title: Deciphering the Mechanisms of CC-122 Resistance in DLBCL via a Genome-Wide Screen

city: San Diego

first: Rajkumar

last: Balakrishnan

state: California

address: 10300 Campus Point Drive, Suite 100,

department: Informatics & Predictive Sciences

institute: Bristol Myers Squibb

summary: CC-122 is a next-generation cereblon E3 ligase modulating agent that has demonstrated promising clinical efficacy in relapsed or refractory diffuse large B‐cell lymphoma (R/R DLBCL) patients. Mechanistically, CC-122 induces the degradation of IKZF1/3, leading to T cell activation and robust cell-autonomous killing in DLBCL. Here, we report a genome-wide CRISPR/Cas9 positive selection screen for CC-122 in a DLBCL cell line SU-DHL-4 with follow-up mechanistic characterization in 6 DLBCL cell lines to identify genes regulating the response to CC-122. Top-ranked CC-122 resistance genes encode not only well-defined members or regulators of the CUL4-DDB1-RBX1-CRBN E3 ubiquitin ligase complex, but also key components of several signaling and transcriptional networks that have not previously been shown to modulate the response to other cereblon modulators. Ablation of CYLD, NFKBIA, TRAF2, or TRAF3 induces hyper-activation of the canonical and/or non-canonical NF-κB pathways and subsequently diminishes CC-122-induced apoptosis in 5 out of 6 DLBCL cell lines. Depletion of KCTD5, the substrate receptor of the CUL3-RBX1-KCTD5 ubiquitin ligase complex, promotes the stabilization of its cognate substrate, GNG5, resulting in CC-122 resistance in HT, SU-DHL-4, and WSU-DLCL2. Furthermore, knockout of AMBRA1 renders resistance to CC-122 in SU-DHL-4 and U-2932, whereas knockout of RFX7 leads to resistance specifically in SU-DHL-4. The ubiquitous and cell line-specific mechanisms of CC-122 resistance in DLBCL cell lines revealed in this work pinpoint genetic alternations that are potentially associated with clinical resistance in patients, and facilitate the development of biomarker strategies for patient stratification, which may improve clinical outcome of CC-122 for R/R DLBCL.

overall_design: SU-DHL-4 cells treated with either DMSO or 10 µM CC-122 were profiled to identify changes in transcriptome-wide expression using RNA-seq. Biological replicates were used for RNA sequencing, where 150 bp PE-reads were obtained using Illumina Hiseq 4000 Sequencer.

first: Zhongying

last: Mo

first: Scott

last: Wood

first: Shawn

last: Namiranian

first: Reina

last: Mizukoshi

first: Stephanie

last: Weng

first: In

last: Jang

first: Celia

last: Fontanillo

first: Joshua

last: Baughman

first: Michelle

last: Slade

first: Arianna

last: Silva-Torres

first: Marwa

last: Khater

first: Kai

last: Wang

first: Mark

last: Rolfe

first: Gang

last: Lu


title: Non-viral Induction of Transgene-free iPSCs from Somatic Fibroblasts of Multiple Mammalian Species [pig]

status: Public on Oct 01 2020

city: Shinjuku

first: Sho

last: Yoshimatsu

state: Tokyo

address: 35 Shinanomachi

department: Physiology

country: Japan

institute: Keio University School of Medicine

type: Expression profiling by high throughput sequencing

summary: Induced pluripotent stem cells (iPSCs) are capable of providing an unlimited source of cells from all three germ layers as well as germ cells. The derivation and usage of iPSCs from various animal models may facilitate stem-cell-based therapy, generation of gene-modified animals, and evolutionary studies assessing interspecies differences. However, there is a lack of species-wide methods for deriving iPSCs, in particular by means of non-viral and non-transgene-integrating (NTI) approaches. Here, we demonstrated the derivation of iPSCs from somatic fibroblasts of multiple mammalian species from three different taxonomic orders, including the common marmoset (Callithrix jacchus) in Primates, the dog (Canis lupus familiaris) in Carnivora, and the pig (Sus scrofa) in Cetartiodactyla, by combinatorial usage of chemical compounds and NTI episomal vectors. Interestingly, the somatic fibroblasts temporarily acquired a neural stem cell (NSC)-like state during the reprogramming procedure. Collectively, our method, robustly applicable to various species, holds a great potential for facilitating stem-cell-based research using various animals in Mammalia.

overall_design: RNA seq of porcine fibroblasts, iNSLCs, and iPSCs

first: Mayutaka

last: Nakajima

first: Hideyuki

last: Okano


title: Non-viral Induction of Transgene-free iPSCs from Somatic Fibroblasts of Multiple Mammalian Species [human]

overall_design: RNA seq of human fibroblasts, iNSLCs, and iPSCs


title: Non-viral Induction of Transgene-free iPSCs from Somatic Fibroblasts of Multiple Mammalian Species [dog]

overall_design: RNA seq of canine fibroblasts, iNSLCs, and iPSCs


title: Non-viral Induction of Transgene-free iPSCs from Somatic Fibroblasts of Multiple Mammalian Species

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

sample_organism:
Callithrix jacchus
Homo sapiens
Canis lupus familiaris
Sus scrofa

overall_design: Refer to individual Series


title: EZH2 Inhibition Activates a dsRNA-STING-Interferon Stress Mechanism that Potentiates Response to PD-1 Check-Point Blockade in Prostate Cancer

city: Boston

first: Xintao

last: Qiu

state: MA

address: 440 brookline ave

country: USA

institute: Dana Faber Cancer Institute

summary: Prostate cancers are considered immunologically ‘cold’ tumors given the very few patients who respond to checkpoint inhibitor therapy (CPI). Recently, enrichment of interferon stimulated genes (ISGs) predicts a favorable response to CPI across various disease sites. The enhancer of zeste homolog-2 (EZH2) is over-expressed in prostate cancer and is known to negatively regulate ISGs. Here, we demonstrate that a majority of ISGs are not directly silenced by EZH2 mediated H3K27me3 deposition, but instead in a poised chromatin state associated with accessible chromatin (ATAC+) and H3K27ac deposition. Inhibition of EZH2 catalytic activity in prostate cancer models activates a dsRNA-STING-ISG cellular stress response upregulating genes involved in antigen presentation, Th-1 chemokine signaling, and interferon (IFN) response, including PD-L1 that is dependent on STING activation. EZH2 inhibition combined with PD-1 CPI significantly enhances anti-tumor response dependent on up-regulation of tumor PD-L1 expression. Further, combination therapy significantly increases intratumoral trafficking of activated CD8+ T-cells and M1 tumor associated macrophages (TAMs) with concurrent loss of M2 TAMs. Our study identifies EZH2 as a potent inhibitor of antitumor immunity and responsiveness to CPI. This data suggests EZH2 inhibition is a novel therapeutic direction to enhance prostate cancer response to PD-1 CPI.

overall_design: Mouse prostate organoids generated from genetically engineered mouse models housing overexpression of cMYC, EZH2 floxed alleles, and an inducible PSA-CreERT2 were generated. The purpose was to compare control verse EZH2 inhibition effects on gene expression changes. Organoid EZH2 activity was inhibited chemically by  treatment with the EZH2 inhibitor DZNep (verse DMSO control) or genetically using Tamoxifen (verse EtOH control). 3 days post treatment, organoids were used to generate RNA and be analyzed by RNA-seq.

first: Leigh

last: Ellis


title: In Situ Maturation and Tissue Adaptation of Type 2 Innate Lymphoid Cell Progenitors

status: Public on Oct 02 2020

city: Freiburg

first: Patrice

last: Zeis

address: Stübeweg 51

country: Germany

institute: Max Planck Institute of Immunobiology and Epigenetics

summary: We analyzed the full spectrum of lung ILC2s at single-cell transcriptome resolution and identified an immature population of Il18r1+ tissue-associated ILCs, that express Tcf7 and Zbtb16, transcription factors known to act in ILC progenitors, and low levels of Gata3, ST2 and ILC2 effector molecules. We revealed a continuous differentiation trajectory and validated the potential of immature Il18r1+ cells to differentiate into Gata3hi ILC2s. We found that immature ILCs were recruited into the adult lung of parabiotic and shield chimeric mice during helminth infection. Intriguingly, recruited cells generated the entire spectrum of ILC2s in the lung, consistent with their potential role in the renewal of tissue ILC2s. Our work identifies local ILC progenitors and highlights their tissue adaptation and in situ differentiation as a mechanism of ILC maintenance and phenotypic diversification

overall_design: Comprehensive single-cell atlas and transcriptome comparison of ILC populations in the BM(Lin-IL18r1+Icos+), and neonatal(liveLin-CD45+Thy1+IL7r+Nk1.1-ST2+/-) and adult lung (LiveLin-CD45+Thy1+IL7r+Nk1.1-) at steady-state, as well as over the course of helminth infection in wild-type, parabiotic and shield chimeric mice,

first: Dominic

last: Grün


title: Exploring the involvement of Adathoda vasica in hypoxia axis  at the molecular level

status: Public on Sep 30 2020

city: Delhi

first: Mitali

last: Mukerji

address: Mathura Road

country: India

institute: CSIR-IGIB

summary:
The role of increased hypoxia has been identified for the many pathological features of SARS-CoV2, such as impaired immunity, airway hyper-inflammation, fibrosis, sepsis, and thrombosis. In this study, we tested the potential effect of Adhatoda Vasica (AV) aqueous extract, an ayurvedic anti-hypoxic, and anti-inflammatory medicine, on SARS-CoV2 infection using genome-wide expression approach.
In this study, we did the genome-wide expression analysis of the lungs of mice treated with AV and compared them with transcriptome data of COVID19 patients. A meta-analysis of transcriptome profiles shows that AV may serve as a potential drug candidate to boost immune response and to prevent the transcriptional response elicited by SARS-CoV2 infection.

overall_design: For the transcriptomic experiment, distilled water (vehicle) or AV administered to naïve BALB/c mice by oral gavage for four consecutive days. Lung total RNA was isolated on 4th day of experiment, and a microarray experiment performed using Affymetrix Gene Chip MTA 1.0 array according to manufacturer’s instruction. Gene expression analysis was performed using Affymetrix® Expression Console and functional analysis done by using the Enrichr tool.

first: Atish

last: Gheware

first: Dhwani

last: Dholakia

first: Bhavana

last: Prasher


title: Exosomes derived from hypertrophic cardiomyocytes induce inflammation in macrophages via miR-155 mediated MAPK pathway.

status: Public on Sep 25 2020

city: Shanghai

first: Li

last: Zhang

address: Haowen Road no.5

country: China

institute: Fulgent Genetics

type: Non-coding RNA profiling by high throughput sequencing

summary: We report the application of RNA sequencing technology for high-throughput profiling of normal cardiomyocytes-derived and hypertrophic cardiomyocytes- derived exosomes. The miRNA expression profiles were determined by high throughput miRNA sequencing, and 635 differentially expressed miRNAs were found. However, compared with the control group, 7 miRNAs were significantly differentially expressed in exosomes released from Ang II-treated cardiomyocytes. A total of 4 miRNAs were upregulated while 3 were downregulated. We used qRT-PCR to characterize relative expression levels of miR-155 and miR-212-3p to validate the data obtained through miRNA sequencing. The results obtained through qRT-PCR and miRNA sequencing were essentially identical. To elucidate the potential role of miRNAs in hypertrophic cardiomyocytes, the prediction of miRNA targets was performed and 11,637 genes were obtained. To reduce the false positives rate of target gene prediction, only the predicted targets within the 3 databases described above were further analyzed. Finally, 5,477 predicted target genes were selected for further investigation. Our results support the concept that exosomal microRNAs have emerged as important inflammatory response modulators regulating the cardiac hypertrophy.

overall_design: Examination of normal cardiomyocytes-derived and hypertrophic cardiomyocytes- derived exosomes.

first: Hui

last: Yu

first: Lei

last: Qin

first: Yunzhi

last: Peng

first: Wenhui

last: Bai

first: Zhanli


title: The vitamin D receptor (VDR) autonomously regulates skeletal muscle mass

status: Public on Sep 29 2020

city: Stirling

first: Iain

last: Gallagher

address: Room 4B133 Cottrell Building, University of Stirling, Airthrey Road

department: Faculty of Health Sciences and Sport

country: United Kingdom

institute: University of Stirling

summary: Vitamin D (VitD) deficiency is estimated to affect ~40% of the world’s population. Notably, VitD deficiency has been associated with impaired muscle maintenance and insulin resistance. VitD exerts its actions through the ubiquitous Vitamin D-receptor (VDR), the expression of which was recently confirmed in fully-differentiated muscle. To seek a possible autonomous role of the VDR in skeletal muscle, we first generated stable VDR-knockdown cells, which exhibited impaired myogenesis (i.e. cell-cycling, differentiation and myotube formation). In vivo VDR-knockdown in rat hind-limbs elicited myofibre atrophy and triggered autophagy pathways. In contrast, in vivo VDR-overexpression yielded myofibre hypertrophy; enhancing translational efficiency (e.g. mTOR-signaling), ribosomal biogenesis and satellite cell content. Neither VDR-knockdown nor overexpression impacted muscle glucose uptake. Crucially, induction of VDR mRNA correlated with muscle hypertrophy in humans following long-term resistance exercise training, but not aspects of insulin sensitivity. The VDR autonomously regulates muscle mass, acting reciprocally to limit atrophy and promote hypertrophy.

overall_design: In vivo electroporation was used to knock out (ko) or knock in (ki) the vitmain D receptor in the tibialis anterior muscle of one hindlimb in rats. VDR-KD groups received VDR shRNA (Table S1) in right leg TAM and scramble shRNA in left leg TAM. VDR-OE groups received pCAGGS-mVDR in right leg TAM, and empty pCAGGS controls in left leg TAM. Immediately following, one HV (900 V/cm, 100 µ-sec pulse), and four LV (90 V/cm, 100 m-sec) pulses were administered across the distal limb via tweezer-electrodes attached to an ECM-830 electroporator (BTX, Holliston, MA). Animals subsequently received a subcutaneous injection of carprofen (50mg/kg), before recovery from anesthesia. RNA-seq was carried out with the contrast of interest being either ko v control limb or ki v control limb.

first: Joseph

middle: J

last: Bass

first: Philip

last: Atherton


title: Comparative analysis of global transcriptional profile of Escherichia coli and Schizosaccharomyces pombe in respiratory and fermentative growth: Inferring the evolution of analog circuits of post-transcriptional regulation

city: Cuernavaca

first: Veronica

last: Jimenez-Jacinto

state: Morelos

address: Av. Universidad 2001. Col. chamilpa

department: Unidad Universitaria de Secuenciación Masiva y Bioinformática

country: Mexico

institute: Universidad Nacional Autonoma de Mexico

summary: Next Generation sequencing techniques have ushered in a new era in life sciences, especially genomic research. Currently, massive RNA sequencing (RNA-seq) has increased knowledge about the "RNA universe" from the global characterization of transcriptomes, differential gene expression studies and the continued discovery of non-coding regulatory transcripts such as non-coding RNAs (ncRNAs). Post-transcriptional gene regulation is a mechanism for controlling gene expression that is present in both prokaryote and eukaryote organisms and regulates multiple biological processes. Examples of ncRNAs that regulate the levels of transcripts of genes related to central carbon metabolism have been described in bacteria and higher eukaryotes. In order to describe whether analogies exist in regulatory circuits mediated by ncRNAs in two cell patterns, a comparative analysis of the transcriptome of Escherichia coli and Schizosaccharomyces pombe in similar growth conditions was carried out using two carbon source, 2 % glucose and a mix of 2% glycerol and 0.2 % sodium acetate. Twelve RNA-seq libraries were performed. The RNA-seq data contained a number unambiguously mapped readings on the genome of each organism ranged from 1-1.3 million. A total of 739 and 743 genes were detected as differentially expressed for E. coli and S, pombe respectively with a module of log FC > 1 and FDR > 0.05. In aerobic glicolisis GO asciated to cell proliferetionweres detected in both organisms. Among the genes differentially expressed in glycerol/acetate general transcription factors such as fliA, 11 literature validated sRNAs, and assembly flagellum maquinary gene were detected in E. coli, whereas in fission yeast, hexose transporters, switches integral membrane proteins and ncRNAs related to cellular stress are highlighted. In general, our study shows that a conserved "foraging behaviour" response is observed in eukaryotic and procaryotic organism when there is no present glycolityc carbon source.

sample_organism:
Escherichia coli
Schizosaccharomyces pombe

overall_design: 12 Ribosomal depleted Total RNA profiles of mid-log phase of E. coli K-12 MG1655  and S. pombe 972 h- in two growth  were generated by deep sequencing, in triplicate, using Illumina Next-seq.

first: Joivier

last: Vichi

first: Emmanuel

last: Salazar

first: Verónica

last: Jiménez

first: Leticia

last: Olvera

first: Ricardo

last: Grande

first: Enrique

last: Morett

first: Edgar

last: Dantán

first: Armando

last: Hernández


title: RNA sequencing analysis of SLC6A15 overexpression in papillary thyroid cancer cells

status: Public on Sep 21 2020

city: Guangzhou

first: Yu

middle: xin

last: Chen

state: Guangdong

address: 58 Zhongshan Road II

department: Department of Endocrinology and Diabetes Center

institute: The First Affiliated Hospital of Sun Yat-sen University

summary: Solute Carrier Family 6 Member 15 (SLC6A15) has been found with dysregulated expression in several kinds of cancers. However, its expression pattern and biological functions in papillary thyroid cancer (PTC) remain unknown. To explore the role of SLC6A15 in PTC, transcription profiling analysis was performed in KTC1 cells with or without SLC6A15 overexpression.

overall_design: The mRNA profiles of KTC1-Vector and KTC1-SLC6A15 cells were generated by deep sequencing, in duplicate, using BGISEQ-500

first: Yuxin

first: Hai

last: Li

first: Hongyu

last: Guan

first: Weiwei

last: Liang

first: Yanbing


title: MicroRNAs and Ascl1 facilitate direct conversion of porcine fibroblasts into induced neurons

status: Public on Sep 22 2020

city: Aarhus C

first: Mette

last: Habekost

state: Midtjylland

address: HØEGH-GULDBERGSGADE 10

department: Department of Biomedicine

country: Denmark

institute: Aarhus University

summary: Direct neuronal conversion describes the process of generating induced neurons from somatic cells such as fibroblasts by overexpressing cell type-specific transcription factors. This was first achieved by expressing Brn2, Ascl1 and MytL1 in mouse fibroblasts, and was later achieved in human cells by inclusion of additional factors such as NeuroD1. Here, we present the first protocol for directly converting porcine fibroblasts into induced neurons. We used lentivirus-mediated delivery of previously identified neuronal-specifying transcription factors and microRNAs and evaluated morphology and neuronal marker expression after ten days of conversion. We found that Ascl1 and microRNAs, miR-9/9* and miR-124 together generated more neuronal cells than other conditions tested. The porcine induced neurons expressed common mature markers such as MAP2 and Synaptophysin after four weeks of conversion. Transcriptomic analysis revealed that fibroblast-specific signatures were silenced early in the conversion process, while the neuronal programs increased and matured during conversion. We generated a heterogenous population of glutamatergic and GABAergic neurons.

overall_design: RNA sequencing of porcine induced neurons from fibroblast at three different timepoints during conversion. Three replicates were used for each timepoint.


title: Distinct roles of Bdh2 in regulating embryonic stem cell fate

city: Hohhot

first: Yuting

last: Fu

address: university west road

institute: Inner Mongolia University

summary: To further gain insights into the role of Bdh2 in cell fate decisions of mouse ESCs, we generated Bdh2 homozygous knockout mouse advanced pluripotent embryonic stem cell (ASC) and embryonic stem cell (ESC) lines by CRISPR/Cas9 genome editing technology. Bdh2 deficiency in ASCs/ESCs exhibited no effect on expression of core pluripotent transcription factors and alkaline phosphatase activity, suggesting dispensability of Bdh2 for self-renewal and pluripotency of embryonic stem cells. Notebly, Bdh2-knockout ASCs/ESCs exhibit higher expression of the endodermal somatic genes, Gata4, Gata6, Sox17,and also colocalizated with Oct4. Importantly, expression of key DNA methylation genes Dnmt3a, Dnmt3b and Dnmt1 decreased in Bdh2-knockout ASCs/ESCs. Overall, we provide insight on understanding of earliest cell fate decicion in ASCs/ESCs and a new avenue to create and preserve pioneer lineage precursors with pluripotency.

overall_design: 3 replicates of mRNA of mouse embryonic stem cells mRNA profiles of wild type (WT) and Bdh2-/- cell lines

first: Jia

last: Zhang

first: Fangyuan

last: Liu

first: Shuo

last: Cao


title: Onco-fetal reprogramming of endothelial cells drives immunosuppressive macrophages in Hepatocellular Carcinoma

status: Public on Sep 24 2020

city: Singapore

first: Ankur

last: Sharma

state: Singapore

address: 60 Biopolis Street, Genome, #02-01

country: Singapore

institute: Genome Institute Of Singapore

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
Refer to individual Series
The linked supplementary file includes:
sheet 1: patient information
sheet 2: Top-25 genes HCC atlas clusters
sheet 3: Top-25 genes T cell clusters
sheet 4: Top-25 genes NK cell clusters
sheet 5: Top-25 genes Fibroblast clusters
sheet 6: Top-25 genes Hepatocyte clusters
sheet 7: Garnett Markers-whole atlas
sheet 8: Top-25 genes MNC clusters
sheet 9: Top-25 genes Endothelial clusters
sheet 10: Garnett Markers-myeloid
sheet 11: Spatial Transcriptomics


title: AKT/mTOR/HER/PI3K family proteins and phospho-proteins as predictors of pCR for early stage breast cancer patients treated with the AKT inhibitor MK2206 in the I-SPY 2 TRIAL (NCT01042379)

status: Public on Sep 20 2020

city: Manassas

first: Julia

last: Wulfkuhle

state: VA

address: 10920 George Mason Circle

institute: George Mason University

summary: The AKT inhibitor MK2206 (M) was one of the experimental agents evaluated in I-SPY 2, a neoadjuvant platform trial for high risk, early stage breast cancer, and graduated in the HER2+, HR-, and HR-HER2+ signatures.  Based on the hypotheses that since MK2206 is an enzymatic inhibitor of AKT, response to MK2206 may be predicted by the relative pre-treatment levels of phosphorylation of AKT kinase substrates, in pre-defined analyses we assessed 26 well known proteins/phospho-proteins  in the HER-AKT-mTOR pathway to test their association with pCR in the MK2206 arm, in both HER2+ and HER2- subsets. We specifically used laser capture microdissection (LCM)- enriched tumor epithelium cell isolates from the clinical biopsy specimens as the sample input for all protein pathway activation analysis, which has been shown to be critical in accurate measurement of ubiquitously expressed and activated signaling proteins.  To the best of our knowledge, this work represents the first rigorous study of association between  direct measurement of AKT pathway activation using phospho-protein activation levels and response to MK2206 performed in early stage breast cancer patients.

overall_design:
Reverse phase protein array (RPPA) array run on isolates from pre-treatment breast cancer biopsy samples that were first LCM-enriched for epithelial cells.
The sample characteristics and their values represent:
HER2: the status of the HER2 gene. 1 = positive; 0 = negative.
HR: the status of hormone (estrogen and progesterone) receptors.  1 = positive; 0 = negative.
pCR: the status of a pathological complete response (pCR).  1 = complete response; 0 = failed complete response
Arm: arm of the trial per patient - MK2206 arm' or 'Control arm'.
RPPA Array:  Platform Array version.  Key to convert from raw to normalized data.

first: Denise

last: Wolf

first: Christina

last: Yau

first: Emanuel

middle: F

last: Petricoin III

first: Laura

last: van ‘t Veer


title: Characterization of small RNAs in early zebrafish PGCs

status: Public on Sep 13 2020

first: XiaoYuan

address: 320 Yueyang road

institute: Shanghai Institute of Biochemistry and Cell Biology,CAS

summary: Microscale small RNA high-throughput sequencing was used to study small RNA distribution in early zebrafish PGCs (primordial germ cells). We find that early zebrafish PGCs have large quantities of piRNAs and small quantities of miRNAs. Among the miRNAs detected, miR-430 accounted for the majority which has been proved to play diverse roles in zebrafish development. In addition, miR-92a-3p and miR-26a-5p have high expression and previous studies show that these two miRNAs can influence cell proliferation in cancer cells. More experiments should be performed in the future to explore whether miR-92a-3p and miR-26a-5p can influence the number of zebrafish PGCs.

overall_design: Small RNA profiles at 6 hpf (hours post fertilization, shield stage), 11 hpf (3-somite stage) and 24 hpf (prim-5 stage) of zebrafish PGCs

first: Xiaoyuan


title: Onco-fetal reprogramming of endothelial cells drives immunosuppressive macrophages in Hepatocellular Carcinoma (scRNA-seq)

summary: Liver dysfunction is associated with diseases ranging from metabolic disorders to hepatocellular carcinomas (HCC). Here we employed single-cell RNA-sequencing to extensively characterise the cellular landscape of human liver, from development to disease. We analysed ~212,000 cells representing human fetal liver, HCC and mouse liver. Our analysis revealed a remarkable fetal-like reprogramming of the tumor microenvironment (TME). Specifically, the HCC ecosystem displayed features reminiscent of fetal development, including the re-emergence of fetal-associated endothelial cells (PLVAP+/VEGFR2+), and fetal-like (FOLR2+) tumor-associated macrophages (TAMs). In a cross-species comparative analysis, we discovered remarkable similarity of gene expression and regulatory networks between mouse embryonically-seeded, fetal-liver and FOLR2+ tumor macrophages. Spatial transcriptomics further corroborated a shared onco-fetal ecosystem between fetal-liver and HCC. Furthermore, gene regulatory analysis, spatial transcriptomics, and in vitro functional assays implicated VEGF and NOTCH signaling in maintaining onco-fetal ecosystem. Taken together, we report a shared immunosuppressive onco-fetal ecosystem between the human fetal-liver and HCC. Our results unravel a previously unexplored onco-fetal reprogramming of tumor ecosystem, provides a novel target for therapeutic interventions in HCC and also opens up avenues for identifying similar paradigms in other cancers and disease states.

overall_design: Analysis of hepatocellular carcinoma (HCC), healthy normal liver, human fetal liver and mouse liver samples

middle: W

last: Seow

first: Charles-Antoine

last: Dutertre

first: Rhea

last: Pai

first: Camille

last: Blériot

first: Archita

last: Mishra

first: Regina

middle: M

last: Wong

first: Gurmit

middle: S

last: Singh

first: Samydurai

last: Sudhagar

first: Shabnam

last: Khalilnezhad

first: Sergio

last: Erdal

first: Hui

last: Teo

first: Ahad

first: Svetoslav

last: Chakarov

first: Tony

middle: K

last: Lim

first: Alexander

middle: Y

last: Chung

first: Alfred

last: Kow

first: Peng

middle: C

last: Cheow

first: Glenn

last: Bonney

first: Brian

last: Goh

first: Jerry

last: Chan

first: Pierce

last: Chow

first: Florent

last: Ginhoux

first: Ramanuj

last: DasGupta


title: Gene expression profile of mouse liver in response to activation of PPARA by WY-14643.

status: Public on Sep 14 2020

first: David

middle: J.

last: Waxman

address: 5 Cummington Mall

department: Department of Biology and Bioinformatics Program

institute: Boston University

summary: Gene expression in adult male mouse liver was assayed by RNA-seq, as part of a study of liver transcriptomic changes in response to activation of PPARA by WY-14643.

overall_design: Twelve independent pools, comprised of RNA isolated from n=3-5 individual 12 wk old male livers per pool, were prepared and used for RNA-seq. Three pools were untreated, intact male mice livers; three pools were treated (WY-14,643), intact male mice livers; three pools were untreated, PPARA-KO mouse livers and three pools were treated, PPARA-KO mice livers. Wy-14,643 treatment was via the diet, at 0.1% for 48 hr.

first: Chad

middle: N

last: Brocker

first: Tisha

last: Melia

first: Frank

last: Gonzalez


title: In vivo transcriptome analysis provides insights into host-dependent expression of virulence factors by Yersinia entomophaga MH96, during infection of Galleria mellonella

status: Public on Sep 01 2020

city: Christchurch

first: Amber

middle: Rose

last: Paulson

address: Lincoln Research Centre, Cnr Springs Road And Gerald Street, Private Bag 4749

department: Forage Science

country: New Zealand

institute: AgResearch Litmited

summary: Genome-wide transcriptional responses of Yersinia entomophaga to shifts in temperature (25 - 37 degrees C) and in vitro and in vivo growth conditions were investigated using RNA-seq.

overall_design: In vivo samples included four biological replicates and in vitro samples included two biological replicates. At 25 degrees, lag, exponential and stationary growth stages were sampled from in vitro and in vivo conditions. At 37 degrees only exponential growth conditions were sampled.

middle: R

first: Paul

middle: B

last: Rainey

first: Xue-Xian

last: Zhang

first: Maureen

last: O'Callaghan

first: Mark

last: Hurst


title: Analysis of genomic aberrations by array comparative genomic hybridization (aCGH) of tumor tissues of mice after 12 months on CD-HFD (n= 9) or 12 months on CD-HFD plus 8 weeks treatment with α-PD-1 (n= 12)

status: Public on Aug 31 2020

city: Munich

first: Kristian

last: Unger

address: Marchioninistr. 15

department: Department of radiation oncology

institute: LMU University Hospital

summary: Genomic DNA was extracted from tumors that developed in C57BL/6 mice fed with CD-HFD for 12 months alone or fed with CD-HFD for 12 months followed by 8 weeks of treatment with anti-PD-1. DNA extracted from five pooled tail-clips of  C57BL/6 mice was used as reference DNA.

overall_design: the tumor profiles of 9 CD-HFD only mice was compared to that of 12 CD-HFD/anti-PD1-treated mice.

first: Dominik

last: Pfister

first: Mathias

last: Heikenwälder

last: Ungr


title: Oocyte RNA-seq data of WT, Tet1-/- and Tet2-/-; RNA seq of PGC at E13.5

status: Public on Sep 08 2020

city: Tianjin

first: wang

middle: hua

last: song

address: Tianjin city, China

institute: nankai university

summary: Differential functions of Tet 1 and Tet2 in oocyte development and quality control link to reproductive aging

overall_design: We generated Tet1/2 knock out mice to study the function of Tet1/2

first: Huasong

last: Wang

first: Linlin

last: Liu

first: Mo

last: Gou


title: Different expression of circRNAs in blood from idiopathic short stature patients and healthy controls

city: Nanchang

first: xijuan

last: liu

state: Jiangxi

address: No. 1 Minde Road

institute: The Second Affiliated Hospital of Nanchang University

summary: Idiopathic short stature is diagnosed by a standing height of less than two standard deviation scores in a specific population adjusted for age and gender and the exclusion of identifiable diseases. A series of studies have confirmed that noncoding RNAs can regulate the chondrocyte proliferation, hypertrophy, and endochondral ossification in the growth plate. In order to analyze and find differentially expressed circRNAs in Idiopathic short stature and healthy controls, we aimed to explore whether differentially expressed circRNAs in idiopathic short stature. Four pairs of blood samples were subjected to microarray analysis using the Arraystar Human CircRNAs Microarray v2 (Arraystar, USA). Compared to normal individuals, in ISS patients, the expression levels of 83 circRNAs were upregulated and those of 62 were downregulated.

overall_design: Four pairs of blood specimens from idiopathic short stature patients and healthy controls were used for circRNAs microarray analysis. RNA samples were prepared for microarray hybridization using Eastep Super RNA simple Total RNA Kit according to the manufacturer’s protocol. Total RNA was digested using RNase R to enrich the sample for circRNAs and eliminate linear RNAs.

first: Xijuan

first: Jingyu

last: Jia


title: The role of circular RNAs in porcine antral follicular degeneration

first: Li

last: Meng

address: Wushan Road 483

institute: South China Agricultural University

summary:
Goal:Circular RNAs (circRNAs) are thought to play important roles in multiple biological processes including apoptosis. In the ovary granulosa cell apoptosis plays an essential role in the follicular phase of the estrous cycle as it determines if the selected follicles will degenerate or escape this fate and ovulate. Whether circRNAs play a role in granulosa cell apoptosis is at present unknown. By investigating the potential role of circRNAs in the granulosa cell apoptotic cascade we hope to gain novel information regarding the regulation of antral follicular apoptosis using the porcine ovary as a model.
Method: Ribosomal-depleted RNA-sequencing was performed to generate circRNA expression profiles from isolated mural porcine granulosa cells of healthy antral (HA) and atretic antral (AA) follicles, respectively.
Results:In total, more than 9,632 circRNAs were identified, of which 62 circRNAs were differentially expressed (DE-circRNAs). GO and KEGG analysis of DE-circRNAs showed that apoptosis, cellular responses to stress and cell cycle pathways, were significantly enriched. Next the characteristics of DE-circRNAs, including back-splicing, RNase R resistance and stability were validated, and miRNA binding sites of DE-circRNAs were predicted.
Conclusion: Based on these observations, we conclude that aberrantly expressed circRNAs and their targeted genes seem to be associated with granulosa cell apoptosis and thus antral follicular atresia.

overall_design: circular RNA profiles of Porcine healthy antral (HA) and atretic antral (AA) were generated by deep sequencing, in triplicate, using Illumina.

first: Shouquan


title: Denatonium as a bitter taste receptor agonist modifies transcriptomic profile and functions of acute myeloid leukemia cells

status: Public on Sep 07 2020

city: Meldola

first: Giorgia

last: Simonetti

state: FC

address: Via Piero Maroncelli, 40

country: Italy

institute: IRCCS Istituto Romagnolo per lo Studio dei Tumori (IRST) "Dino Amadori"

summary: Bitter taste receptors (T2Rs) are typical G-protein coupled receptors expressed in various tissue where they are involved in the regulation of physiological processes, thus suggesting a wider function in sensing microenvironment. We analyzed their expression and role in acute myeloid leukemia (AML). AML cells express functional T2Rs and their stimulation with the agonist, denatonium benzoate, substantially modified the AML cell transcriptomic profile and functions. GEP analysis identified relevant cellular processes affected by denatonium treatment in AML, including cell cycle, survival, migration and metabolism. More precisely, T2R activation reduced proliferation by inducing cell cycle arrest in G0/G1 phase or induced apoptosis via caspase cascade activation; impaired AML cell motility and migratory capacity; inhibited cellular respiration by decreasing glucose uptake and oxidative phosphorylation.

overall_design: THP-1 and OCI-AML3 cell lines and primary AML cells treated with denatonium (or vehicle) for 24h were used for RNA extraction and hybridization on Affymetrix microarrays.

first: Valentina

last: Salvestrini

first: Marilena

last: Ciciarello

first: Antonio

last: Curti


title: CSF B cells in relapsing multiple sclerosis are driven towards an antigen-experienced, inflammatory fate

city: San Francisco

first: Akshaya

last: Ramesh

address: 675 Nelson Rising Lane

institute: UCSF

summary: Central nervous system B cells have several potential roles in multiple sclerosis (MS): secretors of proinflammatory cytokines and chemokines, presenters of autoantigens to T cells, producers of pathogenic antibodies, and reservoirs for viruses that trigger demyelination. To interrogate these roles, single-cell RNA sequencing (scRNA-Seq) was performed on paired cerebrospinal fluid (CSF) and blood from subjects with relapsing-remitting MS (RRMS; n = 12), other neurologic diseases (ONDs; n = 1), and healthy controls (HCs; n = 3). Single-cell immunoglobulin sequencing (scIg-Seq) was performed on a subset of these subjects and additional RRMS (n = 4), clinically isolated syndrome (n = 2), and OND (n = 2) subjects. Further, paired CSF and blood B cell subsets (RRMS; n = 7) were isolated using fluorescence activated cell sorting for bulk RNA sequencing (RNA-Seq). Independent analyses across technologies demonstrated that nuclear factor kappa B (NF-?B) and cholesterol biosynthesis pathways were activated, and specific cytokine and chemokine receptors were up-regulated in CSF memory B cells. Further, SMAD/TGF-ß1 signaling was down-regulated in CSF plasmablasts/plasma cells. Clonally expanded, somatically hypermutated IgM+ and IgG1+ CSF B cells were associated with inflammation, blood–brain barrier breakdown, and intrathecal Ig synthesis. While we identified memory B cells and plasmablast/plasma cells with highly similar Ig heavy-chain sequences across MS subjects, similarities were also identified with ONDs and HCs. No viral transcripts, including from Epstein–Barr virus, were detected. Our findings support the hypothesis that in MS, CSF B cells are driven to an inflammatory and clonally expanded memory and plasmablast/plasma cell phenotype.

overall_design: 5' or 3' Single-cell RNA sequencing was performed on the CSF and blood for a cohort of relapsing MS patients, patients with other neurologic diseases, and healthy controls. 5' Single-cell immunoglobulin sequencing was performed on a subset of these subjects and additional subjects with relapsing MS, clinically isolated syndrome, and patients with other neurologic diseases. Additionally, total RNA from 10 B cell subsets  (5 Blood and 5 CSF) from a separate cohort of relapsing MS patients were sorted using FACS and sequenced using bulk RNA sequencing.

first: A

first: R

middle: D

last: Schubert

middle: L

last: Greenfield

last: Dandekar

last: Loudermilk

first: J

last: Sabatino

first: M

middle: T

last: Koelzer

first: E

last: Tran

first: K

last: Koshal

last: Kim

last: Pröbstel

first: D

last: Banerji

first: C

last: Guo

last: Green

last: Bove

last: DeRisi

last: Gelfand

first: B

last: Cree

first: S

last: Zamvil

middle: E

last: Baranzini

last: Hauser

last: Wilson


title: MBD-seq in ovarian tissues of prenatally androgenized (PNA) mice

status: Public on Aug 28 2020

city: Minhang district of Shanghai

first: yani

last: Kang

address: Dongchuan Road 800

institute: Shanghai Jiaotong University

type: Methylation profiling by high throughput sequencing

summary: To investigate the molecular mechanism of PCOS underlying the hyperandrogenic phenotype, prenatally androgenized (PNA) mice were used to mimic this phenotype in women with PCOS. Methylated DNA binding domain sequencing (MBD-seq) and RNA-seq were performed on PNA mice (n=6) and control group (n=4) and validations were applied on ovarian samples from PNA mice (n=6) and control group (n=6) using MSP (methylation-specific PCR) and qPCR. The immunohistochemistry (IHC) and transmission electron microscope (TEM) profiling was separately conducted on tissue sections and granular cells of PNA mice (n=6) and control group (n=6).  We identified 857 genes with differently methylated promoters and 3317 differently expressed genes in PNA mice and control group. We found that PCOS group had a down-regulation of Dnmt1 gene expression, accompanied by global hypomethylation compared with the control group. The promoter regions of Map3k1(mitogen-activated protein kinase kinase kinase 1) and Map1lc3ka (microtubule-associated protein1 light chain 3) were hypomethylated, accompanied by up-regulation of their mRNA expression, which may be involved in the regulation of PCOS through MAPK/p53 pathway activition and autophagy alteration.

overall_design: MBD-seq combined with RNA-seq analysis to find epigenetic regulation in PCOS

first: Yulan

last: Qin

first: Ting

last: Li

first: Hui

last: Zhao

first: Zhanrui

last: Mao

first: Yani


title: Determination of isoform-specific RNA structure with nanopore long reads

status: Public on Sep 09 2020

city: singapore

first: Aw

last: Jong Ghut Ashley

address: 60 Biopolis St

institute: Genome Institute of Singapore

summary: Current methods for determening RNA structure with short-read sequencing cannot capture most differences between distinct transcript isoforms. Here, we present RNA structure analysis using nanopore sequencing (PORE-cupine),which combines structure probing using chemical modifications with direct long-read RNA sequencing and machine learning to detect secondary structures in cellular RNAs. PORE-cupine also captures global structural features, such as RNA binding protein (RBP) binding sites and reactivity differences at single nucleotide variants (SNVs). We show that shared sequences in different transcript isoforms of the same gene can fold into different structures,  highlighting the importance of long-read sequencing for obtaining phase information. We also demonstrate that structural differences between transcript isoforms of the same gene lead to differences in translation efficiency. By revealing isoform-specific RNA structure, PORE-cupine will deepen our understanding of the role of structures in controlling gene regulation.

sample_organism:
Bacillus subtilis
Saccharomyces cerevisiae
Tetrahymena thermophila
Homo sapiens
Dengue virus
Oceanobacillus iheyensis

overall_design:
90 libraries in total
The libraries consist of two types of samples, unmodified control and modified libraries.
Training libraries were used to train the models.
Test libraries were used to evaluate the parameters obtained from training.
TPP libraries were used to determine structural changes between samples incubated with and without its ligand.
H9 libraries were used to study structural changes in the human transcriptome.
Two replicates of H9 were sequenced with Illumnia sequencing.

first: Jong

middle: G

last: Aw

first: Shaun

last: Lim

first: Jia

middle: X

last: Wang


title: Normal and DNMT3L-overexpressing human neurons

status: Public on Aug 29 2020

city: Shenyang

first: JIE

last: LU

state: Liaoning

address: 77 Puhe Road, Shenbei Xiqu

department: Human Anatomy

institute: China Medical University

type: Methylation profiling by genome tiling array

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

overall_design: Refer to individual Series


title: RNA-seq in ovarian tissues of prenatally androgenized (PNA) mice


title: A CD4 T cell population expand and reshape autoreactive B cells, contributing to autoimmune lung pathology and post-infection autoimmunity in children

status: Public on Aug 26 2020

first: Yuxia

last: Zhang

address: 9 Jinshui Rd, Tianhe District

institute: Guangzhou Women and Children's Medical Center

summary: Community acquired pneumonia (CAP) is a significant risk factor for autoimmune disease development. However, mechanisms underlying infection and autoimmunity remain elusive. Here, we report selective expansion of autoantibody producing CD21-CD19+ B cells by bronchoalveolar PD-1+CCR2+GZMA+RUNX3+ T helper 1 like (aTh1) cells in children with CAP. In bronchoalveolar lavage, the numbers of aTh1, CD21-B cells and the concentrations of IgG-specific autoantibodies correlated with CAP severity. Unexpectedly, PD-1 decreased the ability of CD21- B cells to produce autoantibodies and reshaped their specificity in the course of pathogen invasion. Nevertheless, respiratory infection induced aTh1 and CD21-B cell persistence may trigger autoimmune disease development in susceptible individuals; e.g. they were prevalent in cerebrospinal fluid of children with post-infection Guillain-Barre Syndrome. Thus, we reveal dual roles of aTh1 and CD21-B cells in infection immunity and autoimmune pathology, and demonstrate that PD-1 is a critical checkpoint inhibitor for autoimmune disease development.

overall_design:
The bronchoalveolar lavage fluid (BALF) samples from five patients were mixed and FACS-sorted as the following strategy. Briefly, the CD45+ fraction was retrieved and followed by depletion of CD14+ and CD16+ cells. The fractions of CD3E+ and CD3E- of the remaining cells were isolated for single-cell RNA sequencing. Human V(D)J+5’ Gene Expression product from 10X GENOMICS was used to measure paired gene expression and immune repertoire information (paired TCR or BCR) within single cells. By using Gel Beads-in-emulsion code (GEM-Code) technology, transcripts within each cell were indexed with unique barcode. After reverse transcription, the full-length cDNA was amplified via PCR with primers against common 5’ and 3’ ends added during GEM-RT. With sufficient starting material, the double stranded cDNA library was split into two subsamples for the construction of Single Cell 5’ expression and TCR- or BCR-enriched library, separately. The Single Cell V(D)J and 5’ Gene Expression libraries were sequenced at certain depth and running parameters.

**The approval process for the data release is pending from the Chinese Government***

first: Bingtai

last: Lu

first: Jun

first: Zhanghua

last: Chen

first: Diyuan

last: Yang

first: Xiaohui

first: Yifan

last: Zhan

last: Cui

first: Andrew

middle: M

last: Lew

first: Liwei

first: Min

first: Gen

first: Fan

last: Bai


title: Transcriptome profiling of normal and DNMT3L-overexpressing human neurons [RNA-seq]

summary:
The objective of this study is to test the gene expression changes caused by DNMT3L overexpression in human neurons. The total RNA of each sample was extracted from the lentivirus infected, early differentiated human neuroprogenitors with ZsGreen (n=3) or DNMT3L (n=3) overexpression by using TRIzol reagent. Then the RNA samples were processed for high throughput transcriptome sequencing on Illumina HiSeq 3000 platform.
Results: among 57 905 cleaned RNAs, 7983 RNAs were differentially expressed in DNMT3L overexpressing neurons compared with control neurons, with q-value ≤ 0.05. After limited the cut-off by adding |fold change| ≥ 1.5, the numbers of differential expressed RNAs lowered to 1073, with more down-regulation  (590) than up-regulation (483). The differential expressed genes distributed in all chromosomes, also showing the pattern of more down-regulation than up-regulation in every chromosome except chromosome 21. Functional annotation with DAVID revealed the top functional groups including type I interferon signaling pathway and RNA and protein processing.

overall_design: Transcriptome profiling of 3 control (ZsGreen) and 3 experimental (DNMT3L) samples

first: Jie

first: Lin

last: Qin


title: Ovarian tissues of prenatally androgenized (PNA) mice

first: yani

last: Kang


title: Cell-type-specific profiling of loaded miRNAs from Caenorhabditis elegans reveals spatial and temporal flexibility in Argonaute loading [RNA-seq]

status: Public on Aug 25 2020

city: Brisbane

first: Steven

last: Zuryn

state: Queensland

address: 79 Upland Road

department: Queensland Brain Institute

country: Australia

institute: The University of Queensland

summary: We report the the 'translatome' of the nervous system in c.elegans by immunoprecipitating FLAG- epitope tagged RPL-18 expressed under a nervous system specific promoter. In addition we compared this to the nervous system specific 'translatome' in an alg-2 mutant background. As such we were able to determine the specifc effects of ALG-2 on the nervous sytem gene expression landscape.

overall_design: Cell-type spedific polysome complexes were immunoprecipitated using FLAG conjugated magnetic beads. Immunocomplexes were eluted from beads using the FLAG peptide and mRNA purified using TRIzol reagent (Invitrogen).

first: Christopher

middle: A

last: Brosnan

first: Alexander

middle: J

last: Palmer


title: Cell-type-specific profiling of loaded miRNAs from Caenorhabditis elegans reveals spatial and temporal flexibility in Argonaute loading


title: An unorthodox AR addiction drives enzalutamide resistance in prostate cancer

status: Public on Aug 20 2020

city: San Antonio

first: Zhenqing

last: Ye

state: TX

address: 8403 Floyd Curl Dr

department: 6Department of Population Health Sciences

institute: UT Health San Antonio

type: Genome binding/occupancy profiling by high throughput sequencing


title: Cell-type-specific profiling of loaded miRNAs from Caenorhabditis elegans reveals spatial and temporal flexibility in Argonaute loading [miRNA-seq]

summary: We aimed to characterize the cell-type specific loading patterns of miRNAs in c.elegans. We expressed cell-type specific HA-epitope tagged versions of Argonaute-like 1 (ALG-1) and ALG-2 from three major tissue types i.e. intestine, nervous system and body wall mucle. We found that most miRNAs display highly cell-type specific loading patterns. ALG-1 is more ubiquitously loaded whereas ALG-2 is eniriched for miRNA loading within the nervous system. Addtionally we show that there is flexibility in ALG loading which changes during the aging process.

overall_design: A total of 18 samples were anaysed (9 different stains of c.elegans each with biological replicated including; negative control, ALG-1 and ALG-2 from each of the 3 cell-types intestine, nervous system, body wall muscle and GABAergic neurons). Individual biological samples were immunopreciptiated and purified RNA extracted to assess cell-type specfic loading patterns.


title: RNA-sequencing data from Ctrl (Id2F/F or Id2F/FTcf7F/F), Ncr1Cre Id2F/F, Ncr1Cre Tcf7F/F and Ncr1Cre Id2F/F Tcf7F/F bone marrow NK cells.

status: Public on Aug 12 2020

city: Chicago

first: Barbara

last: Kee

state: Illinois

address: 924 E57th st

department: Pathology

institute: The University of Chicago

summary: The goal of this study was to determine how gene expression changes when Id2 and Tcf7 are deleted in bone marrow natural killer cells as compared to NK cells in which only Id2 or Tcf7 are deleted. We sorted NK1.1+CD49b+ cells from the bone marrow of mice with each of the indicated genotypes. Reads were aligned to the mm10 reference genome by Tophat2.1.0. Reads were assigned to genes using the htseq-count tool from HTSeq v 0.6.1 and gene annotations from Ensembl release 78. Differential expression was calculated across 3 independent replicates by EdgeR. We found that NK cells lacking Id2 had reduced expression of mature NK cell genes and increased expression of immature genes and known E protein target genes. Deletion of Tcf7 in ID2-deficient NK cells partially restored expression of maturation associated genes and decreased immature genes bringing their gene expression profiles closer to that of Ncr1Cre Tcf7F/F or Ctrl NK cells.

overall_design: Expression profiling analysis by RNA-sequencing of bone marrow NK1.1+CD49b+ natural killer cells in Ncr1Cre Id2F/F, Ncr1Cre Tcf7F/F or Ncr1Cre Id2F/F Tcf7F/F mice.  Id2F/F or Id2F/FTcf7F/F bone NK cell were used as controls . 3 independent replicates per sample.

first: Zhong-Yin

last: Li

first: Mikael

last: Sigvardsson

first: Elizabeth

last: Bartom

first: Mark

last: Maienshein-Cline


title: An unorthodox AR addiction drives enzalutamide resistance in prostate cancer (RNA-Seq)

summary: Acquisition of resistance to the next-generation anti-androgen enzalutamide (ENZ) constitutes a major challenge for the treatment of castrate-resistant prostate cancer (CRPC). Most ENZ-resistant (ENZ-R) CRPCs are androgen receptor (AR)-positive (CRPCAR+), but often negative for prostate-specific antigen (PSA), a well-established surrogate of AR activity, implying the emergence of AR-indifferent disease. Through genome-wide ChIP-seq and RNA-seq profiling in disease-relevant ENZ-R CRPCAR+ cells, we identified a unique set of gained AR binding sites (ARBS-G) lacking the canonical DNA binding elements of AR and the pioneer factor FOXA1. ARBS-G loci are highly enriched with CpG islands and significantly overlapped with the binding sites of the unmethylated CpG dinucleotide-binding protein CXXC5 and the partner TET2. Expression of CXXC5 and its downstream targets including inhibitor of differentiation-1 (ID1) was upregulated in ENZ-R CRPCAR+ cell lines, patient-derived xenografts (PDXs) and patient specimens. Genetic depletion of AR, CXXC5 or ID1 restored ENZ sensitivity in human ENZ-R CRPCAR+ cells. Most importantly, ENZ-R CRPCAR+ cells, organoids, xenografts and PDXs were invariably hypersensitive to the novel BET-CBP/p300 dual inhibitor NEO2734. These results reveal that ENZ-R CRPCAR+ cancers are only indifferent to the canonical AR (cAR) activity, but remain addictive to the noncanonical AR (ncAR) function which is selectively vulnerable upon dual inhibition of CBP/p300 and BET family proteins.

overall_design: Examination of RNA expression in: long-term (> two months) treatment with enzalutamide (ENZ) resistant C4-2-ENZ cells and control C4-2-CON cells.

first: Haojie

last: Huang

first: Yundong

last: He


title: Single cell RNA-seq analysis of pMN neural progenitors from zebrafish

status: Public on Aug 11 2020

city: Aurora

first: Bruce

last: Appel

state: CO

institute: University of Colorado School of Medicine

summary: The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so, we sorted fluorescently marked pMN cells obtained from Tg(olig2:EGFP) zebrafish embryos at 24, 36 and 48 hours post fertilization and performed 10X Chromium single cell RNA-seq.

overall_design: Three samples obtained from wild-type zebrafish embryos at three developmental timepoints were analyzed.

first: Christina

last: Kearns

first: Austin

last: Gillen

first: Rebecca

last: O'Rourke


title: Effects of LMTK2 knockdown on total RNA expression in CFBE41o- cells after TGFβ1 or vehicle control (HCl+BSA) treatment

city: Pittsburgh

first: Donald

middle: B.

last: DeFranco

state: PA

address: 7041 Biomedical Sciences Tower 3

department: Department of Pharmacology and Chemical Biology

institute: University of Pittsburgh

summary: The purpose of this study was to increase the current knowledge about the LMTK2 network and its intersection with the TGF-β1 signaling pathway. To understand how the LMTK2 and TGF-β1 pathways interconnect, we knocked down LMTK2 using small(si)RNA-mediated silencing in human bronchial epithelial CFBE41o- cells, treated cells with TGF-β1 or vehicle control, and performed differential gene expression analysis by RNA sequencing (RNAseq).

overall_design: Differential gene expression analysis by RNA sequencing (RNAseq) in parental CFBE41o- cells after LMTK2 knockdown and 24hour treatment with TGFβ1 or vehicle control (HCl+BSA)

first: Daniel

last: Cruz

first: Nilay

last: Mitash

first: Agnieszka

last: Swiatecka-Urban


title: Epigenome analysis of whole-blood samples of septic and healthy preterm neonates

city: Paterna

first: María José

last: Garzón Garzón

address: Carrer del Catedràtic Agustín Escardino Benlloch, 9

country: Spain

institute: EpiDisease SL

summary: Neonatal sepsis is an important health-care concern worldwide occurring more frequently in premature newborns and for which diagnosis is yet a challenge, causing delays in therapy and increasing the risk of death. DNA methylation, involved in regulating gene expression, has been associated with the development and progression of sepsis. Actually, the detection of differentially methylated regions (DMRs) is a promising epigenetic tool used for diagnosis and prognosis in complex diseases. The present study focuses on two different bioinformatic methods, DMRcate and mCSEA, with the aim of obtaining different methylation traits using Illumina Infinium Human Methylation EPIC data of neonatal sepsis patients. The DMR sets obtained by both approaches can also be overlapped to obtain a reliable set of DMRs which can contribute to improve our understanding on the molecular pathways underlying disease.

overall_design: Blood samples from septic (n=17) and healthy (n=6) preterm infants were hybridized by the Illumina Infinium EPIC Human Methylation Beadchip, following DNA extraction and bisulphite treatment.

first: Sheila

last: Lorente-Pozo

first: Paula

last: Navarrete

first: María

last: Garzón

first: Inmaculada

last: Lara-Cantón

first: Jesús

last: Beltrán-García

first: Rebeca

last: Osca-Verdegal

first: Salvador

last: Mena-Mollá

first: Eva

last: García-López

first: Máximo

last: Vento

first: Federico

middle: V

last: Pallardó

first: José Luis

last: García-Giménez


title: Analysis of differentially expressed long noncoding RNAs, circular RNAs and the associated TF‐mRNA network in tongue squamous cell carcinoma

status: Public on Aug 04 2020

city: Fuzhou

first: mi

last: zhang

address: Jiaotong Road number 88

institute: Fujian Medical University

summary:
Purpose: The purpose of this study was to identify aberrantly expressed lncRNAs, circular RNAs and the associated TF‐mRNA network in TSCC.
Methods: Tongue tumor noncoding and mRNA profiles of tongue squamous cell carcinoma and adjacent noncancerous tissues were generated by deep sequencing, using Illumina HiSeq 4000.
Results: Using an optimized data analysis workflow.
Conclusions: we found a profile of dysregulated lncRNAs, TFs and mRNAs that could serve as prospective clinical biomarkers because of their tissue specificity and association with the tumorigenesis and progression of TSCC.

overall_design: tongue tumor noncoding and mRNA profiles of tongue squamous cell carcinoma and adjacent noncancerous tissues were generated by deep sequencing, in triplicate, using Illumina HiSeq 4000

first: Mi

last: Zhang

first: Zexi

last: Chen


title: An unorthodox AR addiction drives enzalutamide resistance in prostate cancer (ChIP-Seq)

first: Zhenqing

last: Ye

overall_design: To determine the molecular mechanism underlying AR growth of ENZ-R cells, we performed chromatin immunoprecipitation sequencing (ChIP-seq) for AR, FOXA1 and histone H3 lysine 27 (H3K27ac), a histone mark of both active enhancer and transcription start site (TSS) regions

first: Haojie

last: Huang

first: Yundong

last: He


title: Lipid remodeling in response to methionine stress in MDA-MBA-468 triple-negative breast cancer cells

city: Irvine

first: Peter

last: Kaiser

state: CA

address: Department of Biological Chemistry, 829 Health Sciences Rd

institute: University of California, Irvine

summary: Methionine is an essential amino acid and critical precursor to the cellular methyl donor S-adenosylmethionine. Unlike non-transformed cells, cancer cells have a unique metabolic requirement for methionine and are unable to proliferate in growth media where methionine is replaced with its metabolic precursor, homocysteine. This metabolic vulnerability is common among cancer cells regardless of tissue origin and is known as “methionine dependence”, “methionine stress sensitivity”, or Hoffman effect. Here, we characterize the response of lipids to methionine stress in the triple negative breast cancer cell line MDA-MB-468 and its methionine stress insensitive derivative, MDA-MB-468res-R8. Lipidome analysis identified an immediate, global decrease in lipid abundances with the exception of triglycerides and an increase in lipid droplets in response to methionine stress specifically in MDA-MB468 cells. Furthermore, specific gene expression changes were observed as a secondary response to methionine stress in MDA-MB-468, resulting in a down-regulation of fatty acid metabolic genes and up-regulation of genes in the unfolded protein response pathway. We conclude that the extensive changes in lipid abundance during methionine stress is a direct consequence of the modified metabolic profile previously described in methionine stress sensitive cells. The changes in lipid abundance likely results in changes in membrane composition inducing the unfolded protein response we observe.

overall_design: RNA profiles of the triple-negative, methionine-sensitive breast cancer line MDA-MB-468 and its methionine-insensitive derivitive cell line MDA-MB-468res-R8 during methionine stress.

first: Stacey

last: Borrego


title: Mutant huntingtin stalls ribosomes and represses protein synthesis in a cellular model of Huntington disease.

status: Public on Jul 22 2020

city: Jupiter

first: Srinivasa

last: Subramaniam

state: FL

address: 130 Scripps Way

institute: The Scripps Research Institute


title: Primate innate immune responses to bacterial and viral pathogens reveals an evolutionary trade-off between strength and specificity

status: Public on Aug 09 2020

first: Luis

last: Barreiro

state: IL

address: 900 East 57th Street

department: Genetic Medicine

institute: Chicago University

summary: We report the whole genome transcriptomic responses of ape species (human, common chimpanzee) and AAMs (rhesus macaque and olive baboon) to bacterial and viral stimulation. We find stark differences in the responsiveness of these groups, with apes mounting a markedly stronger early transcriptional response to both viral and bacterial stimulation, altering the transcription of ∼40% more genes than AAMs. Additionally, we find that genes involved in the regulation of inflammatory and interferon responses show the most divergent early transcriptional responses across primates and that this divergence is attenuated over time. Finally, we find that relative to AAMs, apes engage a much less specific immune response to different classes of pathogens during the early hours of infection, upregulating genes typical of anti-viral and anti-bacterial responses regardless of the nature of the stimulus

sample_organism:
Macaca mulatta
Papio anubis
Pan troglodytes
Homo sapiens

overall_design: Whole blood stimulation of two stimulants that mimic bacterial and viral infections of whole blood obtained from 4 primate species, human, chimp, macaque and baboon.

first: Mohamed

last: Fahmy Hawash

first: Joaquin

last: Sanz-Remón

first: Jean-Christophe

last: Grenier

first: Jordan

last: Kohn

first: Vania

last: Yotova

first: Zach

last: Johnson

first: Robert

last: Lanford

first: Jessica

middle: F

last: Brinkworth


title: Sonic hedgehog accelerates DNA replication to cause replication stress promoting cancer initiation in medulloblastoma (RNA-Seq)

status: Public on Jul 20 2020

city: Montreal

first: Frederic

last: Charron

state: Quebec

address: 110 Avenue des Pins Ouest

country: Canada

institute: IRCM

summary: Cancer is a multi-stage disease caused by sequential mutations; however, the molecular mechanism generating cancer-initiating mutations for many cancers is not well understood. Using the developing cerebellum as model system, here we delineate a molecular mechanism for tumor initiation in medulloblastoma (MB), the most frequent malignant pediatric brain tumor. Activation of the Sonic hedgehog (SHH) pathway is frequent in MB, with mutations in the tumor suppressor PTCH1 (a negative regulator of SHH signaling) being the most common initiating event for SHH-MB. However, how SHH as a developmental mitogen promotes early carcinogenesis in the cells of origin, granule cerebellar progenitors (GCPs), remains to be determined.       Here we report that physiological exposure of GCPs to Shh causes a distinct form of DNA replication stress, altering DNA replication dynamics to increase both origin firing and fork velocity. Shh promotes DNA helicase loading and activation in GCPs, with increased levels of Cdc7-dependent replication origin firing. S-phase duration is reduced and hyper-recombination consequently occurs. Such elevated recombination causes copy-number neutral LOH, an event seen frequently at the PTCH1/ptch1 locus. Moreover, Cdc7 inhibition to attenuate origin firing in a MB mouse model is sufficient to reduce somatic recombination and preneoplastic tumor formation. We therefore establish that tissue-specific replication stress induced by Shh acts to promote LOH, which in tumor-prone Ptch1+/- GCPs can result in loss of this tumor suppressor, as an early cancer initiating event.

overall_design:
RNA extraction followed by high throughput sequencing (RNAseq)
RNAseq in GCPs treated with Control, Shh or Igf1

first: Lukas

last: Tamayo-Orrego

first: Amandine

last: Bemmo


title: Mutant huntingtin stalls ribosomes and represses protein synthesis in a cellular model of Huntington disease [RNA-seq]

summary: The polyglutamine expansion of huntingtin (mHtt) causes Huntington disease (HD) and neurodegeneration, but the mechanisms remain unclear. Here, we found that mHtt promotes ribosome stalling and suppresses protein synthesis. Depletion of mHtt enhances protein synthesis and increases the speed of ribosome translocation, while mHtt directly inhibits protein synthesis in vitro. We found interactions of ribosomal proteins and translating ribosomes with mHtt. High-resolution global ribosome footprint profiling (Ribo-Seq) and mRNA-Seq indicated a widespread shift in ribosome occupancy toward the 5’ and 3’ end and unique single-codon pauses on selected mRNA targets in HD cells, compared to controls. Thus, mHtt impedes ribosomal translocation during translation by promoting ribosome stalling, a novel mechanistic defect that can be exploited for HD therapeutics.

overall_design: RNA-seq: Control, HD-heterozygous, and HD-homozygous striatal cell line.

first: Mehdi

last: Eshraghi

first: Pabalu

last: Karunadharma

first: Juliana

last: Blin

first: Emiliano

last: Ricci

first: Audrey

last: Michel


title: Mutant huntingtin stalls ribosomes and represses protein synthesis in a cellular model of Huntington disease [Ribo-seq]

first: Srinivasa

last: Subramaniam

overall_design: For ribosome footprinting, three independent rounds of cultured stratial cells (i.e. control, HD-heterozygous, and HD-homozygous cells) were used. To resolve ribosome-associated footprints, the lysates were treated with RNase T1 and RNase A for 30 min, separated on 10-50% sucrose gradients and RNA from the 80S monosome fractions were purified.


title: Sonic hedgehog accelerates DNA replication to cause replication stress promoting cancer initiation in medulloblastoma (EdU-Seq)

overall_design: EdU-seq assay followed by high throughput sequencing


title: Sonic hedgehog accelerates DNA replication to cause replication stress promoting cancer initiation in medulloblastoma

overall_design: Refer to individual Series


status: Public on Jul 13 2020

city: Los Angeles

first: Bob

last: Goldberg

address: 610 Charles E Young Drive East

department: Molecular, Cell and Developmental Biology

institute: University of California, Los Angeles

summary: To dissect the gene regulatory networks operating during Scarlet Runner Bean seed development, we identified the binding sites genome-wide for transcription factor in Scarlet Runner Bean seeds during seed  development using ChIP-seq

overall_design: Identification of Scarlet Runner Bean WOX9-like binding sites in Scarlet Runner Bean seeds during seed development. ChIP and total chromatin controls (input) were obtained for two replicates. Samples were sequenced using the Illumina HiSeq2500 and NovaSeq 6000 platform.

first: Nestor

last: Apuya

first: Min

last: Chen


title: Embryo-derived chemokine CXCL12 enhances the pregnancy outcome via improved endometrial receptivity

status: Public on Jul 09 2020

city: SEONGNAM-SI

first: YOUN JUNG

last: KANG

address: 335 PANGYO-RO

department: MEDICINE

country: South Korea

institute: CHA University

summary: Intrauterine infusion of CXCL12 has a beneficial effect on endometrial angiogenesis by recontruction of endometrial vasculature and improvement of endometrial receptivity

overall_design: As a non-invasive treatment strategy, CXCL12 was infused into one side of mouse uterine horns and saline was infused into the other side of horns for the control (N=28). Assessment of CXCL12 impact on the endometrium at day3 and day8.

first: Youn-Jung

last: Kang

first: Hwa

middle: S

last: Koo

first: Min-Ji

last: Yoon

first: Chang-Jin

last: Lee

first: Seon-Hwa

last: Hong

first: Hwijae

last: Cha

first: Jungho

last: Ahn

first: Danbi


title: Musashi1 contributes to glioblastoma development via regulation of a network of DNA replication, cell cycle, and division genes

status: Public on Jul 08 2020

first: David

last: Kadosh

state: Texas

address: 7703 Floyd Curl Dr., MC: 7758

department: Department of Microbiology, Immunology & Molecular Genetics

institute: University of Texas Health Science Center at San Antonio

summary: RNA binding proteins (RBPs) function as master regulators of gene expression. Alterations in their expression levels and function are often observed in tumors with numerous oncogenic RBPs identified in recent years.  Musashi1 (Msi1) is an RBP and stem cell marker gene that controls the balance between self-renewal and proliferation. High Msi1 levels have been observed in multiple tumor types including glioblastoma and are often associated with poor patient outcomes. Msi1 knockdown is known to affect numerous cancer phenotypes and to prevent growth of different tumor types. To dissect Msi1’s main contributions to glioblastoma development, we conducted a comprehensive genomic analysis. We identified a network of cell cycle/division and DNA replication genes and established these processes as its core functions. Msi1 controls this gene network via two mechanisms: direct interaction and indirect regulation mediated by the transcription factors E2F2 and E2F8. We corroborated Msi1’s impact on these processes and determined that glioblastoma lines with Msi1 KO display increased sensitivity to cell cycle and DNA replication inhibitors. As Msi1 inhibitors have been identified, a drug combination strategy (Msi1 + cell cycle/DNA replication inhibitors) could be a viable route to treat glioblastoma.

overall_design: U251 and U343 control and MSI1 KO cells were plated in 96 wells at a density of 8x104. Cells were synchronized by serum starvation for 24 hours. Following synchronization, cells were exposed to 20 nM of Paclitaxel for 24 hours. Cells were fixed in 4% PFA and stained with Alpha-Tubulin and DAPI. The experiment was performed in triplicates and 1,000 cells were counted per replicate. Differences were assessed with a student’s t-test. p < 0.01.

first: Luiz

middle: O

last: Penalva

first: Saket

last: Choudhary


title: Drosophila Exhibit Divergent Sex-Based Responses in Transcription and Motor Function After Traumatic Brain Injury

status: Public on Jul 06 2020

city: Detroit

first: Ekta

last: Shah

state: Michigan

address: 275 E.  Hancock

institute: Wayne State University

summary: Adult male and female w1118 flies were inflicted with traumatic brain injury using high-impact trauma device. Brains were dissected at control, 1, 2 and 4 hours post-TBI. Total RNA was extracted from fly brains and 3'mRNA-seq was performed to assess gene expression changes in response to TBI in both sexes. 3 replicates were sequenced for each condition.

overall_design: To investigate sex-differences in gene expression at control, 1, 2 and 4-hours post-TBI in Drosophila


title: A single-cell transcriptomics CRISPR-activation screen identifies new epigenetic regulators of the zygotic genome activation programme (10X Genomics 3' scRNA-seq of MERVL LTR and Zscan4 CRISPRa dataset)

status: Public on Jul 05 2020

city: Cambridge

first: Felix

last: Krueger

address: Granta Park

department: Bioinformatics

institute: Altos Labs

summary: Zygotic genome activation (ZGA) is a crucial developmental milestone that remains poorly understood. This first essential transcriptional event in embryonic development coincides with extensive epigenetic reprogramming and is orchestrated, in part, by the interplay of transcriptional and epigenetic regulators. Here, we developed a novel high-throughput screening method that combines pooled CRISPR-activation (CRISPRa) with single-cell transcriptomics and applied this method to systematically probe candidate regulators of ZGA-like transcription. We screened 230 epigenetic and transcriptional factors by upregulating their expression with CRISPRa in mouse embryonic stem cells (ESCs). Through single-cell RNA-sequencing (scRNA-seq), we generated approximately 200,000 single-cell transcriptomes of CRISPRa-perturbed cells, each transduced with a unique short-guide RNA (sgRNA) targeting a specific candidate gene promoter. Using multi-omics factor analysis (MOFA) of the perturbation scRNA-seq profiles, we characterized molecular signatures of ZGA and uncovered 24 factors that promote a ZGA-like response in ESCs, both in the coding and non-coding transcriptome. We further validated nine candidate genes by arrayed CRISPRa analysed by bulk transcriptomics, which demonstrates that the combination of CRISPRa with scRNA-seq is a powerful and valid approach to identify regulators of ZGA-like transcription. Additional cDNA overexpression assays for three top hits, the DNA binding protein Dppa2, the chromatin remodeller Smarca5 and the transcription factor Patz1, confirmed these factors as ZGA-like regulators by alternative methods. Supporting these findings, Dppa2 and Smarca5 knock-out ESCs lose expression of ZGA genes and functional experiments revealed that Smarca5’s regulation of ZGA-like transcription is dependent on Dppa2. Together, our single-cell transcriptomic profiling of CRISPRa-perturbed cells provides comprehensive system-level insights into the molecular mechanisms that orchestrate ZGA.

overall_design: SAM mESCs were transduced with short-guide RNAs (sgRNAs) targeting either the murine endogenous retrovirus with leucine tRNA primer (MERVL) long terminal repeats (LTRs) or zinc finger and SCAN domain containing 4 (Zscan4) gene cluster promoters, and single cell transcriptomes were captured using the 10X Genomics 3’ polyA primed scRNA-seq platform

first: Celia

last: Alda-Catalinas

first: Danila

last: Bredikhin

first: Irene

last: Hernando-Herraez

first: Fátima

last: Santos

first: Oana

last: Kubinyecz

first: Mélanie

last: Eckersley-Maslin

first: Oliver

last: Stegle

first: Wolf

last: Reik


title: A single-cell transcriptomics CRISPR-activation screen identifies new epigenetic regulators of zygotic genome activation

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


title: An early cell shape transition drives evolutionary expansion of human forebrain

first: Paula

last: Freire Pritchett

address: MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge Biomedical Campus

institute: Medical Research Council

summary: The human brain has undergone rapid expansion since humans diverged from other great apes, but the mechanism of this human-specific enlargement is still unknown. Here, we use cerebral organoids derived from human, gorilla and chimpanzee cells to study developmental mechanisms driving evolutionary brain expansion. We find that the differentiation of neuroepithelial cells to neurogenic radial glia is a protracted process in apes, involving a previously unrecognized transition state characterized by a change in cell shape. Furthermore, we show that human organoids are larger due to a delay in this transition. Temporally resolved RNA-seq from human and gorilla organoids reveals differences in gene expression patterns associated with cell morphogenesis, and in particular highlight ZEB2, a known regulator of epithelial-mesenchymal transition and cell shape. We show, through loss- and gain-of-function experiments, that ZEB2 promotes the progression of neuroepithelial differentiation, and its ectopic overexpression in human is sufficient to trigger a premature transition. By mimicking the nonhuman ape expression of this human evolution gene, we are able to force the acquisition of nonhuman ape architecture in human organoids, establishing for the first time, an instructive role of neuroepithelial cell shape in human brain expansion.

sample_organism:
Gorilla gorilla
Homo sapiens

platform_organism:
Gorilla gorilla
Homo sapiens

overall_design: Organoids generated from 3 replicate batches of H9 and G1 cell lines were collected at 7 time points for bulk RNA-seq analysis: 0, 2, 3, 5, 10, 15 and 25 days post neural induction. ~300 organoids per replicate were collected at the time points ranging between day 0 and 5, ~150 organoids at day 10, ~100 organoids at day 15, and ~50 organoids at day 25. RNA was isolated using Direct-zol-96 RNA kit. cDNA libraries were generated according to NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (NEB #E7760) and barcoded with NEBNext Multiplex Oligos for Illumina. All 42 bulk RNA samples were pooled and sequenced with single-end 50 base mode using 3 lanes of an Illumina HiSeq 4000. Human and gorilla reads were aligned to GRCh38 and Kamilah_GGO_v0 reference genomes respectively. GTF gene annotation files for both reference genomes were filtered down to 16,763 genes that are annotated with identical gene names in both species. Read counts were normalized to transcripts per million (TPM) within each sample and can be found in the supplementary file, 'hum_gor_combined_tpms_HISAT2.txt'.

first: Silvia

last: Benito-Kwiecinski

last: Freire-Pritchett

first: Madeline

last: Lancaster


title: Transcriptional profiling of CHB8 human embryonic stem cells with insulin receptor knockdown

status: Public on Aug 01 2020

first: William

last: Mallard

state: MA

address: 7 Divinity Ave

department: Stem Cell and Regenerative Biology

institute: Harvard University

summary: While insulin receptor (IR) signaling is involved in the maintenance and proliferation of human embryonic stem cells (hESCs), its functional significance is poorly understood. To elucidate the role of IR in the pluripotency network, we knocked down INSR in CHB8 hESCs and performed RNA sequencing and differential expression analysis. We observe dysregulation of the AKT and ERK1/2 signalling pathways and downregulation of extracellular matrix genes.

overall_design: RNA sequencing of CHB8 hESCs with shRNA knockdown of INSR.

first: Adrian

middle: K

last: Teo

first: Manoj

last: Gupta

first: John

last: Rinn

first: Marina

last: Gritsenko

first: Richard

last: Smith

first: Wei-Jun

last: Qian

first: Rohit

middle: N

last: Kulkarni


title: A single-cell transcriptomics CRISPR-activation screen identifies new epigenetic regulators of the zygotic genome activation programm (10X Genomics CRISPRa screen dataset)

first: Felix

last: Krueger

overall_design: SAM mESCs were transduced with a lentiviral library of 475 short-guide RNAs (sgRNAs) targeting maternal epigenetic factors, in triplicates at a <0.1 multiplicity-of-infection (MOI) , and following selection and expansion of the pool of transduced cells, single-cell transcriptomes were generated using 10X 3’ scRNA-seq and their corresponding sgRNAs further amplified with a specific amplification protocol (Hill et al. 2018). Each transduction replicate was loaded across a full 10X chip Chromium Controller (8 lanes), with 20,000 cells per lane. Samples names are labelled with the replicate number 1-3 and 10X lane 1-8 (e.g: Replicate1_1 refers to transduction replicate 1 run in 10X lane 1). Amplicon sgRNA libraries were performed for each full length 10X cDNA sample (8 per replicate) and are referred to with the replicate number 1-3 and 10X lane 1-8 (e.g. amplicon_sgRNA_1_1 refers to the amplicon sgRNA libraries made on the 10X full length cDNA of sample1_1).

first: Celia

last: Alda-Catalinas

first: Danila

last: Bredikhin

first: Irene

last: Hernando-Herraez

last: Santos


title: A single-cell transcriptomics CRISPR-activation screen identifies new epigenetic regulators of the zygotic genome activation programme (bulk RNA-seq of E14 and SAM mESCs dataset)

overall_design: Bulk RNA-seq of three replicates of E14 mouse embryonic stem cells (mESCs) and two replicates of SAM mESCs. SAM mESCs are a clonal cell line generated from E14s by lentiviral integration of dCas9-VP64 and MS2-p65-HSF1

first: Fátima

first: Oana

last: Kubinyecz

first: Mélanie

last: Eckersley-Maslin

first: Oliver

last: Stegle

last: Reik


title: Ependymoma subpopulation lineages underlie clinical classification and outcome (scRNAseq data set)

status: Public on Jul 31 2020

first: Andrew

last: Donson

department: Pediatrics

institute: CU Anschutz Medical Campus

summary: Ependymoma (EPN) is a brain tumor commonly presenting in childhood that remains fatal in the majority of children. Intra-tumoral cellular heterogeneity in bulk tumor samples significantly confounds our understanding of EPN biology, impeding development of effective therapy. We used single-cell RNA sequencing to identify four neoplastic cell subpopulations in posterior fossa EPN patient samples that underlie traditional subgroup classification and harbor divergent lineage trajectories and clinical outcomes. Neoplastic subpopulations expressed gene signatures associated with normal ependyma, ependymoma, and mesenchymal phenotypes, consequently named ciliated (CEC), transportive (TEC), undifferentiated (UEC), and mesenchymal ependymoma cells (MEC). The abundance of EPN subpopulations as estimated by deconvolution of EPN bulk tumor transcriptomes showed that the ratio of MEC and CEC cell fractions dictated assignment to the two main classification subgroups in PFA. Longitudinal analysis revealed evolution of subpopulation fractions, notably MEC and CEC, which changed between presentation and recurrence. Outcome analyses demonstrated that a higher proportion of UEC or MEC subpopulations was associated with shorter survival, whereas CEC was associated with longer survival. Unsupervised pseudotime analysis revealed divergent subpopulation lineages. The first lineage conformed to a classic cancer stem cell trajectory where UEC progenitors differentiate sequentially to TECs and then CECs. In the second trajectory MECs arise from UECs via TECs, potentially in response to hypoxia-mediated cellular stress, a process that we recreated in vitro in a hypoxia treated EPN cell line. Our study reveals the existence of significant, unappreciated cellular heterogeneity in EPN and provides important resolution of EPN tumor biology.

overall_design: Single-cell transcriptional profiles (10X Chromium) were generated from ten EPN-PF classified as PFA1 (n=6) and PFA2 (n=4) based on bulk tumor methylomic and transcriptomic profiling.

first: Austin

last: Gillen

first: Keny

last: Riemondy

first: Ahmed

last: Gilani

first: Sujatha

last: Venkataraman

first: Krishna

last: Madhavan

first: Eric

last: Prince

first: Bridget

last: Sanford

first: Vladimir

last: Amani

first: Andrea

last: Griesinger

first: Todd

last: Hankinson

first: Michael

last: Handler

first: Rajeev

last: Vibhakar

first: Ken

last: Jones

first: Siddhartha

last: Mitra

first: Jay

middle: R

last: Hesselberth

first: Nicholas

last: Foreman


title: A single-cell transcriptomics CRISPR-activation screen identifies new epigenetic regulators of the zygotic genome activation progrmme (bulk RNA-seq of arrayed CRISPRa validations and bulk RNA-seq of Patz1, Dppa2, Smarca5 and Carhsp1 cDNA overexpression)

first: Felix

last: Krueger

summary: Zygotic genome activation (ZGA) is a crucial developmental milestone that remains poorly understood. This first essential transcriptional event in embryonic development coincides with extensive epigenetic reprogramming and is orchestrated, in part, by the interplay of transcriptional and epigenetic regulators. Here, we developed a novel high-throughput screening method that combines pooled CRISPR-activation (CRISPRa) with single-cell transcriptomics and applied this method to systematically probe candidate regulators of ZGA-like transcription. We screened 230 epigenetic and transcriptional factors by upregulating their expression with CRISPRa in mouse embryonic stem cells (ESCs). Through single-cell RNA-sequencing (scRNA-seq), we generated approximately 200,000 single-cell transcriptomes of CRISPRa-perturbed cells, each transduced with a unique short-guide RNA (sgRNA) targeting a specific candidate gene promoter. Using multi-omics factor analysis (MOFA) of the perturbation scRNA-seq profiles, we characterized molecular signatures of ZGA and uncovered 24 factors that promote a ZGA-like response in ESCs, both in the coding and non-coding transcriptome. We further validated nine candidate genes by arrayed CRISPRa analysed by bulk transcriptomics, which demonstrates that the combination of CRISPRa with scRNA-seq is a powerful and valid approach to identify regulators of ZGA-like transcription. Additional cDNA overexpression assays for three top hits, the DNA binding protein Dppa2, the chromatin remodeller Smarca5 and the transcription factor Patz1, confirmed these factors as ZGA-like regulators by alternative methods. Supporting these findings, Dppa2 and Smarca5 knock-out ESCs lose expression of ZGA genes and functional experiments revealed that Smarca5?s regulation of ZGA-like transcription is dependent on Dppa2. Together, our single-cell transcriptomic profiling of CRISPRa-perturbed cells provides comprehensive system-level insights into the molecular mechanisms that orchestrate ZGA.

overall_design: Bulk RNA-seq of SAM mESCs after CRISPRa of Patz1, Dppa2, Dppa4, Smarca5, Pou2f2, Tsc22d4, Arnt, Sirt1, Smad1 and Carhsp1; bulk RNA-seq of E14 mESCs after 48h transfection with Dppa2, Smarca5, Patz1 and Carhsp1 GFP fusion constructs and FACS sorting of GFP+ cells. Each experiment was done in biological replicates.

first: Celia

last: Alda-Catalinas

first: Danila

last: Bredikhin

first: Irene

last: Hernando-Herraez

first: Fátima

last: Santos

first: Oana

last: Kubinyecz

first: MÃ©lanie

last: Eckersley-Maslin

first: Oliver

last: Stegle

first: Wolf

last: Reik


title: Gene expression profiles in microglia with or without PGC-1α overexpression after acute ischemic stroke (AIS)

status: Public on Jun 20 2020

city: Tianjin

first: Bin

last: Han

address: 154 Anshan Road, Heping District

institute: Tianjin Medical University General Hospital

summary: PGC-1α overexpression in microglia protects against ischemia-induced brain damage in mice. To investigate the underlying mechanism of PGC-1α, we have employed whole mRNA microarray expression profiling as a discovery platform to identify genes with the potential to change the microglial function. Indeed, PGC-1α overexpression alters the gene expression profiles of microglia after AIS, this suggested that microglial PGC-1α might play an important role after ischemic stroke.

overall_design: Primary microglia were isolated from PGC-1αf/f and mPGC-1α mice with AIS using the MojoSortTM mouse P2RY12 selection kit (Biolegend, San Diego, CA, USA) according to the manufacturer’s protocol. Then the samples were processed to perform microarray analysis.

first: Junwei

last: Hao


title: Transcriptomic profiling of the white adipose tissue (WAT)  in ApoE3L.CETP mice fed a high fat diet (HFD) or a low fat diet (LFD) for three different time periods, or chow diet at baseline

status: Public on Jul 14 2020

city: New York City

first: Dimitris

last: Nasias

state: New York

address: 550 1st Avenue

department: Langone Medical School

institute: New York University

summary: The metabolic syndrome (MetS) is characterized by the presence of metabolic abnormalities that include abdominal obesity, dyslipidemia, hypertension, increased blood glucose/insulin resistance, hypertriglyceridemia and increased risk for cardiovascular disease (CVD). The ApoE*3Leiden.human Cholesteryl Ester Transfer Protein (ApoE3L.CETP) mouse model manifests several features of the MetS upon high fat diet (HFD) feeding. Moreover, the physiological changes in the white adipose tissue (WAT) contribute to MetS comorbidities. The aim of this study was to identify transcriptomic signatures in the gonadal WAT of ApoE3L.CETP mice in discrete stages of diet-induced MetS.

overall_design: For this purpose, 12 weeks old male ApoE3L.CETP mice fed for 4, 8 or 12 weeks a low fat diet (LFD) (D12450B, Research diets, New Brunswick, USA) as control groups or a high fat diet (HFD) (D14010701, Research diets, New Brunswick, USA) to induce the MetS. The clinical profile of mice was thoroughly assessed before and after diet administration and has been previously described (doi: 10.1002/jcp.28649). Mice were sacrificed at the corresponding time points and the gonadal white adipose tissues were collected to assess global gene expression changes. Baseline expression was assessed in mice 12 weeks of age that were fed with a chow diet. Total RNA from all mice (LFD or HFD group at 4, 8 and 12 weeks, 3-4 replicates/ Baseline group, 2 replicates) was labeled and hybridized to GeneChip™ Mouse Gene 2.0 ST Affymetrix expression arrays. The total of 25 arrays was analyzed to the Affymetrix Transcriptome Analysis Console (TAC) Software Version 4.0.2. To determine the differentially expressed genes, we applied the Two Way Analysis of Variance specifying two conditions (diet and time) under specific statistical criteria (p-value <0.05 and absolute fold change ≥1.5).

last: Kardassis


title: Targeted protein degradation reveals non-catalytic function of Aurora A kinase

status: Public on Jul 17 2020

city: Kiel

first: Elmar

last: Wolf

state: Schleswig-Holstein

address: Rudolf-Höber-Str. 1

department: Biochemical Institute

institute: Christian-Albrechts-University Kiel

summary: RNA Seq in MV4-11 cells after treatment with Alisertib or JB170

first: Bikash

last: Adhikari

first: Jelena

last: Bozilovic

first: Apoorva

last: Baluapuri

first: Stefan

last: Knapp


title: AtDXO1 possesses deNADding and exonuclease activities and plays pleiotropic roles in gene regulation

status: Public on Jun 16 2020

city: HONG KONG

first: Huan

last: ZHONG

state: HONG KONG

address: T1012 CHA CHI-MING SCIENCE TOWER

department: BIOLOGY

country: Hong Kong

institute: HKBU

summary: RNA capping and decapping tightly coordinate with transcription, translation, and RNA decay to regulate gene expression. The DXO/Rai1 family of proteins have been implicated in mRNA decapping and decay, and mammalian DXO was recently found to also function as a decapping enzyme for NAD+-capped RNAs (NAD-RNA). The Arabidopsis genome contains a single gene encoding a DXO/Rai1 protein, AtDXO1. Here we show that AtDXO1 possesses both NAD-RNA decapping activity and 5’-3’ exonuclease activity and that the atdxo1 mutation increases the stability of NAD-RNAs. The atdxo1 mutant displays severe growth retardation, pale color, and multiple developmental defects. Transcriptome profiling analysis showed that the atdxo1 mutation leads to elevated expression of defense-related genes and down-regulation of photosynthesis genes. The autoimmunity phenotype of the mutant could be suppressed by either eds1 or npr1 mutation. Surprisingly, the various phenotypes associated with the atdxo1 mutant could be complemented by the enzymatically inactive AtDXO1. We found that AtDXO1 interacts with RNA cap methyltransferase (CMT). The atdxo1 mutation apparently enhances post-transcriptional gene silencing by elevating levels of siRNAs. Our study indicates that AtDXO1 regulates gene expression in various biological and physiological processes through its pleiotropic molecular functions in mediating RNA processing and decay.

overall_design: Transcriptome and global small RNA analysis of atdxo1 mutants

first: Shuying

last: PAN

first: Kaien

last: LI

first: Huihui

last: Wu

first: He

last: ZHANG

first: Yiji

last: XIA


title: Suppression of adenosine-to-inosine (A-to-I) RNA editome by death associated protein 3 (DAP3) promotes cancer progression

status: Public on Jun 23 2020

city: Singapore

first: Polly Leilei

last: Chen

address: #12-01, MD6, 14 Medical Drive

department: Department of Anatomy

institute: Cancer Science Institute of Singapore NUS

summary: RNA editing introduces nucleotide changes in RNA sequences. Recent studies have reported that aberrant A-to-I RNA editing profiles are implicated in cancers. Albeit changes in expression and activity of ADAR genes are thought to have been responsible for the dysregulated RNA editome in diseases, they are not always correlated, indicating the involvement of secondary regulators. Here, we uncover DAP3 as a potent repressor of editing and a strong oncogene in cancer. DAP3 mainly interacts with the deaminase domain of ADAR2 and represses editing via disrupting association of ADAR2 with its target transcripts. PDZD7, an exemplary DAP3-repressed editing target, undergoes a protein recoding editing at stop codon [Stop →Trp (W)]. Because of editing suppression by DAP3, the unedited PDZD7WT, which is more tumorigenic than edited PDZD7Stop518W, is accumulated in tumors. In sum, cancer cells may acquire malignant properties for their survival advantage through suppressing RNA editome by DAP3.

overall_design: We constructed strand-specific cDNA libraries of 6 total RNA samples and sequenced the libraries using Illumina Hiseq 4000 platform achieving over 200 millions of mappable reads per sample. Then we examined Adenosine-to-inosine (A-to-I) editing sites in EC109 and KYSE180 cells after depletion of genes of our interest.

first: Chen

last: Leilei

first: Han

last: Jian

first: Omer

last: An


title: The transcriptional landscapes of a hepatoma cell line grown on scaffolds of Extracellular Matrix proteins

status: Public on Jul 07 2020

city: Basel

first: Anastasiya

last: Boersch

address: Klingelbergstrasse 50 / 70

department: Biozentrum

country: Switzerland

institute: University of Basel

summary: The in vivo microenvironment of cells is complex and highly dynamic, resulting from an interplay between matrix proteins with several other cues and biomolecules. Although the effects of the cellular niche have been investigated for a number of cell types using different molecular approaches, a comprehensive information on the global transcriptome at various stiffness levels of 3D scaffolds resulting derived from components of the extracellular matrix (ECM) are still lacking. In this study, we explored the effect of the biomechanical parameters of Collagen I and Matrigel (ECM) on transcriptional gene regulation in a cell culture system. Using Huh7 cells cultured on traditional cell culture plates or on the component of the ECM, we have generated transcriptome sequencing data that may be further explored to understand the differentiation and growth potential of this cell for the development of 3D cultures. Assessment of the hepatocyte phenotype in our data set would also be useful for the development of systems mimicking the in vivo structure and function of liver cells which still remains a challenge.

overall_design: 3 Biological replicates of each growth type were sequenced. There were 6 conditions of matrix scaffolds. A total of 18 samples were sequenced.

first: Souvik

last: Ghosh

last: Börsch

first: Mihaela

last: Zavolan


title: ACME dissociation: a versatile cell fixation-dissociation method for single-cell transcriptomics

status: Public on Jun 03 2020

city: Oxford

first: Nathan

last: Kenny

address: Headington

institute: Oxford Brookes

summary: Single-cell sequencing technologies are revolutionizing biology, but are limited by the need of dissociating fresh samples that can only be fixed at later stages. We present ACME (ACetic-MEthanol) dissociation, a species-versatile cell dissociation approach that fixes cells  as they are being dissociated. ACME-dissociated cells have high RNA integrity, can be cryopreserved multiple times, can be sorted by Fluorescence-Activated Cell Sorting (FACS) and are permeable, enabling combinatorial approaches of single-cell transcriptomics. ACME is based on cheap reagents and it can be done in most labs and even sampling trips. As a proof of principle, we have performed SPLiT-seq with ACME cells to obtain around ~35K cells from two planarian species and identified all previously described cell types in similar proportions. This technique allows fixed, dissociated cells to be obtained from diverse organisms  that can be cryopreserved and subjected to combinatorial barcoding methods for single-cell transcriptomics  and thus will accelerate our knowledge of cell types across the tree of life.

sample_organism:
Dugesia japonica
Schmidtea mediterranea

platform_organism:
Dugesia japonica
Schmidtea mediterranea

overall_design: Adult planarians were pooled, and cells disassociated using the novel ACME technique as described in Garcia-Castro et al 2020 (this manuscript). They were then used to perform SPLiTseq analysis. This sample was split into 3 sub-libraries for sequencing (Illumina Novaseq), however all sublibraries were derived from the same overall population.

first: Helena

last: Garcia-Castro

first: Marta

last: Iglesias

first: Patricia

last: Alvarez-Campos

first: Vincent

last: Mason

first: Anamaria

last: Elek

first: Anna

last: Schonauer

first: Victoria

last: Sleight

first: Jakke

last: Neiro

first: Aziz

last: Aboobaker

first: Jon

last: Permanyer

first: Manuel

last: Irimia

first: Arnau

last: Sebe-Pedros

first: Jordi

last: Solana