incorporating this modification-free RNA library as a negative control into established techniques, we obtain precise and quantitative maps of m6A and m5C after removing the pervasive false positives resulted from other elements such as specific sequence context and RNA secondary structure.

overall_design: In this study, we developed a new method to create a synthetic RNA library resembling the sequence and expression of endogenous transcriptome except modifications. Then we applied the modification-free RNA library in three representative scenarios described above and obtained high confidence, quantitative epitranscriptome maps of m6A and m5C after eliminating the false positive sites.

first: Guanzheng

last: Luo


title: Transcriptomic analyses of lung adenocarcinoma cells

overall_design: Transcriptome analysis in lung adenocarcinoma cells.


title: Secretory MPP3 reinforce myeloid differentiation trajectory and  amplify myeloid cell production

city: Yuseong-gu

first: Hyojung

last: Paik

state: Daejeon

address: 245 Daehak-ro

institute: KISTI

summary: Hematopoietic stem cells (HSC) and downstream lineage-biased multipotent progenitors (MPP) tailor blood production and control myelopoiesis on demand. However, a precise resolution of myeloid differentiation trajectories and cellular heterogeneity in various MPP populations and understanding of their role in controlling myeloid cell production remains largely lacking. Here, we analyze that myeloid-biased MPP3 are functionally and molecularly heterogenous, with a distinct subset of myeloid-primed secretory cells with high endoplasmic reticulum (ER) volume.

overall_design: Examination of 15 Bulk RNA seqs from hematopoietic stem and progenitor populations from mouse bone marrow, and 8 droplet-based single cell RNA seqs from MPP3 cells from mouse bone marrow

first: Yoon-A

first: Si

first: Jonathan

last: Chen

first: Matt

last: Warr

first: Rong

last: Fan

first: Emmanuelle

last: Passegué


title: In-vitro B cell stimulation RNA-seq

status: Public on Aug 17 2021

city: Philadelphia

first: Diego

last: Espinoza

state: PA

address: 3400 Civic Center Boulevard

institute: University of Pennsylvania

summary: RNA-seq profiles of sorted B cells undergoing stimulation of BCR, CD40, both, or none.

overall_design: 3 donors; 4 conditions each; 12 samples total

first: Rui

last: Li


title: RNASeq of Candida-stimulated CD3+ T-cells from individuals with CARD9 mutations and normal controls

status: Public on Aug 12 2021

city: Bethesda

first: Julie

middle: E

last: Niemela

state: MD

address: 10 Center Dr

department: Dept. of Lab Medicine

institute: NIH Clinical Center

summary: CD3+ T cells were enriched using the EasySep human T cell isolation kit (Stem cell technology). T cells from normal controls or patients with CARD9 mutations were co-cultured with monocytes of one normal control in the presence of heat killed candida. After 3 days, T cell were enriched again to remove the monocytes and total RNA was extracted from the T cells for the RNA-sequencing (RNASeq) evaluation. Targeted RNA sequencing library preparation was carried out using the Ion AmpliSeq Transcriptome Human Gene Expression Kit (Life Technologies), which profiles more than 20,000 human genes; each amplicon (~150 bp) represents a unique targeted gene (one transcript per gene). For library preparation, each sample was run in duplicate, and a cDNA library was generated from a minimum of 10 ng of total RNA. The cDNA was barcoded and amplified with Ion AmpliSeq technology, and the amplified cDNA Libraries were evaluated for quality and quantified with Agilent Bioanalyzer high-sensitivity chip. Libraries were then diluted to 100 pM and pooled equally, with 4 individual samples per pool. Pooled libraries were amplified and enriched with the Ion Chef System (Life Technologies). Templated libraries were then sequenced on an Ion Torrent Proton sequencing system (Life Technologies) with Ion PI HiQ kit and chip version 3. We performed gene-level differential expression analysis of targeted RNASeq data using R (v.3.5.3) and the Bioconductor packages DESeq2 (v.1.22.2).

overall_design: RNA sequencing was performed using the Ion AmpliSeq Transcriptome Human Gene Expression Kit  and Ion Torrent Proton sequencing system (Life Technologies). We analyzed RNA samples extracted from human Candida-stimulated CD3+ T-cells from normal controls and patients with CARD9 mutations.

first: Sergio

last: Rosenzweig

first: Hyesun

last: Kuehn

first: Jennifer

last: Stoddard

first: Shubham

last: Goel


title: GABAergic neuronal IL-4R mediates T cell effect on memory [WT + SCID]

last: Kipnis

address: 4515 McKinley Ave

institute: Washington University in Saint Louis

summary: The immune and nervous systems are highly interactive, however, little is known about how immune cells and their derived molecules impact brain function. Here we show that T cells play a critical role in retrieval of contextual fear conditioning (CFC) memory. Single-nuclei sequencing (snRNAseq) of engram neurons and non-engram neurons from the dentate gyrus one day after CFC learning revealed new evidence about differential expression of immune-related and synaptic plasticity-related genes, in SCID vs. wild-type mice. These findings present a comprehensive picture on how T cells and their derived cytokines could regulate memory retrieval and provide new insights into the nature of neuroimmune interactions at the transcriptional level in neurons.  Overall, restoration of T cells or IL-4R signaling might lead to selective rescue of memory retrieval, and proffer a valuable strategy for treating memory loss.

overall_design: This experiment aims to compare the transcriptional profile of nuclei from 2 genotypes (wild-type/prkdc-SCID)  and 2 activation states (engram/nonengram). All nuclei were collected from dentate gyrus of mice after fear conditioning. All mice were housed under standard conditions and injected with labelled viruses to detect the activation state.

first: Jasmin

last: Herz

first: Zhongxiao

last: Fu

first: Taitea

last: Dykstra


title: Transcriptome analysis of immature and mature male and female gametocytes of the malaria parasite Plasmodium falciparum

status: Public on Jul 31 2021

city: Erlangen

first: Michaela

last: Petter

address: Wassertrumstr. 3-5

department: Mirkobiologisches Institut

institute: Univeritätsklinikum Erlangen

summary: In the human host, Plasmodium falciparum parasites propagate asexually in erythrocytes causing the symptoms of malaria. However, in every asexual cycle, a small proportion of the parasites commits to sexual differentiation, which is critical for transmission. During a period of about 10 days, the sexually committed parasites either develop into male or female gametocytes. In this study, gametocytes carrying an endogenously GFP tagged version of the female specifically expressed ABCG2 gene were FACS sorted to separate male (GFP low) and female (GFP high) gametocyte populations at different days of development. RNA was subsequently extracted from these populations as well as from asexual parasites as controls and subjected to directional RNAseq. These data reveal the transcriptional differences between asexual parasite stages and developing gametocytes, but also between male and female gametocytes as well as for each sex during the course of differentiation. In particular, male gametocytes undergo distinct transcriptional transitions when comparing the transcriptome between day 4, 6 and 10 of gametocytogenesis. In contrast, female gametocyte transcriptomes are more static during the course of gametocytogenesis and only show minimal variation. This is the first study to longitudinally monitor gene expression in developing male and female gametocytes

overall_design: RNA-Seq of Plasmodium falciparum parasites in differnet developmental stages: ring stage parasites, schizont stage parasites, day 4, 6, and 10 female gametocytes, and day 4, 6, and 10 male gametocytes

first: Jingyi

last: Tang

first: Shamista

last: Selvarajah

first: Michael

middle: F

last: Duffy

first: Myriam

last: Jeninga


title: RNA-seq of Pluripotent stem cell derived cardiac cells

status: Public on Aug 11 2021

city: Madison

first: Martha

last: Floy

state: Wi

address: 1415 Engineering Hall

institute: University of Wisconsin-Madison

summary: Cardiac fibroblasts (CFBs) support heart function by secreting extracellular matrix and paracrine factors, respond to stress associated with injury and disease, and therefore are an increasingly important therapeutic target to treat heart disease. Here, we profile how developmental lineage of human pluripotent stem cell-derived CFBs, epicardial (EpiC-FB) and second heart field (SHF-FB) impacts their transcriptome. We compared this to human pluripotent stem cell derived cardiomyocytes, primary fetal cardiac fibroblasts, primary adult left ventricular fibroblasts, and primary adult dermal fibroblasts.

overall_design: We used three independent differentiation replicates in H9 hESCs for the hPSC-derived cell types and hPSC-cardiac fibroblasts were sampled at P1. Three technical replicates of primary fibroblasts were used between P3-5.

first: Sean

middle: P

last: Palecek


title: Identify METTL3, 14 and 16-bound transcripts via RIP-seq

status: Public on Oct 28 2021

city: Monrovia

first: Jianjun

last: Chen

institute: Beckman Research Institute of City of Hope

summary: To identify the directly bound transcripts of METTL3, METTL14, and METTL16, RNA immunoprecipitation sequencing (RIP-seq) was conducted HEK293T stablely expressing METTL3, METTL14, and METTL16, respectively. Briefly, HEK293T cells were infected with lentivirus, pmiRNA1-3 x Flag-METTL3, pmiRNA1-3 x Flag-METTL14, and pmiRNA1-3 x Flag-METTL14, to overexpress the three METTL family members with 3 x Flag fused in the N-terminal. Only the GFP-positive cells were used for study and expanded in DMEM medium.

overall_design: The GFP-positive HEK293T cells with overexpression of METTL3, METTL14, and METTL16, were expanded in 150mm dish and collected at 70-80% confluency. The were cross-linked by 0.75 % formaldehyde (F8775, Sigma-Aldrich) with gentle rotation at room temperature for 10 min and the reaction was quenched by 125 nM glycine (final concentration) with shaking at room temperature for 5 min. Then cells were collected, lysed, and subjected to RIP with Flag antibody. Two biological replicates were included to confirm the data quality.

last: Su

first: Lei

last: Dong


title: Single-cell-resolved differentiation of human induced pluripotent stem cells into pancreatic duct-like organoids on a microwell chip

status: Public on Jul 08 2021

city: Neuherberg

first: Sandra

last: Wiedenmann

state: Bavaria

address: Ingolstädter Landstraße 1

department: Helmholtz Pioneer Campus

institute: Helmholtz Zentrum München

summary: Creating  in  vitro  models  of  diseases  of  the  pancreatic  ductal  compartment  requires  a  comprehensive  understanding  of  the  developmental trajectories of pancreas-specific cell types. Here, we report the single-cell characterization of the differentiation of pancreatic duct-like organoids (PDLOs) from human induced pluripotent stem cells (hiPSCs) on a microwell chip that facili-tates the uniform aggregation and chemical induction of hiPSC-derived pancreatic progenitors. Using time-resolved single-cell transcriptional profiling and immunofluorescence imaging of the forming PDLOs, we identified differentiation routes from pan-creas  progenitors  through  ductal  intermediates  to  two  types  of  mature  duct-like  cell  and  a  few  non-ductal  cell  types.  PDLO  subpopulations  expressed  either  mucins  or  the  cystic  fibrosis  transmembrane  conductance  regulator,  and  resembled  human  adult duct cells. We also used the chip to uncover ductal markers relevant to pancreatic carcinogenesis, and to establish PDLO co-cultures with stellate cells, which allowed for the study of epithelial–mesenchymal signalling. The PDLO microsystem could be used to establish patient-specific pancreatic duct models.

overall_design: single cell RNA sequencing of pancreatic duct-like organoids differentation kinetics

first: Markus

last: Breunig

first: Jessica

last: Merkle

first: Christine

last: von Törne

first: Tihomir

last: Georgiev

first: Michel

last: Moussus

first: Lucas

last: Schulte

first: Thomas

last: Seufferlein

last: Sterr

first: Heiko

last: Lickert

first: Stefanie

last: Weissinger

first: Peter

last: Möller

first: Stephanie

last: Hauck

first: Meike

last: Hohwieler

first: Alexander

last: Kleger

first: Matthias

last: Meier


title: Expression data from mouse endothelial cell in human ovarian cancer

status: Public on Jul 25 2021

city: Daejeon

first: Kyunghee

last: Noh

address: 125 gahakro

institute: KRIBB

summary:
To examine potential mechanisms underlying resistance to anti-VEGF antibody (AVA) therapy, we used mouse models to identify tumors that demonstrated growth subsequent to a period of initial response. Specifically, we established orthotopic mouse models of ovarian cancer designed to develop adaptive resistance after treatment with an AVA.
To examine potential mechanisms underlying resistance to anti-VEGF antibody (AVA) therapy, we used mouse models to identify tumors that demonstrated growth subsequent to a period of initial response. Specifically, we established orthotopic mouse models of ovarian cancer designed to develop adaptive resistance after treatment with an AVA.

overall_design: We extracted total RNA from B20-sensitive and B20-resistant tumor endothelial cells with use of an RNeasy mini kit (Qiagen). A NanoDrop spectrophotometer (Thermo Fisher Scientific) was used for assessment of both RNA quality and quantity. Total RNA (700 ng) was labeled and hybridized to Bead Chips according to the manufacturer’s protocols

first: Anil

last: Sood


title: Peripheral blood gene expression according to IFIH1 rs1990760 variants in critically-ill COVID-19 patients

status: Public on Jul 03 2021

city: Oviedo

first: Guillermo

last: Albaiceta

address: Avenida de Roma s/n

institute: Instituto de Investigación Sanitaria del Principado de Asturias

summary:
Background: Outcomes in patients with severe SARS-CoV-2 infection (COVID-19) are conditioned by viral control and regulation of inflammation. Variants in IFIH1, a gene coding the cytoplasmatic RNA sensor MDA5, regulate the response to viral infections.
Methods: Patients admitted to an intensive care unit (ICU) with documented COVID-19 were prospectively included and IFIH1 rs1990760 genotypes determined. Peripheral blood gene expression, cell populations and immune mediators were measured during the first day after ICU admission before steroid therapy. Peripheral blood mononuclear cells from healthy volunteers were exposed ex-vivo to an MDA5 agonist and dexamethasone, and changes in gene expression assessed. ICU discharge and hospital death were modelled using rs1990760 variants and dexamethasone therapy as factors.
Findings: 237 patients were studied. Patients with the IFIH1 rs1990760 TT variant showed a decrease in expression of inflammation-related pathways, an anti-inflammatory cell profile and a decrease in pro-inflammatory mediators. Cells with TT variant exposed to an MDA5 agonist ex-vivo showed an increase in FOXO3 and IL6 when dexamethasone was added. All patients with the TT variant not treated with steroids (n=14) survived their ICU stay (HR 2.49 95% confidence interval 1.29 – 4.79). Dexamethasone therapy in this subgroup (N=50) delayed ICU discharge and increased hospital mortality (HR 2.19, 95% confidence interval 1.01 – 4.87) and serum IL-6 concentrations.
Interpretation: COVID-19 ICU patients with the IFIH1 rs1990760 TT variant show an ameliorated inflammatory response that results in better outcomes than CC/CT variants. Dexamethasone can reverse this anti-inflammatory phenotype, worsening the outcome.
Funding: Instituto de Salud Carlos III.

overall_design: Prospective observational cohort study. RNA was extracted from peripheral blood on day 1 after ICU admission. Forty-two patients were studied.

first: Inés

last: López-Alonso

first: Cecilia

last: López-Martínez

first: Laura

last: Amado-Rodríguez


title: Phloem tissue specific genome-wide open chromatin state in Arabidopsis thaliana.

city: Heidelberg

last: Greb

address: Im Neuenheimer Feld 360

department: Centre for Organismal Studies

institute: Heidelberg University

summary: Transformation of undifferentiated stem cells into cells with special functions is central for organismal development. The phloem tissue mediates long-distance transport of energy metabolites along plant bodies and is characterized by an exceptional degree of cellular specialization. How the phloem-specific developmental program is implemented is, however, unknown. Here we reveal that the ubiquitously expressed PHD-finger protein OBERON3 (OBE3) and the phloem-specific SUPPRESSOR OF MAX2 1-LIKE 5 (SMXL5) protein form a central module for establishing phloem identity in Arabidopsis thaliana (Arabidopsis). By phloem-specific ATAC-seq analyses, we show that OBE3 and SMXL5 proteins establish a phloem-specific chromatin profile.

overall_design: Open chromatin state was examined by ATAC-seq using sorted phloem nuclei (SMXL5+) or non-phloem nuclei (SMXL5-) based on SMXL5pro:H4GFP signal in wildtype, smxl5 mutant, smxl4smxl5 double mutant and smxl5obe3 double mutant of Arabidopsis thaliana seedlings. No replicates included.

last: Luzzietti

first: Dongbo

last: Shi


title: Effects of steroids on peripheral blood gene expression in COVID-19 patients according to rs1990760 variants

summary: Background: Outcomes in patients with severe SARS-CoV-2 infection (COVID-19) are conditioned by viral control and regulation of inflammation. Variants in IFIH1, a gene coding the cytoplasmatic RNA sensor MDA5, regulate the response to viral infections.  Methods: Patients admitted to an intensive care unit (ICU) with documented COVID-19 were prospectively included and IFIH1 rs1990760 genotypes determined. Peripheral blood gene expression, cell populations and immune mediators were measured during the first day after ICU admission before steroid therapy. Peripheral blood mononuclear cells from healthy volunteers were exposed ex-vivo to an MDA5 agonist and dexamethasone, and changes in gene expression assessed. ICU discharge and hospital death were modelled using rs1990760 variants and dexamethasone therapy as factors.  Findings: 237 patients were studied. Patients with the IFIH1 rs1990760 TT variant showed a decrease in expression of inflammation-related pathways, an anti-inflammatory cell profile and a decrease in pro-inflammatory mediators. Cells with TT variant exposed to an MDA5 agonist ex-vivo showed an increase in FOXO3 and IL6 when dexamethasone was added. All patients with the TT variant not treated with steroids (n=14) survived their ICU stay (HR 2.49 95% confidence interval 1.29 – 4.79). Dexamethasone therapy in this subgroup (N=50) delayed ICU discharge and increased hospital mortality (HR 2.19, 95% confidence interval 1.01 – 4.87) and serum IL-6 concentrations.  Interpretation: COVID-19 ICU patients with the IFIH1 rs1990760 TT variant show an ameliorated inflammatory response that results in better outcomes than CC/CT variants. Dexamethasone can reverse this anti-inflammatory phenotype, worsening the outcome.  Funding: Instituto de Salud Carlos III.

overall_design: Prospective observational cohort study. RNA was extracted from peripheral blood on day 4 after ICU admission.


title: Long-term survivors of murine sepsis exhibit enhanced LPS-induced lung injury and proinflammatory immune reprogramming

status: Public on Jun 23 2021

city: Ann Arbor

first: Scott

middle: John

last: Denstaedt

state: MI

address: 109 Zina Pitcher Pl

department: Internal Medicine

institute: University of Michigan

summary: Sepsis is the most common cause of hospitalization worldwide. Millions of people survive sepsis each year and are at risk for rehospitalization and death. Pulmonary complications such as respiratory failure due to pneumonia and exacerbation of chronic respiratory disease are among the most common reasons for rehospitalization in sepsis survivors. In order to prevent additional morbidity and death in patients surviving sepsis, we must establish biomarkers to identify patients at risk for pulmonary complications and develop treatments. Late complications in sepsis survivors, particularly nosocomial infections, are proposed to occur through persistent immune reprogramming after sepsis known as immunoparalysis. However, pro-inflammatory immune reprogramming in the form of primed or enhanced responses to secondary stimuli has also been described and could directly contribute to tissue injury and death. Primed immune responses and their contribution to long-term sepsis complications remains understudied. We hypothesize that primed immune responses to inflammatory stimuli in the lung after sepsis are associated with pulmonary complications in survivors of sepsis. To this end, we developed a model of antibiotic treated sepsis induced by cecal ligation and puncture followed three weeks later by secondary challenge with intranasal lipopolysaccharide to induce inflammatory lung injury. We find that mice surviving sepsis have enhanced lung injury responses in the setting of an exaggerated proinflammatory immune response, including primed Ly6Chi monocyte Tnf expression. Using RNA sequencing, we identified derangements in lung gene expression after CLP prior to LPS administration which may mediate enhanced lung injury in this model. One potential mediator, S100A8/A9, was also found to be elevated in the circulation of human sepsis survivors for up to 180 days after sepsis. These findings validate our model and identify S100A8/A9 as one of many potential biomarkers and therapeutic targets for patients at risk for long-term pulmonary complications after sepsis.  The role of S100A8/A9, monocyte priming, and other factors predisposing to enhanced lung injury responses and pulmonary complications after sepsis warrant further investigation in humans and mice.

overall_design: 12 post-mortem perfused lung tissue samples from male mice (C57BL/6) with and without sepsis, 21 days post CLP infection. Cases are CLP sepsis survivors and controls are unopertated healthy controls.

middle: J

first: Angela

middle: C

last: Bustamante

first: Michael

middle: W

last: Newsteadt

first: Bethany

middle: B

last: Moore

first: Theodore

last: Standiford

first: Benjamin

last: Singer

first: Rachel

middle: L

last: Zemens


title: The long noncoding RNA ELDR serves as an epigenetic driver in aging and injury-induced chondrocyte senescence [chondrocytes part2]

status: Public on Jun 20 2021

city: Nanjing

first: Mingliang

last: Ji

state: Jiangsu

address: Dingjiaqiao Road 87

institute: Zhongda Hospital,school of medicine, southeast university

summary: Using  large-scale patient data sets and genetically engineered (inducible conditional knockout and knockin) mouse models, we show that a novel transcript of long noncoding RNA ELDR is essential for the development of chondrocyte senescence.

overall_design: 3 chondrocytes from controls and 3 chondrocytes from Eldr cKI mice

first: Ming

middle: l

first: Jun

last: Lu


title: Self-reactivity shapes functional diversity of naive CD8+ T cells by co-opting tonic type I interferon

status: Public on Jun 19 2021

city: Hwasun-eup, Hwasun-gun

first: Jae-Ho

last: Cho

state: Jeollanam-do

address: 264, Seoyang-ro

institute: CHONNAM NATIONAL UNIVERSITY

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

overall_design: Refer to individual Series


title: Systemic Transcriptional Phenotypes in Mecp2 Null Mice

status: Public on Sep 01 2021

city: Middlebury College

first: Amanda

last: Crocker

state: Vermont

address: 276 Bicentennial Way

department: Neuroscience

institute: Middlebury College

summary: We report the RNAseq signatures of cortex, hippocampus, striatum, liver, muscle and kidney from wild type and Mecp2 null males mice (B6.129P2(C)-Mecp2 tm1.1Bird/J, Stock No: 003890) at ~44-46 days of postnatal age

overall_design: We compare diffrential gene expression between males wild type and null.

first: Victor

last: Faundez


title: Genomic alterations in Reinke's edema

status: Public on Jun 09 2021

city: Brasília

first: Rolando

middle: Andre Rios

last: Villacis

state: DF

address: Asa Norte

department: Genetics and Morphology

country: Brazil

institute: University of Brasília - UnB

type: Genome variation profiling by genome tiling array

summary: Reinke's edema (RE) is a laryngeal lesion related to excessive tobacco smoking, voice overuse, and laryngopharyngeal reflux. Although the risk of malignancy is low, RE has been classified among premalignant lesions. Array-based comparative genomic hybridization (aCGH) was used to investigate DNA copy number alterations (CNA) and as a tool to identify markers of risk stratification.

overall_design: Genomic profiling of eight surgical specimens of RE was evaluated by aCGH (Agilent 4x180K platform). CNAs were compared with the Database of Genomic Variants (DGV) and genes mapped on altered regions with annotated functions.

first: Luis

last: Móz

middle: A


title: The long noncoding RNA ELDR serves as an epigenetic driver in aging and injury-induced chondrocyte senescence [chondrocytes part1]

overall_design: 3 chondrocytes from controls and 3 chondrocytes from Eldr cKO mice


title: Dynamic chromatin accessibility tuning of long noncoding RNA ELDR accelerates chondrocyte senescence and osteoarthritis


title: tRF-seq landscapes the variation of tRNA derived fragments after mouse liver injury

status: Public on Jun 05 2021

city: Beijing

first: Sunyang

last: YING

state: Beijing

address: 1-5 Beichen West Road, Chaoyang District, Beijing

department: State key laboratory of stem cell and reproductive biology

institute: Chinese Academy of Sciences

summary: In order to intensively elucidate the metabolic characteristics of tRFs after liver injury, we performed tRF-seq by high throughput sequencing. We found that after liver injury, large abundance of tRFs were produced. Especially, the tRF-1s largely vary during the liver injury process and may play important role in the liver disease.

overall_design: WT and NS-KO mice were once or repeated injected CCl4, recorded as Day 3 or Day 27. The control mice of WT or NS-KO were injected corn oil, recorded as Day 0. The mice livers were collected and used for tRF-seq, in duplicate, using Illumina NextSeq 500 system.

first: S

last: Ying

first: Q

last: Zhou


title: ZHX2 Promotes HIF1a Oncogenic Signaling in Triple-Negative Breast Cancer

status: Public on May 25 2021

city: Boston

first: Jeremy

last: Simon

state: MA

address: 450 Brookline Ave

department: Department of Data Science

institute: Dana-Farber Cancer Institute

summary: Triple-negative breast cancer (TNBC) is an aggressive and highly lethal disease. Due to its heterogeneity and lack of hormone receptors or HER2 expression, it is critical to identify novel therapeutic targets in TNBC. Analysis of copy number and gene expression in tumors from The Cancer Genome Atlas suggested that ZHX2 was amplified and overexpressed in breast cancer patients. Correspondingly, we found that ZHX2 was highly expressed in TNBC cell lines and TNBC patient tissues. Functionally, depletion of ZHX2 inhibited TNBC cell growth and invasion in vitro, orthotopic tumor growth and spontaneous lung metastasis in vivo. Mechanistically, ZHX2 bound with Hypoxia inducible factor (HIF) family members and positively regulated HIF1 activity in TNBC by using loss-of-function or gain-of-function studies. Our integrated ChIP-Seq and gene expression profiling demonstrated that ZHX2 co-occupied with HIF1 on transcriptionally active promoters marked by H3K4me3 and H3K27Ac, therefore promoting gene expression. Furthermore, structural simulation and functional studies revealed that multiple residues (R491, R581 and R674) are important in regulating the phenotype of ZHX2 on TNBC tumorigenic potential, which correspond with their roles on controlling HIF1 activity in TNBC cells. These studies establish that ZHX2 activates oncogenic HIF1 signaling, therefore serving as a potential therapeutic target for TNBC.

overall_design: ChIP-seq of ZHX2 and input control, and RNA-seq of ZHX2-knockdown vs control shRNAs in MDA-MB-231 breast cancer cells

middle: M

first: Qing

last: Zhang


title: Self-reactivity shapes functional diversity of naive CD8+ T cells by co-opting tonic type I interferon [RNA-Seq_2]

summary: The strength of the T cell receptor interaction with self-ligands affects antigen-specific immune responses. However, the precise role, functional relationship, and underlying mechanisms have not been elucidated completely. Here, we demonstrate that naive CD8+ T cells with relatively high self-reactivity are phenotypically heterogeneous owing to varied responses to type I interferon (T1IFN), resulting in three distinct subsets, CD5loLy6C–, CD5hiLy6C–, and CD5hiLy6C+ cells. CD5hiLy6C+ cells differ from CD5loLy6C– and CD5hiLy6C– cells in their gene expression profiles and functional properties. Moreover, CD5hiLy6C+ cells demonstrate more extensive antigen-specific expansion upon viral infection, with enhanced differentiation into terminal effector cells and reduced memory cell generation. Such unique features of CD5hiLy6C+ cells are imprinted in a steady-state and observed even for those of monoclonal CD8+ T cell populations in a T1IFN dependent fashion. These findings demonstrate that self-reactivity shapes the functional diversity of naive CD8+ T cells by co-opting tonic T1IFN signaling.

overall_design: Gene profiles of 3 naive (CD44lo) P14 CD8+ T cell subsets (CD5lo, CD5hi Ly6C-, and CD5hi Ly6C+) after LCMV infection (DPI 7)

first: Young-Jun

last: Ju

first: Sung-Woo

last: Lee


title: RNA-seq in murine HSCs and CHIP-seq for H3K27me3 in K562 cells after CUX1 knockdown

status: Public on May 03 2021

city: Chicago

first: Megan

last: McNerney

state: IL

address: 900 E 57th St

department: Pathology

institute: The University of Chicago

summary: Therapy-related myeloid neoplasms (t-MN) are a high-risk, late effect in cancer survivors with poorly understood pathogenesis. It has been postulated that, in some cases, hematopoietic stem and progenitor cells (HSPCs) harboring mutations are selected for by cytotoxic exposures and transform. Here, we elucidate this model in the context of deficiency of CUX1, a transcription factor encoded on chromosome 7q. We report that CUX1 has a critical, early role in the DNA repair process in HSPCs. Mechanistically, CUX1 recruits the histone methyltransferase EHMT2 to DNA breaks to promote H3K27 methylation, phospho-ATM retention, subsequent γH2AX foci formation and propagation and, ultimately, 53BP1 recruitment. Despite significant unrepaired DNA damage sustained in CUX1-deficient murine HSPCs after cytotoxic exposures, they continue to proliferate and expand, mimicking clonal hematopoiesis in patients post-chemotherapy. As a consequence, preexisting CUX1 deficiency predisposes mice to highly penetrant and rapidly fatal therapy-related erythroleukemias. These findings establish the importance of epigenetic regulation of DNA repair and position CUX1 as a gatekeeper in myeloid transformation.

overall_design: RNA-seq in control (Renilla) and Cux1-low murine hematopoietic stem cells (three replicates) and ChIP-seq of H3K27me3 in control (gHPRT) and CUX1-knockout K562 cells (two replicates)

first: Molly

middle: K

last: Imgruet

first: Jeffrey

last: Kurkewich


title: The long noncoding RNA ELDR serves as an epigenetic driver in aging and injury-induced chondrocyte senescence [lncRNA]

first: Mingliang

last: Ji

type:
Expression profiling by array
Non-coding RNA profiling by array

summary:
To investigate lncRNA profile in OA patients and analyze the critical role of lncRNA in  chondrocyte senescence
To investigate lncRNA profile in OA patients and analyze the critical role of lncRNA in  chondrocyte senescence

overall_design: 3 OA patients and 3 controls

first: Ming


title: Matrix-anchored Nanoparticles for MMP13 RNA Interference Block Post-Traumatic Osteoarthritis

status: Public on Apr 17 2021

city: Nashville

first: Sean

last: Bedingfield

state: TN

address: 2301 Vanderbilt Place PMB 351826

department: Biomedical Engineering

institute: Vanderbilt University

summary: Targeting MMP13 directly by RNA interference (RNAi) silenced MMP13 in a murine post-traumatic osteoarthritis (PTOA) and was found to have broad effects on overall joint health, including both the articular cartilage and surrounding hard and soft tissues. To further characterize the global impacts of targeted MMP13 inhibition, the mechanical loading mouse model was applied with samples were harvested at 4 weeks to capture an intermediate stage of disease. The nanoString nCounter Inflammation panel was used to broadly quantify gene expression of the knee joint samples (articular cartilage, meniscal, and synovial tissue).   Abstract: Osteoarthritis (OA) is a debilitating and prevalent chronic disease, but there are no approved disease modifying OA drugs (DMOADs), only pharmaceuticals for pain management. OA progression, particularly for post-traumatic osteoarthritis (PTOA), is associated with inflammation and enzymatic degradation of the extracellular matrix. In particular, matrix metalloproteinase 13 (MMP13) breaks down collagen type 2 (CII), a key structural component of cartilage extracellular matrix, and consequently, matrix degradation fragments perpetuate inflammation and a degenerative cycle that leads to progressive joint pathology. Here, we tested extracellular matrix-binding MMP13 RNA interference (RNAi) nanoparticles (NPs) as a DMOAD. The retention approach pursued here deviates from the convention of targeting specific cell types (e.g., through cell surface receptors) and instead leverages a monoclonal antibody (mAbCII) that targets extracellular CII that becomes uniquely accessible in OA-damaged cartilage. CII monoclonal antibody-functionalized nanoparticles carrying small interfering ribonucleic acid (siRNA) (mAbCII-siNPs) create an in situ NP depot for retention and potent activity within OA joints. The mAbCII-siNPs loaded with MMP13 siRNA (mAbCII-siNP/siMMP13) potently suppressed MMP13 expression (95% silencing) in TNFα-stimulated chondrocytes in vitro and had higher binding to trypsin-damaged porcine cartilage than control NPs. In an acute mechanical injury mouse model of PTOA, mAbCII-siNP/siMMP13 achieved 80% reduction in MMP13 expression (p = 0.00231), whereas control siNP/siMMP13 achieved only 55% silencing. In a more severe, longer-term PTOA model, weekly mAbCII-siNP/siMMP13 treatment provided significant protection of cartilage integrity (0.45+/-.3 vs 1.6+/-.5 on the OARSI scale; p=0.0013) and overall joint structure (1.3+/-.6 vs 2.8+/-.2 on the Degenerative Joint Disease scale; p=0.0418), including reduction of osteophyte formation (siMMP13 vs siNEG: 0.088+/-0.074 vs 0.47+/-0.107 mm femoral osteophyte outgrowth; p<0.0001). Multiplexed gene expression analysis of 254 inflammation-related genes showed that MMP13 inhibition suppressed clusters of genes associated with tissue restructuring, angiogenesis, innate immune response, and proteolysis. Finally, in benchmarking studies, intra-articular mAbCII-siNPs better reduced OA disease progression relative to either one time or weekly treatment with the clinical gold standard steroid methylprednisolone. Here, we introduce the concept of anchoring to unique local extracellular matrix signatures to sustain retention and increase delivery efficacy of biologics with intracellular activity and also validates the promise of MMP13 RNAi as a DMOAD in a clinically-relevant therapeutic context.

overall_design: Groups are either healthy mice or subjected to an acute PTOA model of noninvasive, repetitive joint loading by subjecting the knee joints of 8-week-old C57BL/6 mice to 500 cycles of compressive mechanical loading at 9N (adapted from Poulet B, et al. Arthritis and rheumatism. 2011. PubMed PMID: 20882669). This procedure was repeated five times per week over a period of four weeks. PTOA groups were either left untreated or treated MMP13-specific or negative control small interfering RNA (siRNA) loaded into collagen II-targeting antibodies conjugated to polymer nanopartiples (mAbCII-siNPs/siMMP13 or siNEG).

first: Craig

last: Duvall


title: Leukocyte dynamics after intracerebral hemorrhage in a living patient reveal rapid adaptations to tissue milieu

status: Public on Jun 02 2021

city: Cambridge

first: Dunaduan

last: Ma

state: Ma

address: 77 Massachusetts Ave

department: Koch Institute

institute: MIT

summary: Intracerebral hemorrhage (ICH) is a devastating form of stroke with a high mortality rate and few treatment options. Discovery of therapeutic interventions has been slow given the challenges associated with studying acute injury, particularly over time, in the human brain. Inflammation induced by exposure of brain tissue to blood appears to be a major part of brain tissue injury.  Here we longitudinally profiled blood and cerebral hematoma effluent from a patient enrolled in the Minimally Invasive Surgery with Thrombolysis in Intracerebral Haemorrhage Evacuation (MISTIEIII) trial, offering a rare window into the local and systemic immune responses to acute brain injury. Using single-cell RNA-sequencing, we characterized the local cellular response during ICH in the brain of a living patient at single-cell resolution for the first time. Our analysis revealed rapid shifts in the activation states of myeloid and T cells in the brain over time, suggesting that leukocyte responses are dynamically reshaped by the hematoma microenvironment. Interestingly, the patient had an asymptomatic re-bleed (second local exposure to blood) that our transcriptional data indicated occurred more than 30 hours prior to detection by CT scan. This case highlights the rapid immune dynamics in the brain after ICH and suggests that sensitive methods like scRNA-seq can inform our understanding of complex intracerebral events.

overall_design: We generated single cell RNA-sequencing data on cells isolated longitudinally from patient blood and hematoma drainage. We additionally performed scRNA-seq on patient followup blood 2.5 years after stroke, and on the blood of four age-matched control donors.

first: Brittany

last: Goods


title: Osteosarcoma tumors maintain intratumoral heterogeneity, even while adapting to environmental pressures that drive clonal selection

status: Public on Jul 09 2021

city: Columbus

first: Ryan

last: Roberts

state: OH

address: Research Building II, 5th Floor WA5108, 700 Children's Drive,

department: Center for Childhood Cancers and Blood Disorders

institute: Nationwide Children's Hospital

summary: Osteosarcoma cells were grown in cell culture, as subcutaneous flank tumors, orthotpic primary tumors or experimental metastases to study the role of intra-tumor transcriptional heterogeneity on adaptations in tumor cells that faciliate tissue colonization.

overall_design: single cell RNA-Seq profiles of human and murine osteosarcoma models (OS17, 143B, NCH-OS-2, NCH-OS-7, F420, K7M2) grown in cell culture, subcutaneous flank, primary tumor (bone) or  metasatic lesions (lung)

first: Sanjana

last: Rajan

first: Maren

last: Cam

first: Amy

last: Gross

first: Matthew

middle: V

last: Cannon

middle: D


title: MicroRNA-29 is an essential regulator of non-CG methylation during brain maturation [RNA-seq]

status: Public on Apr 07 2021

first: Jeremy

last: Simon

summary: While embryonic brain development and neurodegeneration have been examined extensively, the events that govern postnatal brain maturation are less understood. During this period, widespread changes in synaptic connectivity are evident1,2 and, perturbations of synaptic development or maturation can have severe neurobehavioral consequences as seen in autism spectrum disorders (ASD), epilepsy and schizophrenia3-6. Recent studies have led to the unexpected discovery that marked increases in non-canonical (non-CG) DNA methylation occur during postnatal brain maturation7-14. This de novo CH methylation (where H= A, C, T), which occurs 1-4 weeks postnatally in mice and 1-15 years in humans7, is important for experience-dependent fine tuning of gene expression15 . The critical period of CH methylation appears to be mediated by Dnmt3a, the DNA methyltransferase that catalyzes CH methylation8,9. During brain maturation, Dnmt3a levels are elevated and then sharply decline7,16, and a strict control of CH methylation seems important as mutations or variants of Dnmt3a are associated with ASD, epilepsy and intellectual disability17-19. Here we identify the microRNA miR-29 as an essential regulator of Dnmt3a and CH methylation during brain maturation. Mice deficient in miR-29 or mice in which miR-29 can no longer target Dnmt3a exhibit enhanced levels of Dnmt3a, global CH hypermethylation, and repression of neuronal genes specifically associated with synaptic activity and neurotransmission. Importantly, these mice appear normal at birth, but subsequently exhibit severe neurological deficits including susceptibility to seizures and premature lethality. Together, these results identify an essential function of miR-29 in defining the critical period of DNA methylation during brain maturation, the absence of which results in neurobehavioral deficits.

overall_design: Bulk RNA-seq of cortices from P40 miR29-Nestin (n=3) and miR29 WT (n=3) mice. Bulk RNA-seq of cortices from 4.5 month old Dnmt3a-mi5 (n=4) and Dnmt3a-WT (n=4) mice

first: Mohanish

last: Deshmukh


title: Single-cell RNAseq of childhood medulloblastoma

status: Public on Jun 04 2021

city: Aurora

first: Andrew

last: Donson

state: CO

department: Pediatrics

institute: CU Anschutz Medical Campus

summary: We explore cellular heterogeneity in 28 childhood medulloblastoma (MB) (1 WNT, 9 SHH, 7 GP3 and 11 GP4) using single-cell RNA sequencing (scRNA-seq), immunohistochemistry and deconvolution of bulk transcriptomic data. Neoplastic cells are broadly clustered according to subgroup, and within subgroups discrete sample clustering is associated with chromosomal copy number variance. Each subgroup contains subpopulations exhibiting mitotic , undifferentiated and neuronal differentiated transcript profiles , corroborating other recent medulloblastoma scRNA-seq studies and identifying new subpopulations. We identify a photoreceptor-differentiated subpopulation that is predominantly found in GP3 medulloblastoma, and in SHH, a subpopulation that constitutes differentiating nodules . Deconvolution of a large transcriptomic dataset shows that neoplastic subpopulations are associated with major and minor subgroup subdivisions, for example, photoreceptor subpopulation cells are more abundant in GP3-alpha. This scRNA-seq dataset also demonstrates medulloblastoma subgroup-specific differences in the tumor microenvironment and immune landscape, and reveals an SHH nodule -associated myeloid subpopulation. Additionally, we perform scRNA-seq on genetically engineered mouse (GEM) models of GP3 and SHH medulloblastoma. These models specifically matched the corresponding human subgroup-specific neoplastic subpopulations. We provide an interactive online resource that facilitates exploration of these MB single cell datasets. Collectively, our findings advance our understanding of the neoplastic and immune landscape of the main medulloblastoma subgroups in both humans and GEM models.

overall_design:
Characterization of childhood medulloblastoma using single-cell RNAseq and methylation profiling.
****Bam files produced by 10x Genomics cellranger were submitted to GEO. These files are available from the SRA database (SRP274357). If needed the bam files can be converted to the original paired-end FASTQs using bamtofastq from 10x Genomics****

first: Kent

last: Riemondy


title: Transcriptomic analyses of gastrulation-stage C57BL/6J and C57BL/6NHsd mouse embryos with differential susceptibility to alcohol

status: Public on Jun 01 2021

summary: Craniofacial and CNS malformations characteristic of Fetal Alcohol Syndrome are caused by alcohol exposure during gastrulation (embryonic day [E]7 in mice; 2nd – 3rd week of human pregnancy). Genetics are a known contributor to differences in alcohol sensitivity in humans, and between different animal model strains such as C57BL/6J and C57BL/6NHsd mice. To investigate how these two genetically similar mouse strains differ in alcohol sensitivity, we profiled gene expression at baseline (E7.0) and 6 or 12 hours after alcohol exposure (E7.25–E7.5) in gastrulation-stage embryos. Many of the baseline transcriptional differences across strains were associated with immune signaling, indicative of their molecular divergence. Alcohol exposure was associated with a more pronounced transcriptional effect in 6J than in 6N, matching the 6J’s increased sensitivity. Alcohol exposure upregulated pathways related to cell death, stress, and hypoxia, and downregulated proliferation and morphogenic pathways. Dysregulated genes were also associated with craniofacial and CNS defects, but only in the 6J embryos. These data demonstrate the genetic variation that can contribute to the effects of prenatal alcohol and identify impacted molecular pathways, including higher baseline expression of inflammatory signaling genes in 6J embryos that likely increases sensitivity to teratogens such as alcohol. Collectively, our data provide a valuable resource that not only enables the discovery of biomarkers pertinent to prenatal alcohol exposure, but also facilitates the transcriptional comparison of two genetically similar mouse strains at three finely resolved developmental timepoints. To this end, we developed a web-based interactive tool to aid in data exploration available at http://parnell-lab.med.unc.edu/Embryo-Transcriptomics/.

overall_design: RNA-seq at baseline (E7.0) and 6 or 12 hours after alcohol exposure (E7.25–E7.5) in gastrulation-stage embryos of C57BL6/J and C57BL6/N mice

first: Karen

last: Boschen

first: Scott

last: Parnell


title: Single Cell Genomechanics: Correlating Cancer Cell Mechanics and Gene Expression

city: Atlanta

first: Katherine

middle: Monnet

last: Young

state: GA

address: 315 Ferst Dr. NW

institute: Georgia Institute of Technology

summary: While cell mechanics and metastatic potential are related, the molecular factors that drive these behaviors remain unknown. Understanding how molecular signaling networks modulate cellular phenotype and mechanotype can help elucidate how metastasis occurs. Therefore, we developed a workflow to measure mechanical properties and gene expression on the single cell level. The process combines atomic force microscopy and optical microscopy to measure the mechanics and morphology of individual ovarian cancer cells, followed by multiplexed RT-qPCR gene expression analysis. Surprisingly, the genes that most strongly correlated with mechanical properties were not cytoskeletal, but rather were markers of epithelial-to-mesenchymal transition and cancer stemness. A dimensionality reduction analysis showed that cells of different metastatic potential were best identified through combining mechanical and gene expression data. Finally, a network analysis revealed master regulators that can predictably stiffen and soften cells while modulating cell migration. The single cell genomechanics methods demonstrate how molecular drivers can disable biophysical processes underpinning metastasis.

overall_design: Multiplexed RT-qPCR gene expression profiling. Single ovarian cancer cells from three cell lines (91 HEY A8 cells, 95 HEY cells, 87 OVCAR3 cells) were isolated through micropipette aspiration and their RNA content was collected for pre-amplification and multiplexed expression analysis with the Fluidigm 96.96 Dynamic Array IFC.


title: Age and dietary fiber effect on mouse brain cells

city: Urbana

first: Mario

middle: Pietro Emilio

last: Vailati Riboni

address: 1207 W Gregory Dr

department: Animal Science

institute: University of Illinois at Urbana-Champaign

summary: With increasing age microglia shift toward a pro-inflammatory phenotype and become hyperresponsive to inflammatory stimuli, disrupting brain homeostasis. Soluble fibers have been suggested as a dietary strategy to prevent or reverse microglia dysregulation, due to the bioactive nature of the short chain fatty acids (SCFA; e.g., butyrate) produced during its fermentation in the colon.

overall_design: One sample per combination of age (adult, aged) and fiber content (high, low), in a 2x2 factorial design.

last: Vailati-Riboni

first: Lauretta

last: Rund

first: Maria

middle: E

last: Caetano-Silva

first: Noah

last: Hutchinson

first: Selena

middle: S

last: Wang

first: Jeffrey

last: Woods

last: Steelman

first: Rodney

last: Johnson


title: Epigenetic prediction of response to anti-PD-1 treatment in non-small-cell lung cancer: a multicenter, retrospective analysis

city: Badalona

first: Manel

last: Esteller

state: Barcelona

address: Ctra. de Can Ruti, Cami de les Escoles s/n

department: Cancer and Leukemia Epigenetics and Biology Program (PEBCL)

institute: Josep Carreras Leukaemia Research Institute (IJC)

type: Methylation profiling by genome tiling array

summary: Anti-programmed death-1 (PD-1) treatment for advanced non-small-cell lung cancer (NSCLC) has improved the survival of patients. However, a significant percentage of patients do not respond. We examined the use of DNA methylation profiles to determine the efficacy of anti-PD-1 treatment in stage IV NSCLC patients.

overall_design: We first established an epigenomic profile based on a microarray DNA methylation signature (EPIMMUNE) in a discovery set of 34 NSCLC tumor samples treated with nivolumab or pembrolizumab. The EPIMMUNE signature was validated in an independent set of 47 NSCLC cases. A derived DNA methylation marker was validated by a single-methylation assay in a validation cohort of 61 NSCLC patients.

first: Michäel

last: Duruisseaux


title: Single cell sequencing identifies serine proteases as regulators of myofibroblast differentiation

status: Public on May 31 2021

city: Vienna

first: Vera

last: Vorstandlechner

address: Waehringer Guertel 18-20

institute: Medical University of Vienna

summary: Despite recent advances in understanding skin scarring, mechanisms triggering hypertrophic scar formation are still poorly understood. In the present study we performed single-cell sequencing of mature human hypertrophic scars and developing scars in mice. Compared to normal skin, we found significant differences in gene expression in most cell types present in scar tissue. Fibroblasts (FBs) showed the most prominent alterations in gene expression, displaying a distinct fibrotic signature. By comparing genes upregulated in murine FBs during scar development with genes highly expressed in mature human hypertrophic scars, we identified a group of serine proteases, tentatively involved in scar formation. Two of them, dipeptidyl-peptidase 4 (DPP4) and urokinase (PLAU), were further analyzed in functional assays, revealing a role in TGFβ1-mediated myofibroblast differentiation and over-production of components of the extracellular matrix (ECM) without interfering with the canonical TGFβ1 -signaling pathway. In this study, we delineate the genetic landscape of hypertrophic scars and present new insights into mechanisms involved in hypertrophic scar formation. Our data suggest the use of serine protease inhibitors for the treatment of skin fibrosis.

overall_design:
Single-cell transcriptome of human hypertrophic scars and human skin, and 6 and 8 weeks old mouse scars
>>> Raw data are unvailable due to patient privacy concerns <<<

last: Laggner

first: Dragan

last: Copic

first: Yiyan

last: Chen

first: Bahar

last: Golabi

first: Werner

last: Haslik

first: Chrisitine

last: Radtke

first: Erwin

last: Tschachler

first: Hendrik

last: Ankersmit

first: Michael

last: Mildner


title: Sequencing-based quantitative mapping of the cellular small RNA landscape

first: Dunaduan

last: Ma

type: Non-coding RNA profiling by high throughput sequencing

summary: Current next-generation RNA sequencing methods do not provide accurate quantification of small RNAs within a sample due to sequence-dependent biases in capture, ligation, and amplification during library preparation. We present a method – AQRNA-seq – that minimizes biases and provides a direct, linear correlation between sequencing read count and copy number for small RNAs in a sample. The library preparation and data processing steps were optimized and validated using a 963-member microRNA reference library, oligonucleotide standards of varying lengths, and northern blots. Application of AQRNA-seq to a panel of human cancer cells revealed >800 detectable miRNAs that varied as a function of cancer progression, while application to bacterial tRNA pools, a traditionally hard-to-sequence class of RNAs, revealed 80-fold variation in tRNA isoacceptor levels, stress-induced site-specific tRNA fragmentation, quantitative modification maps, and evidence for stress-induced tRNA-driven codon-biased translation. AQRNA-seq thus provides a means to quantitatively map the small RNA landscape in cells.

overall_design: 15  human mammary epithelial cell (HMEC)  samples including 3 cell types and 5 replicates each type were analyzed. The three cell types in the HMEC model represent progressive stages of tumorigenesis conferred by engineering the cells with tumor-promoting genes: reactivation of telomerase by expression of telomerase catalytic subunit (hTERT)  immortalizes HMEC 1 cells, additional expression of H-Ras oncoprotein (HRASG12V) further drives aberrant growth in HMEC 2 cells, and further P53 suppression by expression of SV40 large-T and small-t antigens yields HMEC 3 cells that are fully capable of tumor growth in mice.

first: Jennifer

middle: F

last: Hu

first: Daniel

last: Yim

first: Peter

last: Dedon

first: Bo

last: Cao


title: Single-cell Transcriptomics Identifies Gene Expression Networks Driving Differentiation and Tumorigenesis in the Human Fallopian Tube

status: Public on Apr 19 2021

city: Los Angeles

first: Robbin

last: Nameki

state: CA

address: 8700 Beverly Blvd

department: Obstetrics and Gynecology

institute: Cedars-Sinai Medical Center

summary:
The human fallopian tube harbors the cell-of-origin for the majority of high-grade serous ‘ovarian’ cancers (HGSCs), but its cellular composition, particularly of the epithelial component, is poorly characterized. We performed single-cell transcriptomic profiling in 12 primary fallopian specimens from 8 patients, analyzing around 53,000 individual cells to map the major immune, fibroblastic and epithelial cell types present in this organ. We identified 10 epithelial sub-populations, characterized by diverse transcriptional programs including SOX17 (enriched in secretory epithelial cells), TTF3 and RFX3 (enriched in ciliated cells) and NR2F2 (enriched in early, partially differentiated secretory cells). Based on transcriptional signatures, we reconstructed a trajectory whereby secretory cells differentiate into ciliated cells via a RUNX3high intermediate. Computational deconvolution of the cellular composition of advanced HGSCs based on epithelial subset signatures identified the ‘early secretory’ population as a likely precursor state for the majority of HGSCs. The signature of this rare population of cells comprised both epithelial (EPCAM, KRT) and mesenchymal (THY1, ACTA2) features, and was enriched in mesenchymal-type HGSCs (P = 6.7 x 10-27), a group known to have particularly poor prognoses. This cellular and molecular compendium of the human fallopian tube in cancer-free women is expected to advance our understanding of the earliest stages of fallopian epithelial neoplasia.
PAX8, SOX17, MECOM, and WT1 are critical genes in high-grade ovarian cancer tumorigenesis. However, the target genes of these factors in a normal to tumor context is unknown. Depletion of these factors in fallopian tube secretory epithelial and high-grade serous ovarian cancer cells revealed transcription factor and cell-type dependent differential expression patterns.

overall_design: siRNA knock-down of two non-targeting controls, PAX8, SOX17, DUAL(PAX8/SOX17), MECOM,and WT1 in cell lines followed by bulk RNA sequencing. This series includes re-analysis of all samples in GSE150443.

first: Huy

middle: Q

last: Dinh

first: Xianzhi

last: Lin

first: Forough

last: Abbasi

first: Marcela

last: Haro

first: Claire

last: Olingy

first: Heidi

last: Chang

first: Lourdes

last: Hernandez

first: Simon

last: Gayther

first: Kelly

middle: N

last: Wright

first: Paul-Joseph

last: Aspuria

first: Beth

first: Rosario

middle: I

last: Corona

first: Andrew

last: Li

first: B

middle: J

last: Rimel

first: Matthew

last: Siedhoff

first: Fabiola

last: Medeiros

first: Kate

last: Lawrenson