title: An alternative transcriptome shapes cell fate transitions in yeast (TES-seq)

overall_design: 3’ end RNA sequencing analysis of Saccharomyces cerevisiae SK1 strains. All strains used in this study harbor the IME1gene controlled by a copper inducible promoter (pCUP1), the NDT80 gene controlled by a galactose inducible promoter (pGAL) and a constitutively expressed Gal4 transcription factor fused to the hormone binding domain of the estrogen receptor (Gal4-ER). In addition, mutant strains include those with the following genotypes: set2, set3, set2 set3 and Spt16 with a C-terminal degron (SPT16-AID). 3 biological replicates of each condition were analysed for the sporulation and return to growth time courses. For the sporulation time course, cells were grown overnight in YPD medium, diluted, and shifted to sporulation media (SPO). IME1 was induced after 2h in Spo by adding 50 μM CuSO4. Degradation of Spt16-AID was induced at Spo 2h with 500 μM 3-indole-acetid acid (3-IAA, auxin). For the return to growth (RTG) time course, cells in meiotic prophase (Spo 6h) were washed and re-suspended in an equal volume of YPD. Details for library preparation are described below.


title: Coordinated control of adiposity and growth by anti‐anabolic kinase ERK7

status: Public on Dec 14 2020

city: Helsinki

first: Krista

last: Kokki

address: Viikinkaari 9, Biocenter 1, Institute of Biotechnology P.O.Box. 56

department: Department of Biosciences (Genetics)

country: Finland

institute: University of Helsinki

type: Expression profiling by high throughput sequencing

summary: Nutrient availability influences animal growth and metabolism. How starvation inhibits anabolic functions in vivo remains insufficiently  understood. Here we establish an atypical MAP kinase ERK7 as an inhibitor of growth and lipid anabolism with an essential role in the  physiological adaptation to starvation stress. Drosophila null mutants of ERK7 display accelerated growth rate and elevated triacylglycerol  (TAG) stores on rich diet. ERK7 inhibits TAG storage in the Drosophila fat body by negatively regulating the expression of the Gli-similar  transcription factor Sugarbabe. While displaying growth advantage on rich diet, ERK7 mutants fail to cease their growth during starvation  and are consequently starvation sensitive. ERK7 modulates the majority of the starvation-induced gene expression response in a genomewide  setting, influencing, for example, genes involved in insulin signaling and lipid metabolism. In conclusion, ERK7 regulates peripheral  nutrient sensing to control animal growth and lipid metabolism with respect to nutrient availability.

overall_design: Control and ERK7 mutant flies starved and fed

first: Kiran

last: Hasygar

first: Ville

last: Hietakangas


title: Tissue-specific transcriptional imprinting and heterogeneity in human innate lymphoid cells revealed by full-length single-cell RNA-sequencing

city: Huddinge

first: Luca

last: Mazzurana

address: Alfred Nobels Aleè 8

country: Sweden

institute: Karolinska Institutet

summary: The impact of the microenvironment on innate lymphoid cell (ILC)-mediated immunity in humans remains largely unknown. Here we used full-length Smart-seq2 single-cell RNA-sequencing to unravel tissue-specific transcriptional profiles and heterogeneity of CD127+ ILCs across four human tissues. Correlation analysis identified gene modules characterizing the migratory properties of tonsil and blood ILCs, and signatures of tissue-residency, activation and modified metabolism in gut and lung ILCs. Trajectory analysis revealed potential differentiation pathways from circulating and tissue-resident naïve ILCs to a spectrum of mature ILC subsets. In the lung we identified both CRTH2+ and CRTH2- ILC2 with lung-specific signatures, which could be recapitulated by alarmin-exposure of circulating ILC2. Finally, we describe unique TCR-V(D)J-rearrangement patterns of blood ILC1-like cells, revealing a subset of potentially immature ILCs with TCR-d rearrangement. In summary, we provide publicly available data as a resource for in-depth understanding of ILC-mediated immunity in humans, with implications for disease.

overall_design:
In this study, we analyzed human ILCs through full-length scRNAseq, combining data from four different tissues: peripheral blood from three healthy donors,  lung tissue from four patients undergoing lobectomy for lung cancer, and biopsies from the ascending colon obtained from three tumor-screening patients with known genetic predisposition for colon cancer but without any polyps or tumors at the time of sampling. The previously published tonsil data was also used in this study and can be found at GEO: GSE70580  (all GSMs); Sequence Read Archive: SRP060416.

>>>Submitter declares that the raw data for non-tonsil samples will be deposited in EGA due to patient privacy concerns.<<<

first: Jenny

last: Mjösberg


title: Transcriptional effect of early-life adversity in adult hippocampus

status: Public on Dec 15 2020

city: Bethesda

first: Anton

last: Schulmann

state: Maryland (MD)

address: 35 Convent Dr

country: USA

institute: NIH/NIMH

summary: Early-life adversity (ELA) is associated with lifelong memory deficits, yet the responsible mechanisms remain unclear. We imposed ELA by rearing rat pups in simulated poverty, assessed hippocampal memory, and probed changes in gene expression, their transcriptional regulation and the consequent changes in hippocampal neuronal structure. ELA rats had poor hippocampal memory and stunted hippocampal pyramidal neurons, associated with ~140 differentially expressed genes. Upstream regulators of the altered genes included glucocorticoid receptor and, unexpectedly, the transcription factor neuron-restrictive silencer factor (NRSF/REST). NRSF contributed critically to the memory deficits because blocking its function transiently following ELA rescued spatial memory and restored the dendritic arborization of hippocampal pyramidal neurons in ELA rats. Blocking NRSF function in vitro augmented dendritic complexity of developing hippocampal neurons, suggesting that NRSF represses genes involved in neuronal maturation. These findings establish important, surprising contributions of NRSF to ELA-induced transcriptional programming that disrupts hippocampal maturation and memory function.

overall_design: Gene expression was profiled in dorsal hippocampi from 8-week-old male rats that were subjected to either control rearing conditions (n=4) or ELA (n=5), and did not undergo behavioral testing.

first: Jessica

middle: L

last: Bolton

first: Limor

last: Regev

first: Shan

last: Jiang

first: Yuncai

last: Chen

last: Molet

first: Megan

middle: M

last: Garcia-Curran

first: Akanksha

last: Singh-Taylor

first: Noriko

last: Kamei

first: Manlin

last: Shao

first: Yoav

last: Noam

first: Ali

last: Mortazavi

first: Tallie

middle: Z

last: Baram


title: CRNKL1 is a highly selective regulator of intron-retaining HIV-1 and cellular mRNAs

city: Erlangen

first: Han

last: Xiao

address: Schlossgarten 4

country: Germany

institute: Institute of Clinical and Molecular Virology, University Hospital Erlangen, Friedrich-Alexander Universität Erlangen-Nürnberg

summary: The human splicing factor CRNKL1 was identified as a factor influencing splicing and/or export of the HIV RNA.

overall_design: Following a genome-wide screen, CRNKL1 was selectively knocked down in Jurkat cells and cellular and viral RNA were analyzed in bulk RNA-Seq

first: Xiao

last: Han

first: Wyler

last: Emanuel

first: Milek

last: Miha

first: Grewe

last: Bastian

first: Kirchner

last: Philipp

first: Ekici

last: Arif

first: Oliveira

middle: V

last: Silva

first: Ana

last: Beatriz

first: Jungnickl

last: Doris

first: Florian

last: Full

first: Marco

last: Thomas

first: Markus

last: Landthaler

first: Armin

last: Ensser

first: Klaus

last: Überla


title: Meningeal lymphoid structures are activated under acute and chronic spinal cord pathologies

status: Public on Dec 10 2020

city: Rehovot

first: Ido

last: Amit

address: 234 Herzl st.

department: Immunology

country: Israel

institute: Weizmann Institute of Science

summary: Tertiary lymphoid structures (TLS) are organized aggregates of B and T cells formed ectopically during different life periods in response to inflammation, infection, or cancer. Here, we describe formation of structures reminiscence of TLS in the spinal cord meninges under several central nervous system (CNS) pathologies. Following acute spinal cord injury, B and T lymphocytes locally aggregate within the meninges to form TLS, which continue to accumulate during the late phase of repair, with a negative impact on subsequent pathological conditions, such as experimental autoimmune encephalomyelitis. Using a chronic model of spinal cord pathology, the mSOD1 mouse model of amyotrophic lateral sclerosis, we further showed by single cell RNA-sequencing that a meningeal lymphocyte niche forms, with a unique organization and activation state, including accumulation of pre-B cells in the spinal cord meninges. Such a response was not found in the CNS-draining cervical lymph nodes. The present findings suggest that a unique immune response develops in the meninges during various neurological pathologies in the CNS, a reflection of its immune privileged nature.

overall_design: single cell RNA-seq of B and T cells from mSOD1 mouse meninges or lymph node

first: Merav

last: Cohen

first: Amir

last: Giladi

first: Catarina

last: Raposo

first: Mor

last: Zada

first: Baoguo

last: Li

first: Julia

last: Ruckh

first: Aleksandra

last: Deczkowska

first: Boaz

last: Mohar

first: Ravid

last: Shechter

first: Rachel

middle: G

last: Lichtenstein

first: Michal

last: Schwartz


title: The mTOR regulated RNA-binding protein LARP1 requires PABPC1 for guided mRNA interaction

status: Public on Dec 11 2020

city: Glasgow

first: Ania

last: Wilczynska

address: Switchback Road

country: United Kingdom

institute: Beatson Insistute for Cancer Research

summary: The mammalian target of rapamycin (mTOR) is a pivotal kinase responsible for transducing cellular energy signals to regulate a host of metabolic processes including protein synthesis, which in turn regulate cell growth and proliferation. All aspects of mRNA life cycle are controlled by protein/RNA interactions and although several effectors of mTOR signalling have been identified to date, how mTOR re-sculptures the mRNA interactome is unknown. Here we characterise mTOR regulated RNA-binding proteins, identifying LARP1, whose binding to RNA increases upon mTOR inhibition. We identified over 3800 LARP1 bound mRNAs, which can be broken down into two groups, those constantly bound by LARP1, or mRNAs that increase their interaction following mTOR inhibition.  LARP1 has been implicated in the control of TOP mRNA translation and importantly we observe a large number of TOP mRNAs increasing association with LARP1 upon mTOR inhibition. Regarding the regulation of LARP1, we show that LARP1 and PABP show coordinated differential mRNA binding after mTOR inhibition. Importantly we find that LARP1-PABP interaction is important for LARP1 mRNA binding and mutations in the DM15 domain of LARP1 do not perturb its RNA interaction. Lastly we show that mRNAs bound by LARP1 and PABP are translationally repressed, including mRNAs encoding proteins critical for cell growth and survival.

overall_design: RNA-IP was conducted on endogenous LARP1 and PABP with and without Torin treatment for 1h. RNA-IPs were also conducted for overexpressed Flag-LARP

first: Ewan

last: Smith


title: Single-cell RNA sequencing of Tocilizumab-treated peripheral blood mononuclear cells as an in vitro model of inflammation

city: San Francisco

first: George

middle: Christopher

last: Hartoularos

state: CA

address: 513 Parnassus Ave S965F

department: Graduate Division

institute: University of California, San Francisco

summary: COVID-19 has posed a significant threat to global health. Early data has revealed that IL-6, a key regulatory cytokine, plays an  important role in the cytokine storm of COVID-19. Multiple trials are therefore looking at the effects of Tocilizumab, an IL-6  receptor antibody that inhibits IL-6 activity, on treatment of COVID-19, with promising findings. As part of a clinical trial looking  at the effects of Tocilizumab treatment on kidney transplant recipients with subclinical rejection, we performed single-cell RNA  sequencing of comparing stimulated PBMCs before and after Tocilizumab treatment. We leveraged this data to create an in vitro  cytokine storm model, to better understand the effects of Tocilizumab in the presence of inflammation. Tocilizumab-treated cells  had reduced expression of inflammatory-mediated genes and biologic pathways, particularly amongst monocytes. These results  support the hypothesis that Tocilizumab may hinder the cytokine storm of COVID-19, through a demonstration of biologic impact at  the single-cell level.

overall_design: Examination of peripheral blood mononuclear cells (PBMCs) from 15 human kidney transplant recipients, diagnosed with subclinical rejection on their 6-month post-transplant protocol biopsy and randomized to either continue standard of care (Tacrolimus, mycophenolate, and steroid) immunosuppression (control arm, 8 patients) or standard of care plus Tocilizumab (Tocilizumab treatment arm, 7 patients).

first: Arya

last: Zarinsefat

middle: C

first: Dmitry

last: Rychkov

first: Priyanka

last: Rashmi

first: Sindhu

last: Chandran

first: Flavio

last: Vincenti

first: Chun

middle: J

last: Ye

first: Minnie

last: Sarwal


title: Expression profiling in cohesin mutant MCF10A epithelial and CMK leukaemia cells

status: Public on Dec 13 2020

city: Dunedin

first: Jisha

last: Antony

state: Otago

address: 58 Hanover Street

department: Pathology

country: New Zealand

institute: University of Otago

summary: MCF10A cells were CRISPR-Cas9 edited to create heterozygous deletion in RAD21 and SMC3 subunits of cohesin. STAG2 is on the X chromosome, hence CRISPR-Cas9 editing resulted in complete loss of STAG2. Total RNA was sequenced from the MCF10A parental and cohesin mutant MCF10A  lines. The acute megakaryoblastic leukaemia cell line CMK was CRISPR-Cas9 edited to cotain STAG2 R614* mutation. CRISPR-Cas9 edited STAG2 mutant line showed complete loss of STAG2. CMK parental and the STAG2 mutant line were treated with Wnt3a for 4 hours and total RNA was sequenced at in the control or non-treated (con) and following 4 hours of Wnt3a treatment (Wnt3a4hr).

overall_design: RNA sequencing was done in triplicates for each cell type.

first: Chue

last: Chin

first: Gregory

last: Gimenez

middle: A

last: Horsfield


title: m6A methylome analysis reveals that m6A marks function in larval development and caste differentiation in honeybee (Apis mellifera)

city: GuangZhou

first: Yan

last: Li

state: GuangDong

address: No. 135, Xingang Xi Road, Guangzhou, 510275, P. R. China

department: School of Life Science

country: China

institute: Sun Yat-sen University

type: Methylation profiling by high throughput sequencing

summary: Epigenetic modifications are known to profoundly affect the development and behavior of social insects. In the well-known caste differentiation process of honeybee (Apis mellifera), female larvae with identical genomes are fed royal jellydifferently and develop into either normal workers or into very large, long-lived, and extremely fecund queens, and the queen-worker asymmetry of honeybee is known to be result largely to differential genomic imprinting during larval development that involves DNA methylation-based regulation. The discovery of reversible N6-methyladenosine (m6A) RNA methylation modification has defined a new era for RNA-metabolism-related genetic regulation, yet much remains unknown about m6A-mediated post-transcriptional regulatory mechanisms. Here, we report the first honeybee RNA m6A methylome. Specifically, we used the m6A-seq technique to examine the RNA m6A methylomes of honeybee larvae, including queen and worker larvae at multiple instar stages. We identified multiple conserved features of m6A methylation machinery and transcriptome-wide m6A distribution trends among insect species, and observed that m6A marks exert functions in regulating caste differentiation, with apparently particularly strong functional impacts on fifth instar worker larvae. Functional annotation of differentially methylated candidate caste-differentiation-related transcripts revealed many known regulators of caste differentiation (e.g. ILP-2, p110, PI3K, and JHAMT etc.) as well as the widely-studied Vitellogenin gene, which has not previously been implicated in caste differentiation. As ever-more regulatory roles for m6A marks are discovered, honeybees may become an excellent model studying the biology of such epi-transcriptomic regulatory systems, from embryonic development through holometabolous caste-specific development and on towards behavior and the emergent social hierarchies underlying eusociality in animals.

overall_design: The profiles of mRNA m6A modifications of three development stages from queen and worker honeybee

first: Miao

last: Wang

first: Yu

last: Xiao

first: Guan-Zheng

last: Luo

first: Guifang

last: Jia

first: Liming

last: Wu


title: NFATC2 modulates radiation toxicity in dermal fibroblasts from patients with severe side-effects of radiotherapy

city: Evry

first: Nicolas

last: FORTUNEL

address: 2 Rue Gaston Cremieux

country: France

institute: CEA

summary: Although it is well established that 5 to 15% of radiotherapy patients exhibit severe side-effects in healthy tissue, the cellular and molecular mechanisms involved are still poorly known, and the link between cellular and tissue radiosensitivity is still debated. We here studied fibroblasts from normal skin of patients with severe sequels of radiotherapy in order to examine if specific basal cell activities might be involved in susceptibility to side-effects. When compared to control cells, patient fibroblasts exhibited higher radiosensitivity and defects in DNA double strand breaks (DSB) repair and 8-oxoGuanines, abasic sites and glycol damaged base repair. Transcriptome profiling of dermal fibroblast cell strains from 16 radiotherapy patients with severe side-effects and 8 healthy individuals identified 540 genes specifically deregulated in the patients. Nuclear factor of activated T cells 2 (NFATC2) was one of the most differentially expressed genes. This transcription factor involved in immune responses was found poorly expressed at transcript and protein level in patient cells, whereas the NFATC2 gene region was hypermethylated. Furthermore, NFATC2 expression correlated to the cell survival after irradiation. Finally, we observed that the silencing of NFATC2 in normal cells by RNA interference leads to increased cellular radiosensivity and defects in DNA DSB repair. This study demonstrates that patients with clinical hypersensitivity also exhibit intrinsic cellular radiosensitivity in normal skin cells. It further reveals a new role for NFATC2 as a potential regulator of the cellular sensitivity to ionizing radiations.

overall_design: 24 dermal fibroblast samples total (8 from healthy donors and 16 from cancer patients)

first: Jerome

last: Lamartine

first: Michele

last: Martin

first: Joshua

last: Dulong

first: Tatiana

last: Vinasco-Sandoval


title: Expression data comparing control (day 0) and active (day 3) liver regeneration in mice that have undergone partial hepatectomy

status: Public on Jan 01 2021

city: Hong Kong

first: Stephanie

last: Ma

address: Rm 47, 1/F, Lab Block, Faculty of Medicine Bldg, 21 Sassoon Rd, Pok Fu Lam

department: School of Biomedical Sciences

country: Hong Kong

institute: The University of Hong Kong

summary:
Identification of altered gene expression of normal and regenerating liver post partial hepatectomy
Platform used - Affymetrix GeneChip Mouse Genome 430 2.0 Array

overall_design: Day 0 and Dary 3 partial hepatectomy

middle: Y


title: Disome-seq data in yeast cells

city: Beijing

first: Yan-Ming

last: Chen

address: Beichen West Rd.

institute: Institute of Genetics and Developmental Biology Chinese Academy of Sciences

summary: Genome-wide detection of monosome and disome distributions in yeast cells

overall_design: Examination of disome distribution in yeast

first: Taolan

last: Zhao

first: Wenfeng

last: Qian


title: In vivo nuclear RNA structurome reveals the RNA-structure regulation of mRNA processing in plants

city: Norwich

first: Yiliang

last: Ding

address: Colney lane

institute: John Innes Centre

summary: mRNA processing is critical for gene expression. A challenge in regulating mRNA processing is how to recognize the actual mRNA processing sites such as splice sites and polyadenylation sites when the sequence content is insufficient for this purpose. Previous studies suggested that RNA structure affects mRNA processing. However, the regulatory role of RNA structure in mRNA processing remains unclear. Here, we performed in vivo SHAPE chemical profiling on Arabidopsis nuclear RNAs and generated the in vivo nuclear RNA structure landscape. We found that nuclear mRNAs fold differently from cytosolic mRNAs. Notably, we discovered a two-nucleotide single-stranded RNA structure feature upstream of 5’ splice site that regulates splicing and is responsible for the selection of alternative 5’ splice sites. Moreover, we found branch point single-strandedness is associated with 3’ splice site recognition. We also identified an RNA structure feature comprising two close-by single-stranded regions that is specifically associated with both polyadenylation and alternative polyadenylation events. Our work demonstrates an RNA structure regulatory mechanism for mRNA processing.

overall_design: NAI treatment was applied to Arabidopsis seedlings under in vivo and deproteinized conditions. Control samples without NAI treatment  were used for each conditions. RNA-sequencing was performed for NAI treated and control samples by using Illumina Hiseq 4000.


title: Transcriptome analysis of naive B cells from CD19Cre and CD19Cre_STAT6vt mice

first: David

last: Voehringer

address: Wasserturmstrasse 3-5

department: Department of Infection Biology

institute: University Clinic Erlangen

summary: To investigate how activated STAT6 modulates B cells development and function in vivo, we characterized mice that express a constitutively active version specifically in B cells. CD19Cre_STAT6vt mice show spontaneous phosphorylation and nuclear translocation of STAT6 in B cells.  Based on the finding that STAT6vt accumulates in the nucleus of naïve splenic B cells in CD19Cre_STAT6vt mice, we next determined the changes of the global transcriptome by RNAseq analysis.

overall_design: We isolated naïve B cells by cell sorting (CD19+ CD3neg PIneg, purity >95%) from three age matched CD19Cre_STAT6vt and three CD19Cre control mice, purified total RNA and performed transcriptome analysis using the Illumina platform.

first: Paul

last: Haase

first: Daniel

last: Radkte


title: Embryonic transcriptome analysis of wt vs irf6-/- zebrafish at 4hpf

city: Boston

first: Eric

middle: Chienwei

last: Liao

state: Massachusetts

address: 185 Cambridge st czpn 4265

department: Center for Regenerative Medicine

institute: Massachusetts General Hospital

summary: We report transcriptional profiles of 4hpf zebrafish embryos from either wild-type or a CRISPR-targeted irf6 loss-of-function mutant. Irf6 is a transcription factor implicated in syndromic and nonsyndromic forms of cleft palate. To define critical genes involved in palate development, we harvested RNA from irf6-/- embryos before an embryonic lethal stage in zebrafish and carried out RNA-sequencing and subsequent analysis. We identify top differentially expressed genes that include genes involved in periderm development, genes expressed in developmental processes (such as Fgfs and Wnt-pathway genes), and genes involved in orofacial clefting. We identify a crucial gene esrp1 that we chose for further validation and downstream analysis.

overall_design: mRNA profiles of 4hpf wild-type or irf6-/- zebrafish embryos.

first: Edward

middle: B

last: Li

first: Justin

last: Cotney


title: Requirement of DNMT1 to orchestrate epigenomic reprogramming for NPM-ALK driven lymphomagenesis

status: Public on Dec 08 2020

city: Vienna

first: Gerda

last: Egger

address: Waehringer Guertel 18-20

country: Austria

institute: Medical University of Vienna

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

overall_design: Refer to individual Series


title: Gene expression during Arabidopsis ovule development at stages 9-12

city: Shanghai

first: Shi-Xia

last: Yu

address: Dongchuan Road, Shanghai Minhang District No. 800

institute: Shanghai Jiaotong University

summary: To obtain the global gene expression trends during ovule development, we collected samples of the gynoecium in developmental stages 9–10, 11, and 12 with three biological replicates to perform microarray assay.

overall_design: The plants under normal growth conditions, the fresh pistils was collected and removed stigma.

first: Wen-Hui

last: Lin


title: Regulation of mitophagy by O-linked N-acetylglucosamine transferase is essential for hematopoietic stem cell maintenance

status: Public on Dec 09 2020

city: Shinjuku-ku

first: Koichi

last: Murakami

state: Tokyo

address: Shinanomachi 35

country: Japan

institute: Keio University School of Medicine

summary: O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT) catalyzes post-translational modification of target proteins by introducing O-GlcNAc moiety on serine or threonine residues. OGT critically regulates various cellular functions in many cell types. However, its roles in hematopoietic stem and progenitor cells (HSPCs) are totally unknown. Here we report critical roles of OGT in homeostasis of HSPCs.  Ogt is highly expressed in HSPCs and its disruption led to their rapid loss with increased reactive oxygen species and apoptosis. In particular, Ogt-deficient HSCs lost quiescence and showed severe defects in self-renewal and long-term repopulation capacities. Interestingly, Ogt-deficient HSCs accumulated defective mitochondria due to impaired mitophagy with decreased key mitophagy regulators, Pink1 and Bnip3, through dysregulation of host cell factor 1 (HCF1)-H3K4me3 pathway, and overexpression of Pink1 rescued impaired mitophagy and numbers of Ogt-deficient HSCs.  In summary, our results revealed that OGT plays an essential role in HSC maintenance by assuring mitochondrial quality through mitophagy.

overall_design: Analysis of transcriptional profiles in Ogt WT and KO LSK cells

first: K

first: M

last: Oshima

first: S

last: Koide

first: D

last: Kurotaki

first: W

last: Kawase

first: A

last: Iwama

first: T

last: Tamura

first: H

last: Nakajima


title: Nonsense mediated decay and a novel protein Period-2 regulate casein kinase I in an opposing manner to control circadian period in Neurospora crassa

city: Hanover

first: Christina

last: Kelliher

state: NH

address: 7400 Hinman

department: Molecular & Systems Biology

institute: Geisel School of Medicine at Dartmouth

summary: The period-2 (prd-2) mutant of Neurospora crassa is characterized by recessive inheritance of a 26-hour long period phenotype of the circadian clock. PRD-2 encodes a putative RNA-binding protein. To identify PRD-2 target genes and its role in the clock, we performed transcript profiling in delta prd-2 compared to control samples using RNA-Sequencing. Hundreds of genes are affected by loss of prd-2, including decreased levels of Casein Kinase I (ck-1a, NCU00685). The circadian period length in Neurospora is exquisitely sensitive to ck-1a levels, and we attribute the long period length defect in the prd-2 mutant to low levels of ck-1a transcript, decreased ck-1a transcript stability, and lower levels of CKI protein. Thus, Casein Kinase I gene expression is subject to complex regulation in the circadian clock.

overall_design:
To identify the role of the period-2 mutant in the circadian clock, transcript profiling was performed and compared to control samples. RNA-Sequencing was performed by Novogene. Hundreds of genes are affected by loss of the period-2 gene, and the clock-relevant target is downregulation of ck-1a (NCU00685).
RNA-Seq was performed on biological triplicate samples from 25C, Bird 1.8% glucose medium, constant light cultures.

first: Randy

last: Lambreghts

first: Qijun

last: Xiang

first: Christopher

last: Baker

first: Jennifer

last: Loros

first: Jay

last: Dunlap


title: Mutations within FGFR1 and BRAF indicate superior outcome in diffuse midline gliomas, H3F3A K27M mutant

city: Hamburg

first: Julia

middle: Elisabeth

last: Neumann

state: Hamburg

address: Martinistr. 52

department: Institute of Neuropathology

institute: University Medical Center Hamburg-Eppendorf

type: Methylation profiling by genome tiling array

summary: Diffuse midline glioma (DMG) is a devastating disease that is defined by its localization, by the diffuse growth of astrocytic tumor cells, and by K27M mutations within Histone H3 proteins. In a large series of such tumor samples (n=83), we show here that 12 % of these cases (n=10) harbor concomitant hotspot mutations within FGFR1 or BRAF. Respective cases were all located in midline structures and matched with reference cases of DMG using global DNA methylation profiling. TP53 mutations were significantly more frequent in BRAF/FGFR1 wild  type cases. Presence of hotspot mutations within FGFR1 or BRAF was associated with a significantly better overall survival, which was independent of the patients’ age and tumor localization. Thus, our results establish the relevance of FGFR1 or BRAF mutations in DMG as a prognostic marker and open new targeted therapy options in this subgroup of DMG.

overall_design: We performed global DNA methylation analyses (850K) and classified BRAF/FGFR1 MU (n=4) and BRAF/FGFR1 WT cases (n=16) of diffuse midline gliomas, K27M mutant.

first: Ulrich

last: Schüller

first: Peter

last: Iglauer

first: Mario

last: Dorostkar

first: Christian

last: Mawrin

first: Jochen

last: Herms

first: Armin

last: Giese

first: Markus

last: Glatzel

middle: E


title: Transcriptome sequencing analysis of MyoD-induced transdifferentiation in 10T1/2 fibroblasts using RNA-seq

first: Tom

last: Cheung

address: Clear Water Bay, Kowloon

department: Division of Life Science

institute: Hong Kong University of Science and Technology

summary: To understand the effect of LncMyod on MyoD-induced transdifferentiation of 10T1/2 fibroblast cells, we performed RNA-seq on LncMyoD-KO 10T1/2 fibroblasts with MyoD-induced transdifferentiation.

overall_design: The polyA+ RNA from cultured 10T1/2 fibroblast cells were sequenced using DNBSEQ-G400 sequencer (BGI) (2 x 100)

first: Anqi

last: Dong

first: Ran

last: Yi

middle: H


title: Requirement of DNMT1 to orchestrate epigenomic reprogramming for NPM-ALK driven lymphomagenesis [RNA-Seq]

first: Gerda

last: Egger

summary: Malignant transformation depends on genetic and epigenetic events that result in a burst of deregulated gene expression and chromatin changes. To dissect the sequence of events in this process, we used a T cell-specific lymphoma model based on the human oncogenic NPM-ALK translocation. We find that transformation of T cells shifts thymic cell populations to an undifferentiated immunophenotype, which occurs only after a period of latency, accompanied by induction of the MYC-NOTCH1 axis and deregulation of key epigenetic enzymes. We discover aberrant DNA methylation patterns, overlapping with regulatory regions, plus a high degree of epigenetic heterogeneity between individual tumors. In addition, ALK positive tumors show a loss of associated methylation patterns of neighboring CpG sites. Notably, deletion of the maintenance DNA methyltransferase DNMT1 completely abrogates lymphomagenesis in this model, despite oncogenic signaling through NPM-ALK, suggesting that faithful maintenance of tumor-specific methylation through DNMT1 is essential for sustained proliferation and tumorigenesis

overall_design: Evaluation of gene expression and DNA methylation signatures in a transgenic mouse model of ALCL

first: Elisa

last: Redl

first: Raheleh

last: Sheibani-Tezerji

first: Crhistian

last: de Jesus Cardona

first: Patricia

last: Hamminger

first: Gerald

last: Timelthaler

first: Melanie

middle: R

last: Hassler

first: Masa

last: Zrimsek

first: Sabine

last: Lagger

first: Thomas

last: Dillinger

first: Lorena

last: Hofbauer

first: Kristina

last: Draganic

first: Andreas

last: Tiefenbacher

first: Michael

last: Kothmayer

first: Charles

last: Dietz

first: Bernard

last: Ramsahoye

first: Lukas

last: Kenner

first: Christoph

last: Bock

last: Seiser

first: Wilfried

last: Ellmeier

first: Gabriele

last: Schweikert


title: Single cell RNA-seq of neonatal heart regeneration

city: New York

first: Zhaoning

last: Wang

state: New York

address: 701 West 168th Street Hammer 14 - 1410

department: Genetics and Development

institute: Columbia University

summary: The adult mammalian heart has limited capacity for regeneration following injury, whereas the neonatal heart can readily regenerate within a short period after birth. Neonatal heart regeneration is orchestrated by multiple cell types intrinsic to the heart, as well as immune cells that infiltrate the heart after injury. To elucidate the transcriptional responses of the different cellular components of the mouse heart following injury, we performed single cell RNA-sequencing on neonatal hearts at various time points following myocardial infarction, and coupled the results with bulk tissue RNA-sequencing data collected at the same time points. Concomitant single cell ATAC-sequencing exposed underlying dynamics of open chromatin landscapes and regenerative gene regulatory networks of diverse cardiac cell types, and revealed previously unknown mediators of cardiomyocyte proliferation, angiogenesis and fibroblast activation. Together, our data provide a transcriptional basis for neonatal heart regeneration at single cell resolution and suggest new strategies for enhancing cardiac function in response to injury.

overall_design: We performed myocardial infarction (MI) or sham surgeries to mouse hearts at postnatal day 1 or day 8. At 1d and 3d post P1 and P8 surgery, ventricle samples below the surgery plane were collected for single cell RNA-Seq.

first: Miao

last: Cui

first: Wei

last: Tan

first: Eric

last: Olson


title: Requirement of DNMT1 to orchestrate epigenomic reprogramming for NPM-ALK driven lymphomagenesis [RRBS]

first: Gerda

last: Egger

first: Elisa

last: Redl

first: Raheleh

last: Sheibani-Tezerji

first: Crhistian

last: de Jesus Cardona

first: Patricia

last: Hamminger

first: Gerald

last: Timelthaler

first: Melanie

last: Hassler

first: Masa

last: Zrimsek

last: Lagger

first: Christian


title: Histone modifications in LncMyoD-KD mouse muscle stem cells by ChIP-seq

first: Tom

last: Cheung

type: Genome binding/occupancy profiling by high throughput sequencing

summary: We report the application of ChIPmentation on examination of H3K27ac histone mark in cultured satellite cells with siRNA treatment targeting LncMyoD. We observed no difference of H3K27ac mark around transcription start sites (TSS) after LncMyoD KD.

overall_design: Examination of H3K27ac mark in LncMyoD-KD satellite cells.

first: Anqi

last: Dong

first: Kangning

last: Lin


title: Variability of an early developmental cell population underlies stochastic laterality defects

city: Berlin

first: Jan

middle: Philipp

last: Junker

address: Hannoversche Str. 28

department: Berlin Institute for Medical Systems Biology

institute: Max Delbrück Center for Molecular Medicine

summary: Embryonic development seemingly proceeds with almost perfect precision. However, it is largely unknown how much microscopic variability is hidden beneath this macroscopic accuracy. Here, we quantified embryo-to-embryo variability in vertebrate development, by studying cell number variation in the zebrafish endoderm. We noticed that the size of a sub-population of the endoderm, the dorsal forerunner cells (which later forms the left-right organizer), is highly variable between individual embryos. We found that the frequency of left-right laterality defects is increased drastically in embryos with a low number of dorsal forerunner cells, and we observed that these fluctuations are largely stochastic. Hence, a stochastic variation in early development leads to a remarkably strong macroscopic phenotype. These fluctuations appear to be caused by variable deposition of maternal factors involved in specification of the dorsal forerunner cells. In summary, we here dissect cause and consequence of embryo-to-embryo variability in a vertebrate model.

overall_design: Transcriptomic comparison of Individual embryos from two zebrafish strains (AB and TL) at 2.25, 3.25, 4.25 and 5.25 hpf with two biological replicates.

first: Roberto

last: Moreno-Ayala

middle: P


title: Cell-type-specific gene regulatory networks underlying murine neonatal heart regeneration at single-cell resolution

first: Zhaoning

last: Wang


title: A Long Noncoding RNA, LncMyod, Regulates Myogenic Lineage Progression by Modulating Chromatin Accessibility


title: Transcriptome sequencing analysis of LncMyod-knockdown satellite cells by RNA-seq

summary: To understand the effect of LncMyod knockdown on cultured satellite cells, we isolated satellite cells from mouse hindlimb and cultured them in vitro with siRNA treatment targeting LncMyod. Following RNA-seq showed that loss of LncMyod leads to difficiencies in myogenic differentiation.

overall_design: The polyA+ RNA from cultured satellite cells with siRNA treatment targeting LncMyod were sequenced using Illumina NextSeq 500 (2 x 150)

first: Christopher

last: Preusch


title: Chromatin structure profiling of mouse muscle stem cells by ATAC-seq

summary: By comparing ATAC-seq from freshly isolated satellite cells (QSC), satellite cells isolated 2.5 days post-injury (ASC), and satellite cells cultured for differentiation, we observed a gradual opening of satellite cell chromatin structure during activation and  differentiation process. We used ATAC-seq to study the functional role of LncMyod during satellite cell differentiation and identified genes that exhibited decrease of accessibilities to their transcription start sites (TSS). The majority of the regions are myogenesis-related genes, suggesting LncMyod plays a critical role in establishing permissive chromatin environment for myogenic progression.

overall_design: Examination of chromatin accessibility using ATAC-seq in QSC, ASC, satellite cells cultured for differentiation with control siRNA (LncMyod Control), and satellite cells cultured for differentiation with siRNA targeting LncMyod (LncMyod Knockdown).


title: Differential gene expression during embryoid body (EB) formation of mouse embryonic stem cells when Hnrnpll was depleted

status: Public on Dec 06 2020

first: Jia

last: Yu

address: Dong Dan San Tiao 5, Dongcheng District

institute: Peking Union Medical College

summary: This work indicates Hnrnpll depletion mES cells cannot exit from pluripotency programme, showing embryoid body formation deficiency and no spontaneous differentiation withdraw LIF. Hnrnpll promotes multiple ES cell preferred-exon skipping during ES cells differentiation, to reduce the ES cells specific isoform and facilitate ES cells exit from pluripotency towards differentiation.

overall_design: polyA+ RNA profiles of wildtype or Hnrnpll-/- mESC EB differentiation at day 0, 4, 8.

first: Xue

first: Changyun

last: Ping

first: Puwen

last: Tan

first: Dong

first: Yanni

last: Ma

first: Yue

last: Huang


title: Reduction of alternative electron acceptors drives biofilm formation in Shewanella algae

status: Public on Dec 05 2020

city: La Laguna

first: Jose

last: Gonzalez

address: School of Pharmacy

department: Dept. Microbiology

country: Spain

institute: University of La Laguna

summary: Shewanella spp. possess a broad respiratory versatility, which contributes to the occupation of hypoxic/anoxic environmental or host-associated niches. Here we observed a strain-specific induction of biofilm formation in response to supplementation with the anaerobic electron acceptors dimethyl sulfoxide (DMSO) and nitrate in a panel of Shewanella algae isolates. The respiration-driven biofilm response is not observed in DMSO and nitrate reductase deletion mutants of the type strain S. algae CECT 5071, and can be restored upon complementation with the corresponding reductase operon(s) but not by an operon containing a catalytically inactive nitrate reductase. The distinct transcriptional changes, proportional to the effect of these compounds on biofilm formation, include cyclic di-GMP (c-di-GMP) turnover genes. In support, ectopic expression of the c-di-GMP phosphodiesterase YhjH of Salmonella Typhimurium but not its catalytically inactive variant decreased biofilm formation. The respiration-dependent biofilm response of S. algae may permit differential colonization of environmental or host niches.

overall_design: Transcriptomics in media with two different electron acceptors, DMSO and nitrate

middle: M


title: Expression data from DLBCL expression study

status: Public on Dec 07 2020

city: PLEASANTON

first: Rajiv

last: Dua

address: 4300 Hacienda Dr

institute: Roche Molecular Diagnostics

summary: We used microarray data sets to determine concordance of microarray vs qRT-PCR based LCOO cell-of-origin for Diffuse large B-cell Lymphome subtypes.

overall_design: Three independent sets of samples were processed and analyzed for a total of 180 samples. Training sets were developed to build and develop a DLBCL classification model. A subsequent validation data set was used to validate the classification model.

first: Chris

last: Santini

first: Yu

last: Tai

first: Marcel

last: Fontecha


title: Genome-wide methylation analysis of blood cells from homozygotic triplets

city: Solna

first: Klas

last: Strååt

address: Akademiska stråket 1

department: Medicine

institute: Karolinska Institutet

summary:
We report genome-wide methylation profiles of four blood cell populations from homozygotic triplets, of which two has Hodgkin lymphoma (triplets A and B) and one does not (triplet C). DNA was isolated from CD34+ cells, marginal zone-like B cells, naïve B cells and switched memory B cells and analyzed using the Illumina EPIC platform.
We previously reported Hodgkin lymphoma development in two triplets where all three were homozygotic and had a constitutional deletion in the first intron of the MKL1 (megakaryoblastic leukemia 1) gene; see Bjorkholm et al, Blood 2013;121:4807. The significance by the MKL1 gene deletion in the same set of homozygotic triplets has recently been studied further by Record et al (Haematologica 2019, PMID 31582539), whose investigations suggest that dysregulated MKL1 activity may participate in B cell transformation and HL pathogenesis. It is well recognized that first degree relatives of HL patients carry an increased risk of developing the disease. In this study, we aimed to evaluate the genome-wide DNA methylation profile of several B cell populations as well as CD34+ cells in the triplets.

overall_design: Bisulphite treated DNA from a total of 12 samples (4 cell populations from 3 triplets) were hybridized to the Illumina Infinium MethylationEPIC_v-1-0 beadchip with 850.000 CpG site-specific probes.

first: Thale

middle: K

last: Olsen


title: Optimized RNA-targeting CRISPR/Cas13d technology outperforms shRNA in identifying functional circRNAs

city: Torino

first: Pier Paolo

last: Pandolfi

state: TO

address: 8 Via Verdi

institute: University of Torino

summary: Circular RNAs (circRNAs) are widely expressed, but their functions remain largely unknown. To study circRNAs in a high-throughput manner, short hairpin RNA (shRNA) screens1 have recently been used to deplete circRNAs by targeting their unique back-splicing junction (BSJ) sites. Here, we report frequent discrepancies between shRNA-mediated circRNA knockdown efficiency and the corresponding biological effect, raising pressing concerns about the robustness of shRNA screening for circRNA functional characterization. To address this issue, we leveraged the CRISPR/Cas13d system2 for functional study of circRNAs by optimizing the strategy for designing single guide RNAs to deplete circRNAs. We then performed shRNA and CRISPR/Cas13d parallel screenings and demonstrated that shRNA-mediated circRNA screening yielded a high rate of false positive phenotypes. Furthermore, using a CRISPR/Cas13d screening library targeting over 2,500 human hepatocellular carcinomas (HCC) related circRNAs, we identified a group of circRNAs, whose inhibition increased the therapeutic efficacy of sorafenib, an approved HCC drug. Collectively, these data demonstrate that CRISPR/Cas13d system is an effective approach to study the function of circRNAs in a high-throughput manner.

overall_design: Identification of circular RNAs essential for cell survival in hepatocellular carcinoma using CRISPR RNA Cas13d and shRNA screening. CRISPR Cas13d screening was also performed to identify circular RNAs that drive resistance to sorafenib in liver cancer. Comparative analysis on specificity of CRISPR Cas13d and shRNA in knocking circular RNA was also performed using RNA-Seq

first: Yang

last: Zhang

first: Tuan

last: Nguyen

first: Xiao-Ou

first: Limei

first: Tin

last: Phan

first: John

last: Clohessy

first: Pier-Paolo


title: Merkel Cell Polyomavirus Encodes Circular RNAs (circRNAs) Enabling a Dynamic circRNA/microRNA/mRNA Regulatory Network

city: Pittsburgh

first: Bizunesh

last: Abere

state: PENNSYLVANIA

address: 5117 Centre Ave, Research Pav. Suite 1.8

department: Cancer Virology Program

institute: Hillman Cancer Center

summary: Viral noncoding RNAs have acquired increasing prominence as important  regulators of infection and mediators of pathogenesis. Circular RNAs (circRNAs) generated  by backsplicing events have been identified in several oncogenic human DNA  viruses. Here, we show that Merkel cell polyomavirus (MCV), the etiologic cause of ~80%  of Merkel cell carcinomas (MCCs), also expresses circular RNAs. By RNase R-resistant RNA  sequencing, four putative circRNA backsplice junctions (BSJs) were identified from the  MCV early region (ER). The most abundantly expressed MCV circRNA, designated circMCVT,  is generated through backsplicing of all of ER exon II to form a 762-nucleotide (nt) circular  RNA molecule. Curiously, circMCV-T, as well as two other less abundantly expressed  putative MCV circRNAs, overlaps in a complementary fashion the MCV microRNA (miRNA)  locus that encodes MCV-miR-M1. circMCV-T is consistently detected in concert with linear  T antigen transcripts throughout infection, suggesting a crucial role for this RNA molecule  in the regulatory functions of the early region, known to be vital for viral replication.  Knocking out the hairpin structure of MCV-miR-M1 in genomic early region expression  constructs and using a new high-efficiency, recombinase-mediated, recircularized  MCV molecular clone demonstrates that circMCV-T levels decrease in the presence of  MCV-miR-M1, underscoring the interplay between MCV circRNA and miRNA. Furthermore,  circMCV-T partially reverses the known inhibitory effect of MCV-miR-M1 on early gene  expression. RNase R-resistant RNA sequencing of lytic rat polyomavirus 2 (RatPyV2) identified  an analogously located circRNA, stipulating a crucial, conserved regulatory function of  this class of RNA molecules in the family of polyomaviruses.

sample_organism:
Homo sapiens
Rattus norvegicus

platform_organism:
Homo sapiens
Rattus norvegicus

overall_design: Ribodepleted RNaseR treated total RNA from MCV positive MCC derived cell lines, RatPyV2 positve rat parotid tissue or 293 cell transfected with a recircularized MCV-HF genome was used for RNA sequencing

first: Yuan

last: Chang


title: RNA sequencing of wild type and FXR KO CD8+T cells in fed/fasted LCMV-infected mice

status: Public on Dec 04 2020

city: Princeton

first: Yuri

last: Pritykin

state: NJ

address: 245 Carl Icahn Lab, Lewis-Sigler Institute for Integrative Genomics

institute: Princeton University

summary: Conditional ablation of FXR in T cells was achieved with CD4-Cre Nr1h4fl/fl mice. Twenty thousend congenically marked naive (CD44- CD62L+) WT and FXR KO T cells carrying the OT-I transgenic TCR were cotransferred into lymphoreplete hosts. Recipients were challenged with LCMV-OVA and allowed to feed Ad libitum (AL) or fasted (Fs) for 24h beginning at 6pm on D5 post-infection. Splenic CD44+ effector cells of each genotype were FACS-purified (double-sorted) at the end of the starvation period.

overall_design: Effects of genotype (WT vs FXR KO) and feeding status (AL vs Fs)

first: Clarissa

last: Campbell

first: Michael

last: Schizas

first: Joris

last: van der Veeken


title: Genome-wide mapping of 5-hydroxymethylcytosine enrichment in a novel model of 'NAFLD in a Dish'

status: Public on Dec 03 2020

city: Edinburgh

first: Amanda

middle: J.

last: Drake

address: 47 Little France Crescent

department: Centre for Cardiovascular Science

institute: University of Edinburgh

summary: Human pluriopotent H9 cells were differentiated to hepatocyte-like cells and exposed to a cocktail of lactate, pyruvate and octanoic acid (LPO) for 48 hours. This stimulated development of macrovesicular steatosis, which has previously been reported to impact on mitochondrial metabolism. Under our experimental conditions, we identified rewiring of the TCA cycle linked to macrovesicular steatosis. As key TCA cycle metabolites, alpha-ketoglutarate, succinate and fumarate impact on ten-eleven translocation (TET) enzyme activity, we hypothesised that this rewiring would impact on levels of 5-hydroxymethylcytosine (5hmC) enrichment across the genome. To this end, we performed DNA immunoprecipitation sequencing (DIPseq) to map any changes that occurred. Using IonTorrent, we sequenced our samples and generated bam files aligned to Homo sapiens Hg19. We used the MACS2 pipeline to compare our control and steatosis group, and identified a large number of changes in 5hmC enrichment in response to LPO exposure, indicating that TCA cycle wiring may impact on TET activity.

overall_design: 5hmC profiles were generated for ES-derived hepatocyte-like cells following treatment with lactate (10 mM), pyruvate (1 mM) and octanoate (2 mM) to induce intracellular lipid accumulation, resulting in a NAFLD-like phenotype

first: Matthew

last: Sinton


title: Single cell ATAC-seq of neonatal heart regeneration

first: Zhaoning

last: Wang

overall_design: We performed myocardial infarction (MI) or sham surgeries to mouse hearts at postnatal day 1 or day 8. At 3d post P1 and P8 surgery, ventricle samples below the surgery plane were collected for single cell ATAC-Seq.

first: Wei

last: Tan

first: Eric

last: Olson


title: Slow Adaptive Response of Budding Yeast Cells to Stable Conditions of Continuous Culture Can Occur Without Genome Modifications

status: Public on Dec 02 2020

city: Warszawa

first: Marek

last: Skoneczny

email: kicia@ibb.waw.pl, mrksknc@gmail.com

address: Pawińskiego 5A

country: Poland

institute: Institute of Biochemistry and Biophysics PAS

summary: Continuous cultures assure the invariability of environmental conditions and the metabolic state of cultured microorganisms, whereas batch-cultured cells undergo constant changes in nutrients availability. For that reason, continuous culture is sometimes employed in the whole transcriptome, whole proteome, or whole metabolome studies. However, the typical method for establishing uniform growth of a cell population, i.e., by limited chemostat, results in the enrichment of the cell population gene pool with mutations adaptive for starvation conditions. These adaptive changes can skew the results of large-scale studies. It is commonly assumed that these adaptations reflect changes in the genome, and this assumption has been confirmed experimentally in rare cases. Here we show that in a population of budding yeast cells grown for over 200 generations in continuous culture in non-limiting minimal medium and therefore not subject to selection pressure, remodeling of transcriptome occurs, but not as a result of the accumulation of adaptive mutations. The observed changes indicate a shift in the metabolic balance towards catabolism, a decrease in ribosome biogenesis, a decrease in general stress alertness, reorganization of the cell wall, and transactions occurring at the cell periphery. These adaptive changes signify the acquisition of a new lifestyle in a stable nonstressful environment. The absence of underlying adaptive mutations suggests these changes may be regulated by another mechanism.

overall_design: Comparison of transcriptomes of the wild type (WT) Saccharomyces cerevisiae W303 cell populations grown for 40 generations in continuous culture versus cell populations of the same strain grown in the same conditions for 240 generations. Three biological replicates of 40 generations of growth vs 240 generations of growth were performed. Each biological replicate had two technical replicates with dye swapping.

first: Joanna

last: Klim

first: Urszula

last: Zielenkiewicz

first: Anna

last: Kurlandzka

first: Szymon

last: Kaczanowski


title: Histone modifications in activated and quiescent mouse muscle stem cells by ChIP-seq

city: Hong Kong

first: Tom

last: Cheung

summary: We report the application of ChIPmentation on examination of H3K27me3 and H3K4me3 histone marks in freshly isolated satellite cells (QSC) and satellite cells isolated after 2.5 days post-injury (ASC). We observed increase of H3K4me3 marks around transcription start sites (TSS)  and decrease of H3K27me3 marks around TSS during satellite cell activation.

overall_design: Examination of H3K27me3 and H3K4me3 marks in QSC and ASC.

first: Anqi

last: Dong

first: Wai

last: So

first: Christopher

last: Preusch


title: mRNA sequencing analysis of a novel model of 'NAFLD in a Dish'

summary: The goal of this study was to determine the changes to cellular transcriptional programs following induction of intracellular lipid accumulation, with the aim of confirming the utility of this model for exploring mechanisms underlying the pathogenesis of NAFLD. Comparisons with other publically available data confirm that the alterations we observe in mRNA expression are similar to those observed in both humans and rodents.

overall_design: mRNA profiles were generated for ES-derived hepatocyte-like cells following treatment with lactate (10 mM), pyruvate (1 mM) and octanoate (2 mM) to induce intracellular lipid accumulation, resulting in a NAFLD-like phenotype


title: Intestinal Differentiation Involves Cleavage of Histone H3 NTerminal Tails by Multiple Proteases

city: Milano

first: Diego

last: Pasini

address: Via Adamello, 16

country: Italy

institute: European Institute of Oncology

summary: The proteolytic cleavage of histone tails, also termed histone clipping, has been described as a mechanism for permanent removal of post-translational modifications (PTMs) from histone proteins. Such activity has been ascribed to ensure regulatory function in key cellular processes such as differentiation, senescence and transcriptional control for which different histone-specific proteases has been described. However, all these studies were exclusively performed using cell lines cultured in vitro and no clear evidences that histone clipping is regulated in vivo has been reported. Here we show that histone H3 N-terminal tails undergo extensive cleavage in the differentiated cells of the villi fraction of the mouse intestinal epithelium. Combining biochemical methods, 3D organoids cultures and in vivo approaches, we demonstrate that intestinal H3 clipping is the result of multiple proteolytic activities. We identified Trypsins and Cathepsin L as specific H3 tail proteases active in small intestinal differentiated cells and showed that their proteolytic activity is differentially affected by the PTM pattern of histone H3 tails. Together, our findings provide in vivo evidences of H3 tail proteolysis in mammalian tissues directly linking H3 clipping to cell differentiation.

overall_design: Intestinal villi and crypt subjected with antibodies.

first: Daniel

last: Fernandez

first: Karin

last: Ferrari


title: Single-Cell Transcriptional Profiling of Mouse Islets Following Short-Term ObesogenicDietary Intervention

city: Chicago

first: Raghavendra

middle: G

last: Mirmira

state: IL

address: 900 E. 57th Street

institute: University of Chicago

summary: Obesity is closely associated with adipose tissue inflammation and insulin resistance. Dysglycemia and type 2 diabetes results when islet β cells fail to maintain appropriate insulin secretion in the face of insulin resistance. To clarify the early transcriptional events leading to β-cell failure in the setting of obesity, we fed male C57BL/6J mice an obesogenic, high fat diet (60% kcal from fat) or control diet (10% kcal from fat) for one week and islets from these mice (from 4 high fat- and 3 control-fed mice) were subjected to single cell RNA sequencing analysis. Islet endocrine cell types (α cells, β cells, δ cells, PP cells) and other resident cell types (macrophages, T cells) were annotated by transcript profiles and visualized using Uniform Manifold Approximation and Projection for Dimension Reduction (UMAP) plots. UMAP analysis revealed distinct cell sub-populations (11 for β cells, 5 for α cells, 3 for δ cells, PP cells, ductal cells, endothelial cells), emphasizing the heterogeneity of cell populations in the islet. We identified that distinct β cell populations downregulate genes associated with the endoplasmic reticulum stress response and upregulate genes associated with insulin secretion, while others upregulate genes that impair insulin secretion, cellular proliferation, and survival. Moreover, all β cell populations negatively regulate genes associated with immune response activation. Our data indicate that an early transcriptional response in islets to an obesogenic diet reflects an attempt by distinct populations of β cells to augment or impair cellular function, possible harbingers of ensuing insulin resistance.

overall_design: Islet isolation. Male C57BL/6J mouse pancreas were isolated from 8 weeks of age, animals were fed either a LFD (10% kcal from fat; Research Diets D12450B), or HFD (60% kcal from fat; Research Diets D12492) for one week. Islets were isolated from collagenase-perfused pancreata as previously described (Stull, 2017). After isolation, mouse islets were handpicked and digested with Accutase (EMD Millipore Corporation) containing 2U/ml of DNAse during 5min at 37°C sob agitation (1000rpm). Digested cell islets were washed several times with PBS+2%FBS to eliminate DNAse and then filtered using a cell strainer (40mm). Single cells suspension and samples with more than 90% viability were used for scRNAseq.

first: Annie

last: Piñeros

first: Hongyu

last: Gao

first: Wenting

last: Wu

first: Yunlong

last: Liu

first: Sarah

last: Tersey


title: H2AK121ub in Arabidopsis favors a less accessible chromatin state at transcriptional regulation hotspots [RNA-seq]

city: Seville

first: Francisco

middle: José

last: Romero-Campero

address: Americo Vespucio 49

department: Institute for Plant Biochemistry and Photosynthesis

institute: University of Sevilla

summary: Although it is well established that Polycomb Group (PcG) complexes maintain gene repression through the incorporation of H2AK121ub and H3K27me3, little is known about the effect of these modifications on chromatin, which is fundamental to understand PcG function. Here, by integrating chromatin accessibility, histone marks and expression analyses in different Arabidopsis PcG mutants, we show that H2AK121ub marking favors a less accessible but still permissive chromatin at transcriptional regulation hotspots, which can be further reduced by EMF1 and H3K27me3. We found that when acting concurrently, H2AK121ub and H3K27me3 establish an inaccessible but responsive chromatin to marks levels, allowing gene reprograming. Conversely, when H3K27me3 is alone, chromatin is less responsive, indicating that H2AK121ub-marked hotspots are required for transcriptional responses. Interestingly, despite the loss of H2AK121ub and H3K27me3 leads to increased chromatin accessibility, this is not necessarily accompanied by transcriptional activation, indicating that gene expression is not always predictive of accessible chromatin.

overall_design: Arabidopsis seedlings Col-0 wild type, atbmi1abc triple mutant, clf28/swn7 double mutant, emf1-2 mutant, ring1ab double mutant and lhp1 mutant were grown under long-day condition (16 hr of light/8 hr of dark) at 22 °C on MS agar plates containing 1% sucrose and 0.7% agar and collected at 10 DAG (days after germination). Two independent biological replicates were considered for ATAC-seq and ChIP-seq data whereas three independent biological replicates were considered for RNA-seq data.

first: Myriam

last: Calonje

first: Yue

last: Zhue


title: H2AK121ub in Arabidopsis favors a less accessible chromatin state at transcriptional regulation hotspots

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

overall_design: Refer to individual Series


title: COSMIC-seq of bioactive hairpin N-methylpyrrole/N-methylimidazole polyamides

city: Memphis

first: Aseem

last: Ansari

state: TN

address: 262 Danny Thomas Place

department: Chemical Biology & Therapeutics

institute: St. Jude Children's Research Hospital

summary: Regulating desired loci in the genome with sequence-specific DNA-binding molecules is a major goal for the development of precision medicine. Pyrrole–imidazole (Py–Im) polyamides are synthetic molecules that can be rationally de-signed to target specific DNA sequences to both disrupt and recruit transcriptional machinery.  While in vitro binding has been extensively studied, in vivo effects are often difficult to predict using current models of binding energetics.  Determining the impact of genomic architecture and the local chromatin landscape on polyamide-DNA sequence specificity remains an unresolved question that impedes their utility in vivo.  In this report we directly identified polyamide–DNA interaction sites across the entire genome, by covalent crosslinking and capturing these events in the nuclei of human LNCaP cells.  This method, termed COSMIC-seq, confirms the ability of hairpin-polyamides, with similar architectures but differing at a single ring position, to retain in vitro specificities and display distinct genome-wide binding profiles.  These results underpin the development of Py-Im polyamides as DNA-targeting molecules and the potential for utility as synthetic transcription factors (Syn-TFs) capable of functioning in concert with the cellular regulatory circuitry.

overall_design: To directly map polyamide binding sites across the genome in LNCaP cells, we utilized COSMIC-seq (crosslinking of small molecules to isolate chromatin). COSMIC consists of treating live cells with trifunctional derivatives of polyamides. These molecules were comprised of a DNA-binding polyamide, an affinity handle, and a photocrosslinker. The polyamide confers sequence specificity, while the affinity handle, biotin, is used to purify polyamide–DNA adducts with streptavidin-coated magnetic beads. The photocrosslinker, psoralen, enables the temporally-controlled formation of a covalent bond between the polyamide and DNA. Cells were first treated with the polyamide derivative, crosslinked with 365 nm UV light, lysed, sonicated to shear DNA, and streptavidin-coated magnetic beads were used to capture polyamide-DNA adducts. Because psoralen crosslinks are reversible, the captured genomic DNA can be separated from the polyamide, purified, and identified by massively parallel, next-generation sequencing (NGS). Sequencing reads are mapped to their location across the genome, and loci bound by polyamides show enrichment of sequencing reads relative to a reference input sample.

first: Paul

last: Finn

first: Devesh

last: Bhimsaria

first: Asfa

last: Ali

first: Asuka

last: Eguchi

first: Peter

last: Dervan

middle: Z


title: H2AK121ub in Arabidopsis favors a less accessible chromatin state at transcriptional regulation hotspots [ChIP-seq]

middle: J


title: H2AK121ub in Arabidopsis favors a less accessible chromatin state at transcriptional regulation hotspots [ATAC-seq]


title: Transcriptomic profiling between HSC-3 cells and its highly metastatic subline, HSC-3-M3 cells

status: Public on Dec 01 2020

city: Toyoake

first: Kouhei

last: Sakurai

address: 1-98 Dengakugakubo, Kutsukake-cho

department: Department of Joint Research Laboratory of Clinical Medicine, School of Medicine

institute: Fujita Health University

summary: Metastasis to cervical lymph nodes of oral squamous cell carcinoma (OSCC) leads to a poor prognosis. The present study aimed at investigating the pathways and molecules associated with OSCC metastasis. The transcriptome between HSC-3 cells and their highly metastatic subline, HSC-3-M3 cells, was examined using gene expression microarray.

overall_design: Total RNA was isolated from HSC-3 cells and HSC-3-M3 cells for gene expression profiling.

first: Yuka

last: Ideta


title: Two parallel pathways recruit the H3K9me3 HMT in somatic cells, requiring the Argonaut NRDE-3, or the MBT-domain protein, LIN-61

city: Mainz

first: Jan

last: Padeken

address: Ackermannweg 4

institute: Institute of Molecular Biology

type: Non-coding RNA profiling by high throughput sequencing


title: High-pressure-sprayed double stranded RNA does not induce RNA interference of a reporter gene

city: Neustadt-Mußbach

first: Veli Vural

last: Uslu

state: DEUTSCHLAND

address: Epigenetics, Breitenweg 71

department: Epigenetics

institute: AgroScience GmbH AlPlanta-Institute for Plant Research

summary: We report the sRNA profile upon dsRNA spraying on 16C (Nicotiana benthamiana) plants. In this experiment we could detect only the degradation products of the dsRNA, which indicates absence of further processing of the dsRNA by RNAi machinery

overall_design: Examination of 139nt dsRNA-spraying on 16C, WT plants and water-spraying on 16C plants as a control

first: Veli

middle: V

first: M

last: Wassenegger


title: Two parallel pathways recruit the H3K9me3 HMT in somatic cells, requiring the Argonaut NRDE-3, or the MBT-domain protein, LIN-61 (smallRNA-seq)

summary: The establishment and maintenance of chromatin domains shape the epigenetic memory of a cell, with histone H3 lysine 9 methylation defining repressed heterochromatin. We show that in C. elegans the SET-25 (SUV39/G9a) HMT that catalyzes H3K9me1-3, is able to establish repressed domains de novo. We identify here two distinct pathways that recruit SET-25 to its targets. One requires LIN-61 (L3MBTL2), a conserved protein with 4 MBT domains that recognizes H3K9me2 deposited by the HMT MET-2 (SETDB1). The second pathway is MET-2-independent and requires a somatic Argonaut NRDE-3 and 22nt small nuclear RNAs. This NRDE-3 pathway targets ~10% of all SET-25-modified loci genome-wide including intact RNA and DNA transposons. Removal of both pathways in the met-2;nrde-3 double mutant synergistically derepresses transposons in early embryos and elevates embryonic lethality. The redundancy of these pathways illustrates the key role played by chromatin-mediated silencing in protecting the genome against inherent threats.

overall_design: Small RNA purification from early embryos was performed using the Norgen Single Cell RNA purification kit (51800). Total small RNA was treated with 1 mlTAP (Lucigen) for 1h at 37°C, to allow sequencing of all small RNA species. Library preparation including size selection was performed using the QiaSeq miRNA library. Single-end 50-bp reads were generated using an Illumina HiSeq 2500. Adapter sequences were removed using the fastx_clipper and collapsed using the fastx_collapser functions implemented in the FASTX-Toolkit (http://hannonlab.cshl.edu/fastx_toolkit/). The R package QuasR was used to map reads to the C. elegans genome (WS220), normalized to total number of reads and subsequently subseted according to read lengths.

first: Stephen

last: Methot

last: Zeller

first: Colin

last: Delaney

first: Veronique

last: Kalck

first: Susan

last: Gasser


title: Two parallel pathways recruit the H3K9me3 HMT in somatic cells, requiring the Argonaut NRDE-3, or the MBT-domain protein, LIN-61 (RNA-seq)

overall_design: Total RNAseq of RNA isolated from early C. elegans embryos (~1-300 cell stage) in N2 (Bristol), met-2(n4256) III, set-25(n5021) III, met-2(n4256) III;lin-61(tm2649) I,  lin-61(tm2649) I, set-25(n5021) III;lin-61(tm2649) I, met-2(n4256) III;nrde-3(tm1116) X, set-25(n5021) III;nrde-3(tm1116), nrde-3(tm1116)   mutants. Extraction of RNA was performed according to the WormBook protocol (Stiernagle, 2006). Total RNA was purified using RNeasy kit (QIAGEN 74104) including DNase treatment. Depletion of ribosomal RNA was done for 5 ľg of total RNA with Ribo-Zero Margnetic Gold Kit (Epicentre MRGZG12324) and further concentrated with RNA Clean & Concentrator kit (Zymo Research R1015) according to corresponding manufacturers instructions.


title: RNA-sequencing of Control and Bisphenol A (BPA) exposed mice livers.

city: Los Angeles

first: Xia

last: Yang

address: 610 Charles E. Young Drive East

institute: University of California, Los Angeles

summary: To elucidate the molecular perturbations underlying the connection of low-dose prenatal BPA exposure to cardiometabolic diseases, we assessed the liver transcriptome in both male and female mouse offspring exposed to 5ug/kg/day BPA and corn oil (control) during gestation.

overall_design: Liver samples were dissected from mice offspring at 24 weeks and flash frozen then stored at -80C. RNA was extracted using the Qiagen MiRNeasy Mini Kit. There were 4 groups: 1) Male corn oil (control)  2) Male BPA exposed 3) Female corn oil (control) 4) Female BPA exposed.  There were 4 samples per group (Total samples =16).

first: Graciel

last: Diamante


title: Two parallel pathways recruit the H3K9me3 HMT in somatic cells, requiring the Argonaut NRDE-3, or the MBT-domain protein, LIN-61 (ChIP-seq)

overall_design: Libraries were prepared from chromatin IP and input samples using the NEBNext ultra DNA library prep kit for Illumina (NEB # 7370) and the NEBNext Multiplex Oligos for Illumina (NEB # E7335), according to the manufacturers recommendations. No size selection was performed during sample preparation and the libraries were indexed and amplified using 12 PCR cycles, using the recommended conditions. After a final cleanup with Agencourt AmPure XP beads (Beckman # A63881), the library size distribution and concentrations were determined using a BioAnalyzer 2100 (Agilent technologies) and Qubit (Invitrogen) instrument, respectively. The final pools were prepared by mixing equimolar amounts of all individual indexed libraries and then sequenced on a HiSeq 2500 (Illumina) in Rapid mode (Paired-End 50).


title: Pig brain miRNA profiling

city: Balice near Krakow

first: Klaudia

last: Pawlina-Tyszko

address: Krakowska 1

department: Deaprtment of Animal Genomics and Molecular Biology

institute: National Research Institute of Animal Production

summary:
Purpose: The aim of present research was to characterize miRNA profile of the pig brain tissue and identify miRNAs potentially connected with brain functioning.
Methods: miRNA-seq analysis was performed on brain samples collected from 21 male and female pigs belonging to the Polish 990 synthetic line. The miRNA libraries were constructed from total RNA using NEBNext Multiplex Small RNA Library Prep Set for Illumina (New England Biolabs) according to the manufacturer protocol. The quantification of the obtained libraries was performed on a Qubit 2.0 spectrophotometer (Invitrogen, Life Technologies), while a quality control on a TapeStation 2200 instrument (D1000 ScreenTape; Agilent). 100 single-end cycle sequencing was performed on the HiScanSQ platform (Illumina) with the use of TruSeq SR Cluster Kit v3- CBOT-HS and TruSeq SBS Kit v 3 - HS (Illumina). MicroRNA differentially expressed between males and females were identified with the DESeq2 software, while validation was carried out with RT-qPCR.
Results: miRNA-seq approach allowed the identification of 237 known and 286 potentially novel miRNAs. The comparison of miRNA profiles between females and males showed differential expression of 38 microRNAs. Pathway analysis (DIANA mirPath v.3 tool) confirmed that detected miRNAs are engaged in estrogen signaling pathway, prolactin signaling pathway, long-term depression, axon guidance according to KEGG Database, and response to stress, immune system process, catabolic process according to GO.
Conclusions: Obtained results enabled us to characterize the miRNA profile of the pig brain tissue of males and females, with particular emphasis on the influence of sex. Moreover, they can be the basis for future studies in terms of search of candidate miRNAs related with important processes occuring in the brain and its diseases.

overall_design: MicroRNA next generation sequencing of 21 pig brain samples collected from males and females was performed using Illumina HiScan SQ.

first: Maria

last: Oczkowicz


title: H3K27me3-rich genomic regions can function as silencers to repress gene expression via chromatin interactions

city: Singapore

first: Melissa

last: Fullwood

address: 14 Medical Drive

country: Singapore

institute: Cancer Science Institute

summary: Gene repression and silencers are poorly understood. We reasoned that H3K27me3-rich regions (MRRs) of the genome defined from clusters of H3K27me3 peaks may be used to identify silencers that can regulate gene expression via proximity or looping. MRRs were associated with chromatin interactions and interact preferentially with each other. MRR component removal at interaction anchors by CRISPR led to upregulation of interacting target genes, altered H3K27me3 and H3K27ac levels at interacting regions, and altered chromatin interactions. Chromatin interactions did not change at regions with high H3K27me3, but regions with low H3K27me3 and high H3K27ac levels showed changes in chromatin interactions. The MRR knockout cells also showed changes in phenotype associated with cell identity, and altered xenograft tumor growth. MRR-associated genes and long-range chromatin interactions were susceptible to H3K27me3 depletion. Our results characterized H3K27me3-rich regions and their mechanisms of functioning via looping.

overall_design: K562 DMSO/5μM GSK343 ChIP-seq have 2 biological replicates for each type of antibody. HAP1 Wild-type (WT)/EZH2 Knockout (EZH2KO) ChIP-seq have 2 biological replicates for each type of antibody.  K562 DMSO/5μM GSK343 RNA-seq have 2 biological replicates for each condition. K562 DMSO/1μM GSK343 RNA-seq have 3 biological and 3 technical replicates for each condition. HAP1 WT/EZH2KO RNA-seq have 2 biological replicates for each condition. K562 CRISPR Empty Vector (EV)/Knockout (KO) RNA-seq have 2 biological replicates for each CRISPR clone. K562 EV/MRR2-A1 knockout (IGF1KO)ChIP-seq have 2 biological replicates for each type of antibody. 4C experiments have 2 biological replicates except TMCO4 4C, which only has 1 biological replicate.

first: Yichao

last: Cai

first: Ying

last: Zhang

first: Yan

last: Loh

first: Jia

middle: Q

last: Tng

first: Mei

middle: C

last: Lim

first: Zhendong

last: Cao

first: Greg

last: Tucker-Kellogg


title: Epigenome analysis of upper-tract urothelial carcinomas samples and normal adjacent tissue (NAT)

status: Public on Nov 30 2020

city: Houston

first: Hui

last: Yao

state: TX

address: 1515 Holcombe Blvd.

department: Bioinformatics and Computational Biology

institute: MD Anderson Cancer Center

summary: Genome wide DNA methylation profiling of normal and upper-tract urothelial carcinomas tissues. The Illumina Infinium EPIC arrays was used to obtain DNA methylation profiles across approximately 866,091 probes. Samples included 35 upper-tract urothelial carcinomas samples and 8 adjacent normal tissues

overall_design: Bisulphite converted DNA from the 43 samples were hybridised to the Illumina Infinium EPIC Human Methylation Beadchip arrays

first: Gabriel

last: Malouf


title: Homology-mediated inter-chromosomal interactions in hexaploid wheat lead to specific subgenome territories following polyploidization and introgression

city: shanghai

first: yijing

last: zhang

address: 2005 Songhu Road

department: Biochemistry

institute: Fudan University

summary: Polyploidization and introgression are major events driving plant genome evolution and influencing crop breeding. However, the mechanisms underlying the higher-order chromatin organization of subgenomes and alien chromosomes are largely unknown. We probe the three-dimensional chromatin architecture of Aikang 58 (AK58), a widely-cultivated allohexaploid wheat variety carrying the 1RS/1BL translocation chromosome. The regions involved in inter-chromosomal interactions, both within and between subgenomes, have highly similar sequences. Subgenome-specific territories tend to be connected by subgenome-dominant homologous transposable elements (TEs). The alien 1RS chromosomal arm, which was introgressed from rye and differs from its wheat counterpart, has relatively few inter-chromosome interactions with wheat chromosomes. An analysis of local chromatin structures reveals topologically associating domain (TAD)-like regions covering 52% of the AK58 genome, the boundaries of which are enriched with active genes, zinc-finger factor-binding motifs, CHH methylation, and 24-nt small RNAs. The chromatin loops are mostly localized around TAD boundaries, and the number of gene loops is positively associated with gene activity. The present study reveals the impact of the genetic sequence context on the higher-order chromatin structure and subgenome stability in hexaploid wheat. Specifically, we characterized the sequence homology-mediated inter-chromosome interactions and the non-canonical role of subgenome-biased TEs. Our findings may have profound implications for future investigations of the interplay between genetic sequences and higher-order structures and their consequences on polyploid genome evolution and introgression-based breeding of crop plants.

overall_design: We probed the whole-genome chromosomal interactions using HiC along with DNA metholome and transcriptomic data to systematically characterize the 3D structure of bread wheat cultivar AK58.

first: Yijing

first: JiZeng

last: Jia

first: Zhengang

last: Ru

first: Daowen

last: Wang

first: Guangyao

last: Zhao


title: Transcriptome analysis of mouse embrynic fibroblast cells lacking multiple core clock genes

city: Taichung

first: Yi-Ying

last: Chiou

address: No.145, Xingda Rd., South Dist.

department: Graduate Institute of Biochemistry

country: Taiwan

institute: National Chung Hsing University

summary: We established a cell line lacking endougous CRY, PER, and NR1D proteins (CPN_KO).  By comparing the transcriptome of CPN_KO cells and the cells expressing CRY proteins (PN_KO), genes affected by CRY were analyzed.   Parental cells expressing NR1D proteins (CP_KO) were also analyzed to identify genes regulated by NR1D proteins.

overall_design: mRNA-sequencing of MEF cells lacking Cry, Per, Nr1d proteins


title: Single-cell RNA-sequencing of γδ-T cells from peripheral blood and breast tumour microenvironment.

status: Public on Nov 27 2020

first: Nizar

middle: N.

last: Batada

address: Crew Road

institute: Institute for Genetics and Molecular Medicine, University of Edinburgh

summary: γδ-T cells from PBMCs of healthy donors and immune cells in breast tumour microenvironment.

overall_design:
γδ-T cells were extracted from PBMCs of three healthy donors. Cells from donors HD4 and HD5 were pooled at a 1:1 ratio prior to loading onto 10X.
CD45+ cells were extracted from fresh breast tumours of two patients and loaded onto 10X.

Please note that raw data for the 'BC' samples are not provided (more details requested).

first: Katerina

last: Boufea

first: Victor

last: Gonzalez-Huici

first: Marcus

last: Lindberg

first: Stefan

last: Symeonides

first: Olga

last: Oikonomidou


title: Expression profiling with RNA-seq between mTORC1 high and low AML cells

status: Public on Nov 29 2020

city: Boston

first: Toshihiko

last: Oki

state: MA

address: 185 Cambridge Street

department: Center for Regenerative Medicine

institute: Massachusetts General Hospital

summary: We reported the expression profiling between mTORC1 high and low mouse AML cells with MLL-AF9 utilizing a novel fluorescent probe named mVenus-TOSI (TOr-Signal-Indicator). For this purpose, 3 mTOR high (mVenus low) and 4 mTOR low (mVenus high) Mouse AML cells were harvested from 4 mice. The transcriptomic profiles were distinctive and gene ontology analysis revealed that mTORC1 activity was associated with differentially expressed immune response and metabolism related genes. Further, gene signature enrichment analysis (GSEA) validated that the mTORC1 signal signature correlated with mTORC1 activity and that leukemia stem cell (LSC), Myc, and cell cycle related signatures were enriched in mTORC1 high cells.

overall_design: Examining expression profilng between 3 mTORC1 high and 4 mTORC1 low AML cells with RNA-seq

first: Hiroki

last: Kato

first: Francois

last: Mercier

first: David

last: Scadden


title: Physiological effects of water flow induced swimming exercise in seabream Sparus aurata

status: Public on Nov 26 2020

city: Leiden

first: Ron

last: Dirks

address: Sylviusweg 74

country: Netherlands

institute: Future Genomics Technologies BV

summary: A longer on-land rearing period of Gilthead seabream Sparus aurata before transfer to sea-cages would allow the farmer to benefit from exercise-enhanced growth and resilience as induced by increasing water flow in the tanks. In this study, the physiological effects of flow-conditioning were investigated by subjecting large groups of experimental fish to minimal flow or to flow regimes inducing swimming exercise at 1 or 2 Body Length (BL) s-1 for a period of 8 months (Feb.-Oct.) in 1,500 l RAS tanks. Fish size after eight months of flow conditioning was 92 ± 27 g body weight (BW) for fish under minimal flow; 106 ± 24 g BW (+15%) at 1 BL s-1, and 125 ± 27 g BW (+36%) at 2 BL s-1. Although flow enhanced growth linearly with swimming speed, the number of malformed fish was significantly higher for swimming at 2 BL s-1 indicating that the mechanical load imposed by exercise was too high for a developing juvenile. Fish representing the three treatment groups were then used for: (1) a stress challenge netting test and plasma cortisol measurement (baseline, peaking and recovery levels); (2) blood plasma measurements of glucose, triglycerides, lactate, cholesterol, growth hormone (GH) and insulin-like growth factor I (IGF-I), and heart and muscle gene expression of the GH and IGF-I receptors and the muscle transcriptome by deep RNA sequencing (RNAseq). Flow conditioning at 1 BL s-1 provided optimal conditions for growth and stress coping without energetic depletion and morphological deformation. The absence of important differences in plasma GH and IGF-I, and expression levels of the receptors in heart and white skeletal muscle, indicated that other factors may be involved in growth enhancement. RNAseq of the white skeletal muscle showed that transcription regulators play an important role. Also expression of immune genes is strongly up-regulated. Whereas muscle of fish conditioned at 1 BL s-1 shows up-regulated expression of structural genes such as troponins and myosins, muscle of fish conditioned at 2 BL s-1 shows up-regulated expression of skeletal genes instead which may reflect the mechanism behind morphological deformations.

overall_design: Deep-sequencing transcriptome analysis of muscle samples derived from sea bream (sparus aurata) specimens. The control group (group 1, n=3) was exposed to 8-months minimal water flow, group 2 (n=3) was exposed to an 8-months swimming exercise at 1 body length per second and group 3 (n=3) was exposed to an 8-months swimming exercise at two body lengths per second.

first: Arjan

last: Palstra


title: Comparison of genes related with tertiary lymphoid tissue formation between control and diseased model mice

city: Sapporo

first: Md.

middle: Abdul

last: Masum

state: Hokkaido

address: Kita 18, nishi 9

department: Basic Veterinary Sciences

institute: Faculty of Veterinary Medicine

summary: Identification of genes responsible for formation and function of vasculature-associated lymphoid tissue in lupus nephritis mouse model

overall_design: The whole kidney was collected from 3 MRL/MpJ (control) and 3  MRL/MpJ-Fas lpr/lpr (diseasead) female mice at six months of age

first: M

middle: A

first: Osamu

last: Ichiii


title: Genome-scale CRISPR Screen Reveals Essential Biological Process Determining Human Lineage Specification

city: Shanghai

first: yanhua

last: du

address: 227 South Chongqing Road, Huangpu District

department: Shanghai Institute of Immunology

institute: Shanghai Jiaotong University School of Medicine

summary: Human pluripotent stem cells (hPSCs) have an unlimited capacity to self-renewal and generate any somatic cell types in the human body, offer a promising cell resource for disease modeling and transplantation. However, the systematic mechanism leading the conversion of hPSCs into lineage specification, which is the earliest cellular event during human embryonic development, is elusive. Here, combining a lineage-specific reporter with inducible CRISPR/Cas9 system, we conducted a genome-scale screening in hESCs for overall drawing the track of cell fate decision. We defined eight functional modules containing embryonic development, mRNA processing, metabolism process, epigenetic regulation, cell cycle and others, that participate in full to the lineage commitment.

overall_design: We performed genome-wide CRISPR/iCa9 loss-of-function (LOF) screen in hPSCs with lineage reporter.

first: Yanhua

last: Du

first: Xiangjie

last: Xu


title: Expression profiling with RNA-seq between mouse AML cells with or without chemotherapy

summary: We reported the expression profiling between mouse AML cells with or without chemotherapy. For this purpose, mouse AML cells were harvested from mice treated with or without chemotherapy drugs on day 7. Gene expression analysis of AML cells with or without chemotherapy identified clearly distinguishable transcriptomic profiles. GO analysis revealed that gene alterations after chemotherapy were related to immune response, RNA biology and several metabolomic processes. mTORC1 signal-related GSEA signatures were also enriched in AML cells after chemotherapy including those of stemness and impaired differentiation.

overall_design: Examining expression profilng bwtween the AML cells with or without chemothrapy


title: A functional connection between translation elongation and protein folding at the ribosome exit tunnel in Saccharomyces cerevisiae

status: Public on Nov 25 2020

city: Solna

first: Vicent

last: Pelechano

address: Nobels väg 16

department: MTC

country: Sweden

institute: ScilifeLab - Karolinska Institutet

summary: Proteostasis is a fundamental network of cellular pathways that ensures the optimal concentration and composition of correctly folded proteins within cells in normal and stress conditions. Among key components of this network are the molecular chaperones, which mediate protein folding but also act as modulators of protein synthesis. We have reported on a functional link between translation and de novo folding of proteins in the yeast Saccharomyces cerevisiae by uncovering a specific synthetic-lethal interaction between apparent unrelated mutant variants, the uL3[W255C] variant of the ribosomal protein uL3 and the null mutants of Zuo1 and Ssz1. Zuo1 and Ssz1 are components of the chaperone system named as ribosome-associated complex. Here, we performed a genome-wide analysis of ribosome dynamics by 5PSeq (Pelechano et al. 2015 PMID 2604644) in strains harbouring either wild-type uL3 or mutant uL3[W255C] in the presence or absence of Zuo1 or Ssz1. This method allows the study of ribosome dynamics, by sequencing 5’ phosphorylated mRNA co-translational degradation intermediates. Our results indicate that the rpl3[W255C] mutant is slightly impaired in translation elongation, defect that is significantly enhanced when combined with the deprivation of either Zuo1 or Ssz1.

overall_design: HT-5Pseq was performed with 3 replicates each for 6 strains of S. cerevisiae.

last: Rodríguez-Galán

first: Juan

last: García-Gómez

first: Iván

last: Rosado

first: Wu

last: Wei

first: Alfonso

last: Méndez-Godoy

first: Benjamin

last: Pillet

first: Lars

last: Steinmetz

first: Dieter

last: Kressler

first: Jesús

last: de la Cruz

first: Alisa

last: Alekseenko


title: Characterization of the immunologic impact of sarcoidosis in peripheral blood mononuclear cells via single-cell RNA-seq

city: Oklahoma City

first: Courtney

last: Montgomery

state: OK

address: 825 NE 13th St

department: Genes and Human Disease Research Program

country: USA

institute: Oklahoma Medical Research Foundation

summary: Sarcoidosis is a systemic inflammatory disease characterized by infiltration of immune cells into granulomas. Previous gene expression studies using heterogeneous cell mixtures lack insight into cell-type-specific immune dysregulation. We performed the first single-cell RNA-sequencing study of sarcoidosis in peripheral immune cells in 48 patients and controls. Following unbiased clustering, differentially expressed genes were identified for 18 cell types and bioinformatically assessed for function and pathway enrichment. Our results reveal persistent activation of circulating classical monocytes with subsequent upregulation of trafficking molecules. Specifically, classical monocytes upregulated distinct markers of activation including adhesion molecules, pattern recognition receptors, and chemokine receptors, as well as enrichment of immunoregulatory pathways HMGB1, mTOR, and ephrin receptor signaling. Predictive modeling implicated TGFβ and mTOR signaling as drivers of persistent monocyte activation. Additionally, sarcoidosis T cell subsets displayed patterns of dysregulation. CD4 naïve T cells were enriched for markers of apoptosis and Th17/Treg differentiation, while effector T cells showed enrichment of anergy-related pathways. Differentially expressed genes in regulatory T cells suggested dysfunctional p53, cell death, and TNFR2 signaling. Using more sensitive technology and more precise units of measure, we identify cell-type specific, novel inflammatory and regulatory pathways. Based on our findings, we suggest a novel model involving four convergent arms of dysregulation: persistent hyperactivation of innate and adaptive immunity via classical monocytes and CD4 naïve T cells, regulatory T cell dysfunction, and effector T cell anergy. We further our understanding of the immunopathology of sarcoidosis and point to novel therapeutic targets.

overall_design: We performed  single-cell RNA-sequencing study of sarcoidosis in peripheral immune cells in 35 patients and 13 controls. Cell capture was performed using the 10x Genomics Chromium system followed by sequencing on an Illumina HiSeq 3000, each according to the respective manufacturer’s protocol. Briefly, peripheral blood mononuclear cells (PBMCs) and neutrophils were collected using LSM (Gibco) and Polymorphprep™ (Alere Technologies, Oslo, Norway). Fresh PBMCs (target 4,000/patient) and neutrophils (target 1,000/patient) were combined and loaded at a concentration of 1000 cells/µL into a 10x Genomics Single Cell A v2 chip into the Chromium controller. Following single cell emulsion generation, uniquely identifiable first strand template cDNA was generated by emulsion PCR and after emulsion was broken, the second cDNA strand was generated and Illumina compatible adapters were ligated. Following qPCR quantification, final libraries were loaded onto single respective lanes of a HiSeq 3000 using read lengths of 26 bp for the first read, 98 bp for the second read, and an 8 base index read. Upon completion of sequencing the raw bcl files were processed using the 10x Genomics Cell Ranger (v3.0.2) informatics pipeline.

first: Lori

last: Garman

first: Richard

last: Pelikan

first: Stuart

middle: B

last: Glenn


title: Transcriptome profiling following focal ischemic stroke in female rats

city: Calgary

first: Simona

middle: Denise

last: Frederiksen

state: Alberta

address: 3330 Hospital Dr NW

country: Canada

institute: University of Calgary

summary: Background. Women account for 60% of all stroke deaths and are more often permanently disabled than men, despite their higher observed stroke incidence. Considering the clinical population affected by stroke, an obvious drawback is that many pre-clinical and clinical studies only investigate young males. To improve therapeutic translation from bench to bedside, we believe that it is advantageous to include both sexes in experimental models of stroke. The aims of this study were to identify early cerebral vascular responses to ischemic stroke in females, compare the differential gene expression patterns with those seen in males, and identify potential new therapeutic targets. Results. Transient middle cerebral artery occlusion (tMCAO) was used to induce stroke in both female and male rats, the middle cerebral arteries (MCAs) were isolated 3 hours post reperfusion and RNA was extracted. Affymetrix whole transcriptome expression profiling was performed on female (n = 12) MCAs to reveal differentially expressed genes. In total, 1076 genes had an increased expression and 879 genes a decreased expression in the occluded MCAs as compared with the control MCAs from female rats. An enrichment of genes related to apoptosis, regulation of transcription, protein autophosphorylation, inflammation, oxidative stress, and tissue repair and recovery were seen in the occluded MCA. The high expression genes chosen for qPCR verification (Adamts4, Olr1, JunB, Fosl1, Serpine1, S1pr3, Ccl2 and Socs3) were all shown to be upregulated in the same manner in both females and males after tMCAO (p < 0.05; n = 23). When comparing the differentially expressed genes in female MCAs (occluded and non-occluded) with our previous findings in males after tMCAO, a total of 297 genes overlapped (all groups had 32 genes in common). Conclusions. The cascades of processes initiated in the vasculature following reperfusion are complex. Dynamic gene expression alterations were observed in the occluded MCAs, and to a less pronounced degree in the non-occluded MCAs. Dysregulation of inflammation and blood-brain barrier breakdown are possible pharmacological targets. The sample of genes (<1% of differentially expressed genes) validated for this microarray did not reveal any sex differences.

overall_design: The study was designed as a case-control study with the occluded and non-occluded MCAs (after tMCAO) as case groups (n = 6, 11 samples) and the MCAs from healthy rats (non-operated) as control group (n = 6, 6 samples), and the primary study outcome was differential gene expression (gene expression in cases relative to controls).

first: Mimmi

last: Rehnström

middle: D

first: Saema

last: Ansar

last: Edvinsson


title: Clonal diversity of mTORC high and low AML before and after chemotherapy

first: Toshihiko

last: Oki

summary: There are two possible mechanisms for the shift of gene expression to a high mTORC1 activity signature after chemotherapy; one is the selection of mTORC1 high cells and the second is the induction of mTORC1 activity following chemotherapy.  We assessed these two possible scenarios by introducing genetic barcodes using the ClonTracer library that allowed for clonal tracking of ~0.2 million AML cells. We found that a significant proportion of unique barcodes are represented in both the mTORC1 high and low cell populations. Since barcodes only infect a single cell which has to exist in a high or low state, a significant proportion of barcodes that are represented by cells in both states suggests that induction occurs. Otherwise the proportion should be close to an extreme (0 or 1) meaning most of the populations exist only in mTORC1 high or mTORC1 low states. Additionally, clonal abundance based on barcodes revealed more similarity with respect to the cumulative proportion of represented barcodes between mTORC1 high and low cells in treated cohorts compared with non-treated cohorts. These data suggest a more similar distribution of mTORC1 high and low cells in these populations which would arise primarily due to induction rather than selection.

overall_design: Clonal abundance between four hundred thousand mTORC1 high (mVenus low) or mTORC1 low (mVenus high) AML cells harvested from the mouse bone marrow  before (day 0) or at either of the 3 different time after chemotherapy (day 5, 7, and 14) were compared.

first: Francois

last: Mercier

first: Hiroki

last: Kato

first: Thomas

middle: O

last: McDonald

first: Franziska

last: Michor

first: David

middle: T

last: Scadden


title: Transcription Factor GATA2 Promotes Chromatin Remodeling at the Super-Enhancers of the Key Mast Cell Identity Genes and Primes Enhancers to Respond to Antigenic Stimulation

city: Denver

first: Hua

last: Huang

state: CO

address: 1400 Jackson St

department: Biomedical Research

institute: National Jewish Health

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

overall_design: Refer to individual Series