title: MicroRNA expression profile in response to CDM-3008 treatment in human hepatocytes.

status: Public on Apr 26 2024

city: Wako

first: Luc

last: Gailhouste

state: Saitama

address: 408 Main Research Building, 2-1 Hirosawa

department: Cluster for Pioneering Research

institute: RIKEN

summary: Chronic hepatitis B virus (HBV) infections represent a major global health burden requiring effective therapeutic interventions. This study investigates the antiviral potential of microRNAs (miRNAs) targeting the HBV entry receptor, sodium-taurocholate cotransporting polypeptide (NTCP). Using an experimental model of primary human hepatocytes (PHHs), we highlighted a set of candidate antiviral miRNAs induced by interferon (IFN) alpha analog treatment. Notably, predictive analysis identified miR-29b-1-5p as interacting with the 3’-untranslated region (3’-UTR) of NTCP, suggesting a post-transcriptional regulatory mechanism. Functional analysis indicated that miR-29b-1-5p directly targeted the NTCP 3’-UTR, leading to significant inhibition of NTCP transcripts. Consistently, hepatocytes overexpressing miR-29b-1-5p showed a remarkable reduction in HBV genome levels after infection. A rescue assay demonstrated that miR-29b-1-5p anti-HBV effect was specifically mediated by NTCP targeting. In summary, these findings underscore the therapeutic potential of miR-29b-1-5p against HBV, advocating for further exploration of miRNA-based therapies in the treatment of human viral infections.

overall_design: To identify miRNAs with potential anti-HBV activity, primary human hepatocytes were treated with 10 µM of the IFN alpha analog CDM-3008 for 8 hours, and total RNA was extracted for miRNA microarray analysis.


title: Sequence and parent-of-origin dependent m6A contribute to allele-specific gene expression [GLORI]

city: Guangzhou

first: Ying

last: Zhang

state: Guangdong

address: No. 135, Xingang Xi Road

country: China

institute: Sun Yat-sen University

summary: Multiple regulatory layers influence allele-specific expression (ASE), particularly through sequence-dependent and parent-of-origin-dependent mechanisms at the transcriptional level. However, little is known about allele-specific gene regulation at the post-transcriptional level. Here, we conduct transcriptome-wide analysis of allele-specific m6A in mice. Using early postnatal cerebellum and cerebrum samples from reciprocal crosses of two divergent mouse strains, we employed quantitative m6A assays to measure allelic differences in m6A at single-base resolution. Our study reveals widespread sequence-dependent allelic imbalance in m6A methylation, identifying thousands of allele-specific m6A (ASm6A) sites with statistically significant and reproducible allelic methylation differences across diverse samples. We find evidence of potential cis-regulatory variants within 50-nt flanking regions of these ASm6A sites, with the highest enrichment at the motif positions. Intriguingly, we detect parental effects on allelic methylation across m6A sites exhibiting parent-of-origin-dependent ASE. For both sequence- and parent-of-origin-dependent allelic m6A methylation, we observe opposing allelic preferences between methylation and expression, suggesting a potential role of ASm6A in regulating ASE through negative effects on gene expression. Overall, our findings reveal that both cis-acting and parent-of-origin effects influence ASm6A, offering new insights into post-transcriptional mechanisms of ASE regulation.

overall_design: To identify allele-specific m6A methylation, we carried out reciprocal crosses between two mouse strains with distant genetic backgrounds (PWK/PhJ (PWK) and C57BL/6J (C57)) and conducted GLORI (glyoxal and nitrite-mediated deamination of unmethylated adenosines) assays on early postnatal cerebellum and cerebrum tissues from F1 hybrids. These hybrids were generated through both initial (F1i; PWK mother, C57 father) and reciprocal (F1r; C57 mother, PWK father) crosses.

first: Zeyu

first: Guanzheng

last: Luo


title: Sequence and parent-of-origin dependent m6A contribute to allele-specific gene expression

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

overall_design: Refer to individual Series


title: Sequence and parent-of-origin dependent m6A contribute to allele-specific gene expression [m6A-seq]

overall_design: To identify allele-specific m6A methylation, we carried out reciprocal crosses between two mouse strains with distant genetic backgrounds (PWK/PhJ (PWK) and C57BL/6J (C57)) and conducted multiplexed m6A-seq assays on early postnatal cerebellum and cerebrum tissues from F1 hybrids. These hybrids were generated through both initial (F1i; PWK mother, C57 father) and reciprocal (F1r; C57 mother, PWK father) crosses.


title: METI: Deep profiling of tumor ecosystems by integrating cell morphology and spatial transcriptomics

status: Public on May 23 2024

city: Houston

first: Linghua

last: Wang

address: 1881 East Road,

country: USA

institute: MD Anderson Cancer Center

summary: The recent advance of spatial transcriptomics (ST) technique provides valuable insights into the organization and interactions of cells within the tumor microenvironment (TME). While various analytical tools have been developed for tasks such as spatial clustering, spatially variable gene identification, and cell type deconvolution, most of them are general methods lacking consideration of histological features in spatial data analysis. This limitation results in reduced performance and interpretability of their results when studying the TME. Here, we present a computational framework named, Morphology-Enhanced Spatial Transcriptome Analysis Integrator (METI) to address this gap. METI is an end-to-end framework capable of spatial mapping of both cancer cells and various TME cell components, robust stratification of cell type and transcriptional states, and cell co-localization analysis. By integrating both spatial transcriptomics, cell morphology and curated gene signatures, METI enhances our understanding of the molecular landscape and cellular interactions within the tissue, facilitating detailed investigations of the TME and its functional implications. The performance of METI has been evaluated on ST data generated from various tumor tissues, including gastric, lung, and bladder cancers, as well as premalignant tissues. Across all these tissues and conditions, METI has demonstrated robust performance with consistency.

overall_design: Two muscle-invasive bladder cancer samples (T2-T4) and four gastric adenocarcinoma samples were used for the Visium Spatial transcriptomics.

first: Jiahui

last: Jiang

first: Jian

last: Hu


title: TIM-3 + CD8 T cells with a terminally exhausted phenotype retain functional capacity in hematological malignancies

status: Public on Apr 19 2024

city: Seattle

first: Geoffrey

middle: R

last: Hill

address: 1100 Fairview Ave N

institute: Fred Hutch Cancer Center

type: Expression profiling by high throughput sequencing

summary: Chronic antigen stimulation is thought to generate dysfunctional CD8 T cells. Here we identify a CD8 T cell subset in the bone marrow tumor microenvironment that, despite an apparent terminally exhausted phenotype (TPHEX), expressed granzymes, perforin, and IFNγ. Concurrent gene expression and DNA accessibility revealed that genes encoding these functional proteins correlated with BATF expression and motif accessibility. IFNγ+ TPHEX effectively killed myeloma with comparable efficacy to transitory effectors, and disease progression correlated with numerical deficits in IFNγ+ TPHEX. We also observed IFNγ+ TPHEX within CD19-targeted chimeric antigen receptor T cells, which killed CD19+ leukemia cells. Importantly, an IFNγ+ TPHEX gene signature was recapitulated in TEX cells from human cancers, including myeloma and lymphoma. Here we characterize a TEX subset in hematological malignancies that paradoxically retains function and is distinct from dysfunctional TEX found in chronic viral infections. Thus, IFNγ+ TPHEX represent a potential target for immunotherapy of blood cancers.

overall_design:
Mouse polyclonal CD8 T cells were harvested from BM and were sequenced for gene expression (3'; 1 sample) and for concurrent ATAC and RNA seq (Multi-ome; 3 samples). Mouse CD19 CAR T cells were harvested from BM and sequenced for gene expression and TCR repertoire (5'; 1 sample)
Please note that the processed data for the 'scMO_MM_CD8* samples are linked as Series supplementary files.

first: Simone

last: Minnie

first: Olivia

last: Waltner

first: Scott

last: Furlan


title: Single-cell multiomics profiling reveals heterogeneous transcriptional programs and microenvironment in Desmoplastic Small Round Cell Tumors [ChIP-seq]

status: Public on May 22 2024

city: Villejuif

first: Clémence

last: Henon

address: 114 rue Edouard Vaillant

country: France

institute: Gustave Roussy

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Desmoplastic Small Round Cell Tumor (DSRCT) is a rare, aggressive sarcoma driven by the EWSR1-WT1 chimeric transcription factor. Despite this unique oncogenic driver, DSRCT displays a polyphenotypic differentiation of unknown causality. Using single-cell multi-omics on samples from patients, we find that DSRCT tumor cells cluster into consistant subpopulations with partially overlapping lineage- and metabolism-related transcriptional programs.

overall_design: ChIP-seq was performed in the JN-DSRCT-1 cell line using the GTX15249 WT1 C-terminal antibody or a rabbit isotype control. A second assay was performed in the same cell line after a 48-hour siRNA-mediated silencing using a control non-targeting siRNA or a  EWSR1::WT1 siRNA, using H3K9ac or H3K27ac antibodies in both conditions.

first: Julien

last: Vibert

first: Léo

last: Colmet-Daage

first: Sophie

last: Postel-Vinay


title: Chromatin-focused genetic and chemical screens identify BRPF1 as a targetable vulnerability in Taxol-resistant triple-negative breast cancer [CRISPR]

status: Public on May 14 2024

city: İstanbul

first: Tugba

last: Bagci-Onder

address: Rumelifeneri Yolu, Sarıyer

department: Koç University Research Center for Translational Medicine

country: Turkey

institute: Koç University

summary: Triple-negative breast cancer (TNBC) stands out as a particularly aggressive and frequently recurring form of breast cancer. Due to the absence of hormone receptors, the available treatment avenues are constrained, making chemotherapy the primary approach. Unfortunately, the development of resistance to chemotherapy poses a significant challenge, further restricting the already limited therapeutic alternatives for recurrent cases. Understanding the molecular basis of chemotherapy resistance in TNBC is pivotal for improving treatment outcomes. Here, we generated two different Taxol-resistant TNBC cell lines with dose-escalation method to mimic chemotherapy resistance in vitro. These cells exhibited hallmark features of resistance, including reduced cell growth, altered morphology, and resistance to apoptosis. Transcriptome analysis uncovered elevated ABCB1 expression and multidrug-resistant phenotype in the resistant cells. To comprehensively investigate the key epigenetic regulators of Taxol resistance, we conducted epigenome-wide CRISPR/Cas9 and chemical probe library screens. Both screens pinpointed Bromodomain and PHD Finger Containing 1 (BRPF1) which is the reader protein in MOZ/MORF histone acetyl-transferase complex, as the regulator of Taxol resistance in TNBC cells. Knockout of BRPF1, but not the other members of the MOZ/MORF complex, sensitized resistant cells to Taxol. Additionally, BRPF1 inhibitors, PFI-4 and OF-1, in combination with Taxol significantly reduced cell viability. Transcriptome analysis upon BRPF1 loss or inhibition revealed a negative impact on ribosome biogenesis-related gene sets, resulting in a global decrease in protein translation in Taxol-resistant cells. Our ChIP-qPCR analysis further demonstrated that active BRPF1 directly interacts with the ABCB1 promoter, enhancing its expression and inducing a multidrug-resistant phenotype. Conversely, knockout or inhibition of BRPF1 leads to decreased ABCB1 expression. This dual mechanism critically sensitizes Taxol-resistant TNBC cells to chemotherapy. Our findings uncover a comprehensive molecular framework, highlighting the pivotal role of epigenetic reader protein BRPF1 in Taxol resistance and providing potential avenues for therapeutic intervention in TNBC.

overall_design: To mimic chemotherapy resistance in vitro, we generated Taxol-resistant SUM159PT cell lines, namely T1-160 and T2-450. We first performed RNA-seq to uncover the differentially expressed genes in resistant cells when compared to parental cells. Later, we identified BRPF1 as a regulator of taxol resistance in T1-160 cells through epigenome-wide CRISPR/Cas9 screening and performed validation experiments using inhibitors or knockouts.

first: Ozlem

middle: Y

last: Bayram

first: Ahmet

last: Cingöz

first: Ebru

last: Yilmaz

first: Ali

middle: C

last: Aksu

first: Beril

last: Esin

first: Nareg

last: Pinarbasi-Degirmenci

first: Ayse

middle: D

last: Cavga

first: Beyza

last: Dedeoglu

first: Buse

last: Cevatemre

first: Hamzah

last: Syed

first: Martin

last: Philpott

first: Adam

middle: P

last: Cribbs

first: Udo

last: Oppermann

first: Nathan

middle: A

last: Lack

first: Ceyda

last: Ayhan

first: Tamer

middle: T

last: Önder


title: Transcriptional changes of the extracellular matrix in chronic thromboembolic pulmonary hypertension governs right ventricle remodeling and recovery

status: Public on Apr 18 2024

city: Bad Nauheim

first: Carsten

last: Kuenne

address: Ludwigstrasse 43

department: Bioinformatics

country: Germany

institute: Max Planck Institute for Heart and Lung Research


title: Transcriptional changes of the extracellular matrix in chronic thromboembolic pulmonary hypertension governs right ventricle remodeling and recovery [ext395_rnaseq]

summary:
The present prospective study included a total number of 71 patients with a final diagnosis of chronic thromboembolic pulmonary hypertension (CTEPH), who were treated by pulmonary endarterectomy (PEA) at the Kerckhoff Heart and Thorax Center between 2016 and 2020.
Biopsies of the free right ventricular wall from 71 patients were collected at baseline (BL) during PEA. In a subgroup of 24 patients, myocardial interventricular septum biopsie were obtained during right heart catheterization (RHC) 12 months after PEA (postPEA)
In this case, to account for technical and safety aspects, the specimens were taken from the myocardial interventricular septum.
Patients were grouped in severe, intermediat and moderate risk according to 1 year mortality risk by applying a modified version of the European Society of Cardiology (ESC) guidelines  risk stratification model according to their BL characteristics.
The aim of the Study was: a) A comparative characterization of RNA-profiles in CTEPH patients with disease status at baseline prior to PEA therapy; b): An assessment of dynamics of the RNA-profiles in CTEPH patients pre and post PEA.

overall_design:
Confirmatory cohort:
a): Comparison of RNA profile of CTEPH patients: severe (n=18) , intermediate (n=23), and moderate (n=30) risk groups
b): 22 prePEA specimens with their corresponding postPEA specimens (postPEA vs. prePEA)

first: Christoph

last: Liebetrau

first: Soni

last: Pullamsetti

first: Eckhard

last: Mayer

first: Peter

last: Dorfmüller

first: Leili

last: Jafari

first: Christian

last: Troidl

first: Stefan

last: Guenther


title: Transcriptional changes of the extracellular matrix in chronic thromboembolic pulmonary hypertension governs right ventricle remodeling and recovery [ext128_rnaseq]

summary:
The present prospective study included a total number of 9 patients with a final diagnosis of chronic thromboembolic pulmonary hypertension (CTEPH), who were treated by pulmonary endarterectomy (PEA) at the Kerckhoff Heart and Thorax Center between 2016 and 2020.
Biopsies of theright ventricularfreewall from9patients were collected at baseline (BL) before PEA surgery (prePEA).
Patients were grouped in severe and moderate risk groups according to 1 year mortality risk by applying a modified version of the European Society of Cardiology (ESC) guidelines  risk stratification model according to their BL characteristics.
The aim of the Study was a comparative characterization of RNA-profiles in CTEPH patients with disease status at baseline prior to PEA therapy

overall_design:
Overall design Exploratory Cohort: Comparison of RNA profile of CTEPH patients: severe
(n=5), moderate (n=4), and intermediate (n=5) risk groups.


title: Transcriptional changes of the extracellular matrix in chronic thromboembolic pulmonary hypertension governs right ventricle remodeling and recovery [ext327_rnaseq]

summary:
The present prospective study included a total number of 3 patients with a final diagnosis of chronic thromboembolic pulmonary hypertension (CTEPH), who were treated by pulmonary endarterectomy (PEA) at the Kerckhoff Heart and Thorax Center between 2016 and 2020.
Biopsies of the myocardial interventricular septum from 3 patients were collected at baseline (BL) before PEA surgery (pre-septal-PEA)
In this case, to account for technical and safety aspects, the specimens were taken from the myocardial interventricular septum.
The aim of the Study was a comparative characterization of RNA-profiles of septum in CTEPH patients after PEA surgery (postPEA) compared to before PEA surgery (pre-septal-PEA).

overall_design:
Confirmatory cohort:
Comparison of RNA profile of CTEPH patients: pre-septal-PEA (n=3) with their corrsponding postPEA (n=3) specimens


title: Assessment of chromatin state changes during human cortical interneuron development

city: Saint Louis

first: Kristen

last: Kroll

state: MO

address: 320 McDonnell Sciences/660 S. Euclid Ave.

department: Developmental Biology

institute: Washington University School of Medicine

summary: We have used our protocol for generating cortical interneurons from human embryonic stem cells to study chromatin state changes during cortical interneuron development. This allows for the identification of cis-regulatory elements and transcription factors which play important roles in this process. Samples were collected at day 0 (hESCs), day 15 (ventral telencephalic patterned medial ganglionic eminence-like (MGE) progenitors), day 35 (immature interneurons), and day 60 (mature interneurons). Day 15 dorsal telencephalic cortical-like neural progenitors were also obtained by using a dual Smad inhibition protocol for comparison with ventral telencephalic MGE-like progenitors.

overall_design: For each time point and histone modification (H3K27ac, H3K4me3, H3K27me3 and H3K9me3), 3-6 biological replicates were generated from independant differentiations of hESCs

first: Gareth

last: Chapman


title: Effect of RFX3 or RFX4 knockdown on ventral telencephalic medial ganglionic eminence-like (MGE-like) neuronal precursors

summary: RFX3 and RFX4 were defined as transcription factors (TFs) with enriched TF binding sites in cis-regulatory elements that are activated during the conversion of human embryonic stem cells (hESCs) into ventral telencephalic MGE-like patterned neuronal precursors. Therefore, we conducted CRISPRi-mediated knockdown of RFX3 or RFX4 and profiled changes in gene expression in MGE-like ventral telencephalic progenitors by RNA-seq. This analysis revealed that RFX3 and RFX4 play important roles in cortical interneuron development, specifically during the transition from mitotically cycling neuronal precursors into post mitotic neurons.

overall_design: Clonal hESC populations were transduced with plasmids carry dCas9-KRAB alone or in combination with gRNA for RFX3 or RFX4 and for each biological condition there are 4 biological replicates generated from indipendant differentiations


title: Assessment of gene expression changes during human cortical interneuron development

summary: We have used our protocol for generating cortical interneurons from human embryonic stem cells (hESCs) to study gene expression changes during this process, to identify regulatory networks critical to cortical interneuron development. Samples were collected at day 0 (hESCs), day 15 (ventral telencephalic patterned medial ganglionic eminence-like progenitors), day 35 (immature interneurons), and day 60 (mature interneurons).

overall_design: For each time point, 3-4 biological replicates generated from independant differentiations of hESCs were used for transcriptomic analysis by RNA-sequencing.


title: Genetics, energetics, and allostery in proteins with randomized cores and surfaces

status: Public on May 13 2024

city: Barcelona

first: Albert

last: Escobedo

state: Catalonia

address: Carrer Dr Aiguader 88

department: Systems and synthetic biology

country: Spain

institute: CRG

summary: A lack of systematic experimental data limits our understanding of protein evolution. In this study, we experimentally characterized proteins with randomized sequences. Vast numbers of amino acid combinations constitute stable protein cores and surfaces. However, alternative cores frequently disrupt protein function by indirect allosteric effects. Both protein stability and binding can be predicted by using simple additive energy models with a small contribution from pairwise energetic couplings. Indeed, energy models trained on one protein can predict functional cores and surfaces across more than a billion years of evolution, with only rare energetic couplings that we experimentally identify limiting the transplantation of cores between highly diverged proteins. Our results reveal the simple energetic architecture of proteins and suggest that allostery is an important constraint on sequence evolution.

overall_design: We built combinatorial libraries in the hydrophobic cores of three small protein domains (FYN-SH3, CI-2A and CspA) using a reduced alphabet consisting of the amino acids F, L, M, V, I encoded by the DTS degenerate codon. By bottlenecking and pooling the libraries, in the sparse_DTS_core_mutagenesis experiment we sparsely measured the intracellular abundance of protein variants in yeast cells using abundancePCA, a protein complementation assay that couples cell growth rate with query protein intracellular abundance under selection by methotrexate. For the SH3 domain of the human FYN kinase, we also randomized 15 buried and 21 exposed sites using reduced amino acid alphabets that were optimized ad-hoc for each site (FYN-SH3_buried_n_exposed_randomization experiment). Also for FYN-SH3, we selected a few query core amino acid combinations that are severely deleterious in abundance fitness and designed a suppressor "permissivity" library by introducing non-core mutations associated with SH3 domains naturally carrying such query core combinations that are deleterious in FYN (FYN-SH3_core_permissivity experiment). For the same protein (FYN-SH3), we assessed the impact of core reconfiguration in function by measuring the binding to its short linear motif ligand PRD1super using bindingPCA, a protein complementation assay that couples cell growth rate with query variant intracellular binding to an interacting partner under selection by methotrexate (FYN-SH3_core_DTS_binding experiment).

first: Gesa

last: Voigt

first: Andre

middle: J

last: Faure

first: Ben

last: Lehner


title: Regulation of Microprocessor assembly and localization via Pasha’s WW domain in C. elegans

status: Public on Apr 16 2024

city: Fort Collins

first: Taiowa

last: Montgomery

state: CO

address: 1878 Campus Delivery

department: Department of Biology

institute: Colorado State University

type: Non-coding RNA profiling by high throughput sequencing

summary: Primary microRNA (pri-miRNA) transcripts are processed by a protein complex called the Microprocessor comprised of the ribonuclease Drosha and its RNA binding partner DGCR8/Pasha. We sequenced small RNAs from animals containing a mutation in the WW domain of C. elegans pash-1. We found that these mutants have a modest but widespread reduction in miRNA levels when grown at 20˚C, which is further enhanced when grown at 25˚C. The results demonstrate a requirement for the WW domain in processing miRNAs in C. elegans.

overall_design: Small RNA high-throughput sequencing was done using RNA isolated from wild type and pash-1(ram33)mutant whole gravid adult C. elegans grown at 20˚C or 25˚C. The tinyRNA pipeline was used to quantitatively assess miRNA levels.


title: Stress granule assembly impairs macrophage efferocytosis [RNA-seq]

status: Public on May 01 2024

city: Shanghai

first: Ye

last: Zhou

address: Xiangyin Road

department: National Key Laboratory of Immunity & Inflammation

institute: Naval Medical University

summary: Stress granules (SGs) assembly in response to various stress, has been demonstrated in the regulation of anti-viral immune response and tumor progression. However, lack of evidence to illustrate the relation between SGs formation and allergic diseases. Applying RNA-seq in primary macrophages  from G3bp1f/f and G3bp1mac-/- mice, differential expressed genes were defined.

overall_design: RNA-seq is undertaken to investigatethe differential expressed genes in primary macrophages between G3bp1f/f and G3bp1mac-/- mice.

first: Zhou

last: Ye

first: Yang

last: Zixuan

first: Hou

last: Jin


title: Single-cell multiomics profiling reveals heterogeneous transcriptional programs and microenvironment in Desmoplastic Small Round Cell Tumors [snMultiome]

status: Public on Apr 15 2024

first: Clémence

last: Henon

overall_design:
Fresh tumor material from one patient with DSRCT was profiled using the single-cell Multiome ATAC + Gene Expression 10x Genomics Chromium assay.

***************************************************************
Raw data were not submitted due to privacy concerns.
***************************************************************

first: Julien

last: Vibert

first: Léo

last: Colmet-Daage

first: Sophie

last: Postel-Vinay


title: Single-cell multiomics profiling reveals heterogeneous transcriptional programs and microenvironment in Desmoplastic Small Round Cell Tumors


title: Single-cell multiomics profiling reveals heterogeneous transcriptional programs and microenvironment in Desmoplastic Small Round Cell Tumors [scRNA-Seq]

overall_design:
Fresh tumor material from patients with DSRCT was profiled using the 3’-end scRNA-seq 10x Genomics Chromium assay.

***************************************************************
Raw data were not submitted due to privacy concerns.
***************************************************************


title: SRRM2 splicing factor modulates cell fate in early development [RNA-Seq 2]

status: Public on Apr 12 2024

city: Berlin

first: Alexander

last: Kukalev

address: Hannoversche Str., 28

department: Berlin Institute for Medical Systems Biology

institute: Max-Delbrueck Center for Molecular Medicine

summary: Srrm2 splicing factor is a novel gene implicated in developmental disorders and diseases. However, the role of Srrm2 in early mammalian development remains unexplored. Here, we show that Srrm2 expression dosage is critical for maintaining embryonic stem cell pluripotency and cell identity. Srrm2 heterozygosity promotes loss of stemness characterized by the coexistence of cells expressing naive and formative markers, together with large gene expression shifts, including in serum-response transcription factor targets and differentiation-related genes. Depletion of Srrm2 by RNA interference in embryonic stem cells identified splicing misregulation of specific genes, often linked to exon skipping. These results show that Srrm2 dosage is key in controlling stemness and cell fate decisions. Our findings unveil Srrm2’s molecular and cellular implications in development, shedding light on the involvement of splicing regulators in early embryogenesis, developmental diseases and tumorigenesis.

overall_design: To investigate the role of Srrm2 levels in pluripotency, we obtain mouse embryonic stem cells (mESC) E14tg2a.4 Srrm2 heterozygous knockout and mESC E14tg2a.4 parental wild-type cells from MMRC UCDavis. We then performed gene expression profilling analysis from total RNA-seq of 3 replicates per condition.

first: Silvia

last: Carvalho

first: Tsz Ching Chloe

last: Tang

first: Paulo

last: Caldas

first: Ana

last: Grosso

last: Pombo


title: SRRM2 splicing factor modulates cell fate in early development [scRNA-Seq]

overall_design: Whole-cell 10x Genomics single cell RNA samples were collected from mouse embryonic stem cells (mESC) E14tg2a.4 WT and Srrm2 heterozygous knockout, in 2 independent experiment for 2 replicates of each cell line.

first: Luna

last: Zea Redondo


title: SRRM2 splicing factor modulates cell fate in early development [RNA-Seq]

overall_design: To investigate the acute role of Srrm2 depletion in mouse embryonic stem cells (mESC), we treated mESC E14tg2a.4 with RNA interference targetting Srrm2 and a control region (GL2). siRNA-treated samples were collected 24 or 72 h upon treatment, in three independent experiments. We then performed gene expression profilling analysis by mRNA-seq of 3 replicates for each condition.


title: Immuno-Oncologic Profiling by Stage-Dependent Transcriptome and Proteome Analyses of Spontaneously Regressing Canine Cutaneous Histiocytoma [RNA-Seq]

status: Public on Apr 10 2024

first: Alina

middle: Katharina

last: Loriani Fard

address: Robert-von-Ostertag Straße 15

department: Department of Veterinary Medicine

institute: Freie Universität Berlin

summary: Canine cutaneous histiocytoma (CCH) is a tumor originating from dermal Langerhans cells, especially affecting young dogs. The common spontaneous regression of CCH makes it an interesting model in comparative oncology research. Previous studies have indicated that anti-tumor immune responses may be involved but details remain speculative to date. Here, we asked which specific immuno-oncological dynamics underlie spontaneous regression of CCH on mRNA and protein levels. QuantSeq 3' mRNA sequencing with functional overrepresentation analysis and an nCounter RNA hybridization assay were employed on 21 formalin-fixed, paraffin-embedded CCH samples representing three different tumor stages. Nine additional samples were subjected to matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI). Surprisingly, only minor stage-specific differences were found. When we investigated expression of B7 family ligands and CD28 family receptors holding co-stimulatory and -inhibitory functions respectively, we found higher abundance of CD80, CD86 and CD28, which trigger activation of lymphocyte-mediated immune responses. CD80 and CD86 expressing cells were further quantified by in situ hybridization and compared with data from three cases of canine histiocytic sarcoma (HS), a malignant tumor variant originating from antigen presenting cells. A stage-specific increase of CD80 expression was recorded in CCH from the tumor bottom to the top, while CD86 was continuously and homogenously expressed at high levels. Expression of CD80 in CCH was similar to that in HS (73.3 ± 37.4% versus 62.1 ± 46.4% positive cells), while CD86 was significantly higher expressed in CCH (94.7 ± 10.3%) when compared to HS (57.6 ± 11.0%). Our data suggest that major immuno-oncological pathways are seemingly not regulated during regression of CCH on the mRNA or protein levels as detectable by the methods used. We speculate that key signals leading to tumor regression may occur at an interactome level or at an earlier time point than examined here. Our data further supports a role of immune stimulatory B7 family ligands and CD28 family receptors in CCH regression.

overall_design: To investigate stage-dependent differences of canine cutaneous histiocytoma (CCH) at the transcriptome level, RNA was isolated from formalin-fixed, paraffin-embedded (FFPE) material. Afterwards gene expression profiling was performed using RNA-Seq data from 4 different groups. Tumor cell-enriched groups (2a vs. 1) and lymphocyte-enriched groups (3 vs. 2b) were compared.

middle: K

middle: F

last: Haake

first: Vladimir

middle: M

last: Jovanovic

first: Sandro

last: Andreotti

first: Michael

last: Hummel

first: Benjamin-Florian

last: Hempel

first: Achim

last: Gruber


title: OCA-B/Pou2af1 promotes autoimmune demyelination through control of stem-like CD4+ T cells [Relapse scRNAseq]

status: Public on Apr 09 2024

city: Salt Lake City

first: Erik

middle: Peter

last: Hughes

state: Utah

address: 2000 Cir of Hope Dr.

department: Pathology

institute: University of Utah

summary: The identification of druggable therapeutic targets that preferentially promote autoimmunity remains a key goal in the field. Recent work shows that populations of memory/stem-like T cells drive autoimmunity, but the factors that generate and sustain these populations are incompletely understood. The lymphocyte-restricted transcriptional cofactor OCA-B/Pou2af1 is a regulator of CD4+ T cell memory. Here we show that T cell-intrinsic loss of OCA-B protects mice from experimental autoimmune encephalomyelitis (EAE) in both chronic and relapsing-remitting mouse models while preserving acute responses to infection with a neurotropic coronavirus. In adoptive transfer EAE driven by antigen re-encounter, T cell-specific OCA-B loss largely eliminates Th1- and Th17-mediated CNS infiltration, proinflammatory cytokine production and disease. Using an OCA-B-mCherry reporter mouse, we show that OCA-B expressing CD4+ T cells within the CNS of mice with EAE preferentially display a memory-like phenotype. Transferring these OCA-Bhi memory-like CD4+ T cells preferentially confers disease, identifying OCA-B as a marker of encephalitogenic autoreactive CD4+ T cells. Notably, in a relapsing-remitting EAE model, OCA-B T cell-deficient mice show specific protection at relapse, highlighting the potential to target OCA-B in MS patients to limit disease progression. During remission, OCA-B promotes the expression of Tcf7, Slamf6, and Sell in proliferating T cell populations. At relapse, OCA-B loss results in the accumulation of an immunomodulatory CD4+ T cell population expressing Ccr9 and Bach2. These results highlight OCA-B as a driver of pathogenic stem-like T cells responsible for relapse and promising MS therapeutic target.

overall_design: NOD Ocab(fl/fl) and Ocab(fl/fl);CD4-Cre mice were induced with EAE. At disease relapse, brain and spinal cord CD3e+ cells were isolated by FACS and processed for single cell RNA sequencing.

first: Amber

last: Syage

first: Elnaz

last: Mirzaei Mehrabad

first: Thomas

middle: E

last: Lane

first: Benjamin

last: Spike

first: Dean

last: Tantin


title: Macrophages THP-1: Control vs Colpvp

status: Public on Apr 01 2024

city: Mexico City

first: Krötzsch

last: Edgar

address: Calz México-Xochimilco 289, Coapa, Col. Arenal de Guadalupe, Tlalpan

country: Mexico

institute: Instituto Nacional de Rehabilitación LGII

summary: Transcriptional array of macrophages cells comparing control untreated vs colpvp treated for 24hrs

overall_design: Two-condition experiment, control  vs colpvp cells. Single samples, one replicate per array

first: Edgar

last: Krotzsch

first: Hugo

last: Mendieta-Zerón

first: Estaban

last: Cruz-Arenas

first: Salvador

last: Díaz-Meza

first: Alejandro

last: Cabrera-Wrooman

first: Edna

last: Mandujano-Tinoco

first: Daniel

last: Muñiz-García

first: Laura

last: Orozco-Castañeda

first: Sonia

last: Hernández-Enríquez

first: Miriam

last: Rodríguez-Piña

middle: S

last: Mulia-Soto

first: José

last: Meneses-Calderón

first: Rosa

last: Salgado

first: Paul

last: Mondragón-Terán


title: Fibroblasts hTERT-BJ1: Control vs Colpvp

summary: Transcriptional array of fibroblasts comparing control untreated vs colpvp treated for 24hrs

first: Ana


title: OCA-B/Pou2af1 promotes autoimmune demyelination through control of stem-like CD4+ T cells [Remission scRNAseq]

first: Erik

last: Hughes

overall_design: NOD Ocab(fl/fl) and Ocab(fl/fl);CD4-Cre mice were induced with EAE. At disease remission, brain and spinal cord CD3e+ cells were isolated by FACS and processed for single cell RNA sequencing.

first: Amber

last: Syage

first: Elnaz

last: Mirzaei Mehrabad

first: Thomas

last: Lane

first: Benjamin

last: Spike


title: Immuno-Oncologic Profiling by Stage-Dependent Transcriptome and Proteome Analyses of Spontaneously Regressing Canine Cutaneous Histiocytoma [nCounter]

first: Alina

last: Loriani Fard

first: Alexander

last: Haake

first: Vladimir

last: Jovanovic

first: Sandro

last: Andreotti

first: Michael

last: Hummel

first: Benjamin-Florian

last: Hempel

first: Achim

last: Gruber


title: Strong Correlation of Gene Counts and Differentially Expressed Genes Between a 3′ RNA-Seq and RNA Hybridisation Platform in Transcriptome Analyses from Canine Archival Tissue [RNA-Seq]

status: Public on Mar 21 2024

summary: 3' RNA-Seq and direct RNA hybridisation are conceptually different approaches for high throughput transcriptome analysis. The former generates next-generation sequencing libraries by targeting and amplifying 3'-end sequences of 60 to 80 nucleotides in length, whereas the latter quantifies transcripts of 800 pre-selected genes. Since both methods are applicable to short mRNA fragments, they are thought to allow for transcriptome analyses from formalin-fixed, paraffin-embedded (FFPE) archival tissue containing partially degraded RNA. In this study, results of both approaches were compared on sample- and gene-wise count levels, gene expression strength and direction, as well as the overlap of differentially expressed genes (DEGs). Using two different oncological perspectives, i.e. a stage-dependent and entity-contrasting comparison on 35 FFPE canine tumours, both methods proved suitable for their use on archival tissues. A moderately to very strong overall count correlation was found (range of Pearson and Spearman means: 0.66 – 0.87). Of note, the gene-wise count correlations depended on gene expression strength. Finally, in the entity-contrasting comparison, expression direction correlated very strongly (range: 0.88 – 0.91) but DEGs only moderately overlapped (Jaccard index: 0.53). Aside from these figures, different practically relevant aspects of the two technologies offer distinct advantages, depending on the objectives and design of a study.

overall_design:
To determine the correlation of gene expression data generated by the nCounter® Canine IO Panel and QuantSeq 3' from three types of canine tumours. FFPE tissue samples were obtained from the archive (spanning the years 2013 to 2021) and total RNA was extracted. After QuantSeq 3' and hybridisation to the nCounter® Canine IO Panel with bioinformatic processing, counts and differential gene expression data were compared to calculate correlations between the two technologies employed.

Attention: This GEO submission includes the 3' RNA-Seq data from the stage-contrasting comparison (n = 10). The data for this comparison from the RNA hybridisation platform (nCounter®) was submitted separately to GEO (262020). The other entity-contrasting tumour samples (n = 25) used in the publication were also submitted separately to GEO (nCounter®: 261791; 3' RNA-Seq: 261790).


title: Spatially preserved multi-region transcriptomic subtyping and biomarkers of outcome with chemoimmunotherapy in extensive-stage small cell lung cancer [CANTABRICO_DSP cohort]

city: Madrid

first: Jon

last: Zugazagoitia

address: Avenida de Cordoba s/n

department: Medical Oncology

institute: Hospital Universitario 12 de Octubre

summary: Transcriptomic subtyping holds promise for personalized therapy in extensive stage small cell lung cancer (ES-SCLC). In this study, we aimed to assess intratumoral transcriptomic subtype diversity and to identify biomarkers associated with long-term chemoimmunotherapy benefit in human ES-SCLC. Our work highlights that high intratumoral heterogeneity, lack of consistent association with outcome, and unclear subtype-specific target expression are major challenges for SCLC subtype-based precision oncology. Pre-existing IFNϒ-driven immunity and mitochondrial metabolism seem key correlates of long-term efficacy for chemoimmunotherapy in ES-SCLC.

overall_design:
We used tumor samples from 26 ES-SCLC patients enrolled in the CANTABRICO clinical trial (EudraCT Number: 2020-002328-35) with adequate tumor material for spatially preserved transcriptomic analysis. The CANTABRICO study was a phase IIIb single arm multicenter trial evaluating the safety and effectiveness of PD-L1 blockade (durvalumab) in combination with platinum-etoposide as first-line treatment in patients with ES-SCLC.
We retrieved pre-treatment, formalin-fixed and paraffin-embedded (FFPE), tumor specimens, which were centrally reviewed by an expert pathologist at the 12 de Octubre research facilities (Madrid, Spain).
Spatial mRNA profiling was done using the GeoMx Digital Spatial Profiling (DSP) system (NanoString). For this study, we used an mRNA assay that contained oligo-nucleotide-labeled, photocleavable, RNA-binding probes targeting the expression of ~1800 genes related to cancer pathways and anti-tumor immunity (Cancer Transcriptome Atlas, CTA, NanoString) plus additional custom-designed probes targeting the expression of three SCLC subtype-defining transcription factors not included in the CTA panel (NEUROD1, POU2F3, and YAP1).


title: scRNA-seq for Klf5 or SE deletion of mESC differentiation

status: Public on Apr 14 2024

city: Tianjin

first: Bohan

last: Chen

address: NO.94 Weijin Road

institute: Nankai University

summary: To investigate the effect of Klf5 or super enhancer deletion for mESC differetiation and development

overall_design: scRNA was performed for WT/KLF5KO/SEKO differentiation cells  respectively.

first: Wange

last: Lu


title: Analysis of retroviral MLT1 enhancer-driven genes identifies over-expression of EPS8L1 as an early-stage initiator of both oxidative stress and poor trophoblast invasiveness in the pregnancy disorder preeclampsia

status: Public on Apr 08 2024

first: Amit

last: Pande

address: Robert-Rössle-Straße 10

institute: Max Delbruck Center for Medicine

summary: In the human-specific disorder of pregnancy preeclampsia (PE) we find dysregulation of multiple genes associated with the cis-regulatory elements (CREs) derived from mammalian-specific endogenous retrovirus (ERV3-MLT1)

overall_design: To identify the regulatory effect of the EPS8L1 enhancer derived from mammalian-specific endogenous retrovirus (ERV3-MLT1).

first: Laurence

last: Hurst

first: Zsuzsanna

last: Izsvak

first: Rabia

last: Anwar


title: Cellular heterogeneity and dynamics of the human uterus in healthy premenopausal women

city: Ann Arbor

last: Vargo

state: United States

address: 4909 Buhl Building, 1241 E. Catherine St.

department: Human Genetics

institute: University of Michigan

summary: We applied single-cell RNA sequencing to profile >50K uterine cells from both the endometrium and myometrium of 5 healthy premenopausal individuals, and jointly analyzed the data with a previously published dataset from 15 subjects. The resulting normal uterus cell atlas contains >167K cells representing the lymphatic endothelium, blood endothelium, stromal, ciliated epithelium, unciliated epithelium, and immune cell populations. Focused analyses within each major cell type and comparisons with subtype labels from prior studies allowed us to document supporting evidence, resolve naming conflicts, and to propose a consensus annotation system of 39 subtypes. We find many subtypes show dynamic change over different phases of the cycle, and identify multiple potential progenitor cells at multiple levels: compartment-wide progenitors for each major cell type, transitional cells that are upstream of other subtypes, and potential cross-lineage multipotent stromal progenitors that may be capable of replenishing the epithelial, stromal, and endothelial compartments. When compared to the healthy premenopausal samples, a postpartum and a postmenopausal uterus sample revealed substantially altered tissue composition, involving the rise or fall of stromal, endothelial, and immune cells. The cell taxonomy and molecular markers we report here are expected to inform studies of both basic biology of uterus function and its disorders.

overall_design: Creation of cell atlas: Characterizing cellular composition and heterogeneity of healthy premenopausal, postmenopausal, and postpartum human uterus using 10x scRNA-seq analysis

first: Nicole

last: Ulrich

first: Qianyi

last: Ma

first: Yu-chi

last: Shen

first: Jun

middle: Z

last: Li

first: Saher

last: Hammoud


title: OCA-B/Pou2af1 promotes autoimmune demyelination through control of stem-like CD4+ T cells

first: Erik

last: Hughes

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

overall_design: Refer to individual Series


title: Strong Correlation of Gene Counts and Differentially Expressed Genes Between a 3′ RNA-Seq and RNA Hybridisation Platform in Transcriptome Analyses from Canine Archival Tissue [nCounter]

first: Alina

last: Loriani Fard

summary: Canine cutaneous histiocytoma (CCH) is a tumor originating from dermal Langerhans cells, especially affecting young dogs. The common spontaneous regression of CCH makes it an interesting model in comparative oncology research. Previous studies have indicated that anti-tumor immune responses may be involved but details remain speculative to date. Here, we asked which specific immuno-oncological dynamics underlie spontaneous regression of CCH on mRNA and protein levels. QuantSeq 3' mRNA sequencing with functional overrepresentation analysis and an nCounter® RNA hybridization assay were employed on 21 formalin-fixed, paraffin-embedded CCH samples representing three different tumor stages. Nine additional samples were subjected to matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI). Surprisingly, only minor stage-specific differences were found. When we investigated expression of B7 family ligands and CD28 family receptors holding co-stimulatory and -inhibitory functions respectively, we found higher abundance of CD80, CD86 and CD28, which trigger activation of lymphocyte-mediated immune responses. CD80 and CD86 expressing cells were further quantified by in situ hybridization and compared with data from three cases of canine histiocytic sarcoma (HS), a malignant tumor variant originating from antigen presenting cells. A stage-specific increase of CD80 expression was recorded in CCH from the tumor bottom to the top, while CD86 was continuously and homogenously expressed at high levels. Expression of CD80 in CCH was similar to that in HS (73.3 ± 37.4% versus 62.1 ± 46.4% positive cells), while CD86 was significantly higher expressed in CCH (94.7 ± 10.3%) when compared to HS (57.6 ± 11.0%). Our data suggest that major immuno-oncological pathways are seemingly not regulated during regression of CCH on the mRNA or protein levels as detectable by the methods used. We speculate that key signals leading to tumor regression may occur at an interactome level or at an earlier time point than examined here. Our data further supports a role of immune stimulatory B7 family ligands and CD28 family receptors in CCH regression.

overall_design:
To determine the correlation of gene expression data generated by the nCounter® Canine IO Panel and QuantSeq 3' from three types of canine tumours. FFPE tissue samples were obtained from the archive (spanning the years 2013 to 2021) and total RNA was extracted. After QuantSeq 3' and hybridisation to the nCounter® Canine IO Panel with bioinformatic processing, counts and differential gene expression data were compared to calculate correlations between the two technologies employed.

Attention: This GEO submission includes the nCounter® data from the stage-contrasting comparison (n = 10). The data for this comparison from 3' RNA-Seq (QuantSeq 3') was submitted separately to GEO (262022). The other entity-contrasting tumour samples (n = 25) used in the publication were also submitted separately to GEO (nCounter®: 261791; 3' RNA-Seq: 261790).

first: Alexander

last: Haake

first: Vladimir

last: Jovanovic

first: Sandro

last: Andreotti


title: Aged human serum induces changes in barrier function in an isogenic venule model

city: Baltimore

first: Peter

last: Searson

state: MD

address: 3400 North Charles Street

institute: Johns Hopkins University

summary: Vascular aging and dysfunction are significant contributors to age-related cardiovascular and neurodegenerative diseases, both of which encompass major causes of death in the aged population. In particular, aging heavily impacts small vessels, damages vascular integrity leading to leakage events and inflammation, and can be further exacerbated by environmental factors that may result in more severe symptomatic presentation of disease. Here, we evaluate generalized, isogenic, co-culture in vitro models of microvasculature under perfusion with aged human serum over just four days, which results in a transcriptomic recapitulation of an aging phenotype, transcript and protein changes in junctional mediators, and functional loss of paracellular and transcellular barrier integrity.  Additionally, in comparison with endothelial monoculture, we identify critical changes to basement membrane composition and aging-cue-mediated cell cycle shifts with pericyte co-culture. This modular approach reveals key impacts to further our understanding of vascular aging to leverage in designing therapeutic and preventative approaches.

overall_design: total samples collected n = 18; n = 3 for each conditions across two models (control microvessel vs. venule model) and three perfusion conditions (media vs. young serum vs. aged serum supplementation)

first: Tracy

last: Chung


title: Defining Cis-regulatory Elements and Transcription Factors that Control Human Cortical Interneuron Development

status: Public on Apr 17 2024

first: Kristen

last: Kroll


title: Proteasome inhibition reprograms chromatin landscape in breast cancer (Start-Seq)

status: Public on Mar 13 2024

city: Durham

first: Brian

last: Bennett

address: 111 T.W. Alexander Drive

institute: NIEHS

summary: The 26S proteasome regulates general protein homeostasis and controls vital cellular processes, including cell division and transcriptional regulation. Cancer cells are dependent on the proper functioning of the proteasome to modulate gene expression networks that promote tumor growth. Proteasome inhibition has emerged as an effective therapeutic strategy, although mechanisms by which this approach achieves its outcomes remain unclear.  We performed an integrative analysis of the transcriptome and chromatin landscape of MCF-7 breast cancer cells treated with a model proteasome inhibitor, MG132, to examine the consequences of proteasome inhibition on gene regulation. MG132 treatment initiated dynamic changes in chromatin accessibility at specific loci termed Differentially Open Chromatin Regions (DOCRs).  DOCRs with increased accessibility were primarily distal to transcription start sites (TSS), while those with decreased accessibility were promoter proximal and distal to TSS.  Promoter proximal DOCRs showed a unique chromatin architecture associated with distinct divergent transcription patterns.  Conversely, DOCRs distal to TSS were enriched in oncogenic super enhancers that are predominantly accessible in non-basal compared to basal breast tumor subtypes.  These data define distinct chromatin states and RNAPII transcription patterns, revealing molecular mechanisms by which the proteasome modulates the expression of gene networks intrinsic to breast cancer biology.

overall_design: Start-seq was used to characterize transcription initiation in MCF-7 breast cancer cells treated with either vehicle (UNTR) or proteasome inhibitor MG132 (1 uM) for 4 and 24 hr

first: Trevor

last: Archer

first: H

last: Kinyamu


title: Strong Correlation of Gene Counts and Differentially Expressed Genes Between a 3′ RNA-Seq and RNA Hybridisation Platform in Transcriptome Analyses from Canine Archival Tissue [nCounter®]

middle: F. H.

address: Robert-von-Ostertag-Straße 15

overall_design:
To determine the correlation of gene expression data generated by the nCounter® Canine IO Panel and QuantSeq 3' from three types of canine tumours. FFPE tissue samples were obtained from the archive (spanning the years 2013 to 2021) and total RNA was extracted. After QuantSeq 3' and hybridisation to the nCounter® Canine IO Panel with bioinformatic processing, counts and differential gene expression data were compared to calculate correlations between the two technologies employed. - Attention: The data in this GEO submission comes from the entity-contrasting comparison of the two most common glandular, perianal tumours in the dog – the hepatoid gland adenoma (HGA, n = 10) and apocrine gland anal sac adenocarcinoma (AGASAC, n = 15). The other stage-contrasting tumour samples (n = 10) used in the publication were submitted separately to GEO.

Attention:
This GEO submission includes the nCounter® data from 25 of the 35 samples used in this study! The data from the from 3' RNA-Seq (QuantSeq 3'; GSE261790) and the other 10 stage-contrasting tumour samples (n = 10) were submitted separately to GEO (nCounter®: GSE262020, 3' RNA-Seq: GSE262022).

first: Achim

last: Gruber


title: OCA-B/Pou2af1 promotes autoimmune demyelination through control of stem-like CD4+ T cells [Bulk RNAseq]

overall_design: Triplicate samples of mCherry positive and negative CD4 T cells which were FACS sorted from Ocab-mCherry reporter mice primed with MOG35-55 in CFA for 14 days and processed for RNA sequencing

first: Amber

last: Syage

first: Elnaz

last: Mirzaei Mehrabad

first: Tom

last: Lane

first: Benjamin

last: Spike

first: Dean

last: Tantin


title: Tumor DNA methylation profiling of patients with Adamantinomatous craniopharyngioma (ACP)

status: Public on Mar 14 2024

city: Ribeirão Preto

first: Junier

middle: Marrero

last: Gutiérrez

state: São Paulo

address: Av Bandeirantes, 3900

department: Pediatric

country: Brazil

institute: Ribeirão Preto Medical School-University of São Paulo

type: Methylation profiling by genome tiling array

summary: Abnormal DNA methylation plays a role in tumor progression and is becoming recognized as a prognostic marker in various types of cancers. In this study, we aimed to investigate the association between DNA methylation and clinicopathological features, prognosis, and outcomes in patients with Adamantinomatous craniopharyngioma (ACPs). To achieve this, we analyzed the global methylation profile of 35 tumors using the Illumina Infinium HumanMethylation850 BeadChip.

overall_design: DNA from 35 microdissected, fresh frozen tumor samples from was extracted using the QIAamp DNA Mini Kit (QIAGEN, Hilden, Germany) according to the manufacturer's instruction. DNA was commisioned to the University of Southern California Keck Genomics Platform (Los Angeles, CA, USA), where it was bisulfite-converted using tht EZ DNA methylation kit (Zymo Research, Irvine, CA) , as provided by the manufacturer, and hybridized to the Illumina Infinium HumanMethylation850 BeadChip as specified by Illumina.

first: Ana

last: Bueno

first: Clarissa

last: Martins

first: Margaret

last: de Castro


title: Putative looping factor ZNF143/ZFP143 is an essential transcriptional regulator with no looping function [PRO-seq]

status: Public on Mar 07 2024

city: Cambridge

first: Domenic

last: Narducci

state: MA

address: Building 56 Room 787B, 32 Vassar Street

department: Biological Engineering

institute: Massachusetts Institute of Technology

summary: Interactions between distal loci in mammalian genomes, including those involving enhancers and promoters, are thought to be a central mechanism of gene regulation in mammals, yet the protein regulators of these interactions remain largely undetermined. The zinc finger transcription factor ZNF143/ZFP143 has been strongly implicated as a factor that regulates chromatin interactions, functioning either with or without CTCF. However, ZNF143/ZFP143’s role in this process and its function with or without CTCF are not well understood. Here, we tagged both CTCF and ZNF143/ZFP143 with dual-purpose degron/imaging tags to combinatorically assess their loop function and effect on each other. We find that ZNF143/ZFP143 possesses no general looping function, and that it largely functions independently to CTCF. Instead, ZNF143/ZFP143 is an essential and highly conserved transcription factor possessing an extremely stable chromatin residence time (>20 min) that regulates an important subset of mitochondrial and ribosomal genes.

overall_design: We generated two replicates of Micro-C, ZFP143 ChIP-seq, CTCF ChIP-seq, and PRO-seq per cell line and condition in two mouse embryonic stem cell (mESC) cell lines with the following conditions: untreated, three-hour ZFP143 depletion by 5-Ph-IAA/HaloPROTAC3 treatment, three-hour CTCF depletion by dTag-13 treatment, or three-hour depletion of both ZFP143 and CTCF. We also generated two replicates of Micro-C, ZFP143 ChIP-seq, CTCF ChIP-seq, and PRO-seq per condition in an additional mESC cell line with untreated and three-hour ZFP143 depletion by dTag-13 conditions. Additionally, we performed two replicates of Micro-C per condition in a HEK293T cell line with untreated and three-hour ZNF143 depletion by IAA conditions.

middle: N

first: Anders

last: Hansen


title: Programmable transcript-specific enrichment for single-cell sequencing enables profiling of rare cell states

status: Public on Mar 26 2024

city: New York

first: Caleb

last: Lareau

state: New York

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

institute: Memorial Sloan Kettering

summary: The widespread application of single-cell genomics technologies has accelerated our understanding of the breadth and depth of heterogeneity of cell states across diverse contexts. As single-cell RNA sequencing (scRNA-seq) has been the most popular modality used for profiling, many populations have been described primarily based on specific marker transcript profiles compared to classical cytometry approaches relying on protein expression. Additionally, many single cell studies require the isolation of nuclei from tissue, eliminating the ability to enrich learned rare cell states based on extranuclear protein markers. To address this limitation, we describe Programmable Enrichment via RNA Flow-FISH by sequencing (PERFF-seq), a scalable assay that enables single cell and single nuclei RNA-seq profiling from subpopulations of complex cellular mixtures solely defined by the abundance of RNA transcripts. Across vignettes of immune cell populations as well as nuclei from fresh frozen and formalin-fixed paraffin-embedded brain tissue, we demonstrate the enrichment of cell populations via RNA-based cytometry upstream of high-throughput scRNA-seq. Together, our approach provides a rational, programmable method for studying cell identities and transcriptional heterogeneity of rare populations identifiable by as few as one marker transcript, advancing the rational study of cellular diversity across fresh and archived tissue materials.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
Human PBMCs, mouse brain nuclei, and human GBM FFPE blocks were processed with HCR-FlowFISH reagents and profiled via the 10x Genomics Flex library kit

Three donors of different ages were profiled with PERFF-seq and pooled using the Flex in-line barcoding 4plexing technology. Cells sorted positive had Y chromosome signa, and the negative cells were further divided based on SSC (top and bottom)

first: Tsion

last: Abay

first: Robert

last: Stickels

first: Ronan

last: Chaligne


title: Putative looping factor ZNF143/ZFP143 is an essential transcriptional regulator with no looping function [ChIP-seq]


title: RNA-seq on FACS sorted paneth cells from the murine duodenum, jejunum and ileum

city: Gent

first: Steven

last: Timmermans

address: Technologiepark 71

country: Belgium

institute: VIB-UGent

summary: The small intestinal crypts harbour secretory Paneth cells (PCs), which express bactericidal peptides that are crucial for maintaining intestinal homeostasis. Considering the diverse environmental conditions throughout the course of the small intestine, multiple subtypes of PCs are expected to exist. We applied single cells RNA-sequencing of PCs combined with deep bulk RNA-sequencing on PC populations of different locations (duodenum, jejunum, ileum) and discovered several expression-based clusters. Some of these are discrete and resemble tuft cell-like PCs, goblet cell-like PCs, PCs expressing stem cell markers, and senescent PCs. Other clusters are less discrete, but based on bulk RNA-seq appear to be derived from different locations along the small intestinal tract. Furthermore, a comprehensive spatial analysis was conducted using the Resolve bioscience technique, identifying different PCs clusters along the small intestine, but not along the crypts themselves.

overall_design: We isolated PCs from the three regions using a pool of 8 mice, and repeated this in total 4 times independently. RNA as isolated and sequenced followed by differentail gene expression profiling

first: Charlotte

last: Wallaeys

first: Lotte

last: Polaris

first: Natalia

last: Garcia-Gonzalez

first: Yvan

last: Saeys

first: Claude

last: Libert


title: Putative looping factor ZNF143/ZFP143 is an essential transcriptional regulator with no looping function [Micro-C]


title: Surface CD52, CD84 and PTGER2 mark mature PMN-MDSCs from cancer patients and G-CSF-treated donors

city: Verona

first: Barbara

last: Mariotti

state: Verona

address: strada le grazie 8

department: Medicine

country: Italy

institute: University of Verona


title: Surface CD52, CD84 and PTGER2 mark mature PMN-MDSCs from cancer patients and G-CSF-treated donors [bulk RNA-seq]

summary:
In this study, we reported the identification of a specific gene signature expressed
by mPMN-MDSCs from NSCLC/HNC patients and G-CSF-treated donor (GDs) by bulk-RNA-seq that clearly reflects their metabolic/functional reprogramming. ScRNA-seq experiments further confirmed that the mPMN-MDSC gene signature is significantly enriched in both mNDNs/mLDNs from GDs.

overall_design: CD66b+CD10+CD11b+CD16+ GD-mLDNs, GD-mNDNs and control HD-mNDNs were isolated from the peripheral blood (PB) of GDs or healthy donors (HDs) by flow cytometry sorting. CD66b+CD10+CD11b+CD16+ mPMN-MDSCs and autologous mNDNs from NSCLC or HNC patients, as well as control HD-mNDNs, were isolated from the PB by cell sorting via CD66b+ beads or by flow cytometry sorting.   CD66b+CD10-CD11b+CD16+ band cells (BCs), CD66b+CD10-CD11b+CD16+/dim metamyelocytes (MMs), CD66b+CD10-CD11bdimCD16- myelocytes (MYs) and CD66b+CD10-CD11b-CD16- promyelocytes (PMs) from either the PB of GDs by flow cytometry sorting.   Total RNA was then extracted and subjected to Smart-seq2 protocol for transcriptome analysis.

first: Marco

last: Cassatella

first: Patrizia

last: Scapini

first: Nicola

last: Tamassia

first: Sven

last: Brandau


title: Strong Correlation of Gene Counts and Differentially Expressed Genes Between a 3′ RNA-Seq and RNA Hybridisation Platform in Transcriptome Analyses from Canine Archival Tissue [3' RNA-Seq]

first: Alexander

last: Haake

overall_design:
To determine the correlation of gene expression data generated by the nCounter® Canine IO Panel and QuantSeq 3' from three types of canine tumours. FFPE tissue samples were obtained from the archive (spanning the years 2013 to 2021) and total RNA was extracted. After QuantSeq 3' and hybridisation to the nCounter® Canine IO Panel with bioinformatic processing, counts and differential gene expression data were compared to calculate correlations between the two technologies employed. - Attention: The data in this GEO submission comes from the entity-contrasting comparison of the two most common glandular, perianal tumours in the dog – the hepatoid gland adenoma (HGA, n = 10) and apocrine gland anal sac adenocarcinoma (AGASAC, n = 15).

Attention:
This GEO submission includes the 3' RNA-Seq data from 25 of the 35 samples used in this study! The data from the RNA hybridisation platform (nCounter®; GSE261791) and the other 10 stage-contrasting tumour samples (n = 10) were submitted separately to GEO (nCounter®: GSE262020, 3' RNA-Seq: GSE262022).

first: Alina

last: Loriani Fard

first: Vladimir

last: Jovanovic

first: Sandro

last: Andreotti

first: Achim

last: Gruber


title: Gene expression analysis using RNA-Seq of dithranol-treated psoriasis

status: Public on Mar 06 2024

city: Graz

first: Theresa

last: Benezeder

address: Auenbruggerplatz 8

department: Department of Dermatology and Venereology

country: Austria

institute: Medical University of Graz

summary: Topical dithranol (anthralin) has remained an effective anti-psoriatic agent. Although dithranol has been used for many years, it has never been fully revealed how dithranol exerts its anti-psoriatic mechanisms.Thus, we studied the therapeutic mechanisms of this drug in serial biopsies from human psoriatic lesions.

overall_design: To investigate the effect of topical dithranol treatment on mRNA expression in psoriasis, dithranol-treated psoriasis skin biopsy were analyzed with RNA Seq and compared to untreated psoriasis and non-lesional skin.

first: Natalie

last: Bordag

first: Peter

last: Wolf


title: Liebenberg syndrome severity arises from variations in Pitx1 locus topology and ectopically transcribing cells

status: Public on Mar 04 2024

city: Geneva

first: Guillaume

last: Andrey

address: Rue Michel-Servet 1

department: Department of Genetic Medicine and Development

country: Switzerland

institute: University of Geneva

summary: Enhancer hijacking, a common cause of gene misregulation linked to disease, occurs when non-matching enhancers and promoters interact ectopically. This interaction is made possible by genetic changes that alter the arrangement or insulation of gene regulatory landscapes. While the concept of enhancer hijacking is well understood, the specific reasons behind the variation in phenotypic severity or the point at which those phenotypes become evident remain unexplored. In this work, we expand on the ectopic activation of the hindlimb-specific transcription factor Pitx1 by one of its own enhancers, Pen, in forelimb tissues that causes the Liebenberg syndrome. We combine a previously developed in-embryo cell-tracing approach to a series of inversions and relocations to show that reduction in Pitx1-Pen relative genomic positioning leads to increased proportions of Pitx1 forelimb-expressing cells and more severe phenotypical outcomes. We demonstrate that the Pitx1 locus assumes an active topology when enhancer-promoter contacts are required for transcription and that its promoter generates consistent transcription levels across different alleles. Finally, we show that changes in 3D chromatin structure and enhancer-promoter contacts are not the result of Pitx1 transcriptional activity. In summary, our work shows that variation in enhancer-promoter interactions can lead to pathogenic locus activation in variable proportions of cells which, in turn, define phenotypic severity.


title: Surface CD52, CD84 and PTGER2 mark mature PMN-MDSCs from cancer patients and G-CSF-treated donors [scRNA-seq]

summary: In this study, we reported the identification of a specific gene signature expressed by mPMN-MDSCs from NSCLC/HNC patients and G-CSF-treated donor (GDs) by bulk-RNA-seq that clearly reflects their metabolic/functional reprogramming. ScRNA-seq experiments further confirmed that the mPMN-MDSC gene signature is significantly enriched in both mNDNs/mLDNs from GDs

overall_design: Validation of the defined mPMN-MDSC gene signature using scRNA-seq data of CD66b+ neutrophils from the LDN and NDN fractions of peripheral blood from G-CSF-treated donors, and the NDN fraction of peripheral blood or the LDN fraction of BM from healthy donors (HDs) as controls.

first: Chiara

last: Lattanzi

first: Francesca

last: Pettinella


title: Illuminating the terminal nerve: Uncovering the link between GnRH-1 neuron and olfactory development

status: Public on Mar 11 2024

city: Albany

first: Paolo

middle: Emanuele

last: Forni

state: NY

address: 1400 Washington Avenue

department: Biology

institute: University at Albany

summary:
During embryonic development, the olfactory placode (OP) gives rise to various populations of neurons; these include putative olfactory pioneer neurons, different neurons of unknown identity and function, cells of the terminal nerve, and the Gonadotropin-releasing hormone-1 (GnRH-1) neurons. In mice, the GnRH-1 neurons are first detectable in the developing olfactory system around mid-gestation. From here, the GnRH-1 neurons migrate, along the axons of the terminal nerve (TN), to various regions of the developing brain. Once in the brain, the GnRH-1 neurons play a central role in controlling the hypothalamic-pituitary-gonadal (HPG) axis. Early migratory neurons forming from the olfactory placode have been proposed to play a vital role in inducing olfactory bulb morphogenesis.
Kallmann syndrome is a condition characterized by defective development of the olfactory system and infertility.
Murine studies have demonstrated the critical role of the Prokineticin 2-Prokinteicin Receptor 2  pathway in olfactory bulb morphogenesis and GnRH-1 neuronal migration. Loss-of-function Prokr2 mutations cause both Kallmann syndrome associated with bulb agenesis and normosmic idiopathic hypogonadotropic hypogonadism (nIHH).
Following  Prokr2 expression and lineage tracing, we found that Prokr2 is not expressed by the cells of the developing olfactory bulb but by migratory putative pioneer/terminal nerve neurons. Performing single-cell-RNA-sequencing, we identified genes enriched in the migratory cells of the putative terminal nerve. By integrating genetic lineage tracing and single-cell transcriptomics, we identified previously undescribed populations of migratory neurons that appear to be enriched in the expression of several genes related to olfactory defects, GnRH migratory deficiencies and infertility.

overall_design: Whole embryonic noses were dissected from E14.5 wild-type mice, cells were isolated then analyzed using scRNAseq. ProKR2iCre/R26tdTomato E14.5 mice had noses dissected, then cells were isolated using Flourescence-activated cell sorting (FACS), then scRNAseq was performed.

first: Enrico

last: Amato Jr

first: Ed-Zandro

last: Taroc


overall_design: We used capture Hi-C enriching a 3,3Mb region at the Pitx1 locus in Mus musculus. We analysed GFP postive and negative cells from Pitx1GFP;Inv1, Pitx1GFP;Inv2, Pitx1GFP;Rel1, Pitx1GFP;Rel2 and Pitx1GFP;Rel3 forelimbs; dCas9P300-Pitx1TSSsgRNAs and dCas9P300-noSgRNAs, Prx1-CRE, Eedflox/- and wildtype bulk forelimbs; and Pitx1GFP;Inv1, Pitx1GFP;Inv2 and Pitx1GFP mESC cells to compare the structural changes at the locus.

first: Olimpia

last: Bompadre


overall_design: We used chromatin immunoprecipitation against H3K27ac chromatin marks in Mus musculus forellimb cells. We analysed GFP postive  from from Pitx1GFP;Inv1, Pitx1GFP;Rel2 and Pitx1GFP;Rel3 cellsto compare changes in activity level at the locus.


title: Single-cell analysis of Rohon-Beard neurons implicates Fgf signaling in axon maintenance and cell survival

status: Public on Mar 01 2024

city: Portland

first: Alex

last: Nechiporuk

state: OR

address: 3181 SW Sam Jackson Park Road

institute: Oregon Health and Science Univ

summary: Peripheral sensory neurons are a critical part of the nervous system that transmit a multitude of sensory stimuli to the central nervous system. During larval and juvenile stages in zebrafish, this function is mediated by Rohon-Beard somatosensory neurons (RBs). RBs are optically accessible and amenable to experimental manipulation, making them a powerful system for mechanistic investigation of sensory neurons. Previous studies provided evidence that RBs fall into multiple subclasses; however, the number and molecular make up of these potential RB subtypes have not been well defined. Using a single-cell RNA sequencing (scRNA-seq) approach, we demonstrate that larval RBs in zebrafish fall into three, largely non-overlapping classes of neurons. We also show that RBs are molecularly distinct from trigeminal neurons in zebrafish. Cross-species transcriptional analysis indicates that one RB subclass is similar to a mammalian group of A-fiber sensory neurons. Another RB subclass is predicted to sense multiple modalities, including mechanical stimulation and chemical irritants. We leveraged our scRNA-seq data to determine that the fibroblast growth factor (Fgf) pathway is active in RBs. Pharmacological and genetic inhibition of this pathway led to defects in axon maintenance and RB cell death. Moreover, this phenotype can be phenocopied by treatment with an FDA-approved Fgf inhibitor dovitinib, which is used in clinic and causes peripheral neuropathy. Importantly, dovitinib-mediated axon loss can be suppressed by loss of Sarm1, a positive regulator of neuronal cell death and axonal injury. This offers a molecular target for future clinical intervention to fight neurotoxic effects of this drug.

overall_design: Thirty-hour old TgBAC(neurod1:EGFP)nl1 zebrafish embryos were collected and euthanized in 1.7 ml microcentrifuge tubes. Embryos were deyolked using a calcium-free Ringer’s solution (116 mM NaCl, 2.6 mM KCl, 5 mM HEPES pH 7.0), by gently pipetting up and down with a P200 pipet. Embryos were incubated for 5 minutes in Ringer’s solution. Embryos were transferred to pre-warmed protease solutions (0.25% trypsin, 1 mM EDTA, pH 8.0, PBS) and collagenase P/HBSS (100 mg/mL) was added. Embryos were incubated at 28° C for 15 minutes and were homogenized every 5 minutes using a P1000 pipet. The Stop solution (6X, 30% calf serum, 6 mM CaCl2, PBS) was added and samples were centrifuged (350xg, 4° C for 5 minutes). Supernatant was removed and 1 mL of chilled suspension solution was added (1% FBS, 0.8 mM CaCl2, 50 U/mL penicillin, 0.05 mg/mL streptomycin, DMEM). Samples were centrifuged again (350g, 4° C for 5 minutes) and supernatant was removed. 700 μl of chilled suspension solution was added and cells were resuspended by pipetting. Cells were passed through a 40 μm cell strainer into a FACs tube and kept on ice. GFP and RFP+ cells were FAC sorted on a BD Symphony cell sorter into sorting buffer (50 μl PBS/ 2% BSA) in a siliconized 1.5mL tube.

first: Adam

last: Tuttle

first: Lauren

last: Miller

first: Lindsey

last: Royer

first: Jimmy

last: Kelly

first: Nicholas

middle: L

last: Calistri

first: Laura

last: Heiser

middle: V


overall_design: Whole forelimb buds were prepared for single cell RNA-seq analysis to identify which cell types are affected by ectopic gene expression.


title: A diverse network of pericoerulear neurons control arousal states

city: Seattle

first: Xiaonan

last: Fu

state: WA

address: 1705 NE Pacific St

department: Biochemistry

institute: University of Washington

summary: As the primary source of norepinephrine (NE) in the brain, the locus coeruleus (LC) regulates both arousal and stress responses1,2. However, how local neuromodulatory inputs contribute to LC function remains unresolved. Here we identify a network of transcriptionally and functionally diverse GABAergic neurons in the LC dendritic field that integrate distant inputs and modulate modes of LC firing to control arousal. We define peri-LC anatomy using viral tracing and combine single-cell RNA sequencing and spatial transcriptomics to molecularly define both LC and peri-LC cell types. We identify several cell types which underlie peri-LC functional diversity using a series of complementary approaches in behaving mice. Our findings indicate that LC and peri-LC neurons comprise transcriptionally and functionally heterogenous neuronal populations, alongside anatomically segregated features which coordinate specific influences on behavioral arousal and avoidance states. Defining the molecular, cellular and functional diversity in the LC provides a road map for understanding the neurobiological basis of arousal alongside hyperarousal-related neuropsychiatric phenotypes.

overall_design: six sections from female adult wild-type mouse and five sections from male adult mouse were used for spatial transcriptome analysis

first: Andrew

last: Luskin

first: Li

last: Li

first: Kelsey

last: Barcomb

first: Taylor

last: Blackburn

first: Esther

first: Akshay

last: Rana

first: Rhiana

middle: C

last: Simon

last: Sun

first: Alexandria

middle: D

last: Murry

first: Sam

middle: A

last: Golden

first: Garret

last: Stuber

first: Christopher

middle: P

last: Ford

first: Liangcai

last: Gu

first: Michael

middle: R

last: Bruchas


title: Mezigdomide is effective alone and in combination with Menin inhibition in pre-clinical models of KMT2A-r and NPM1c AML

status: Public on Feb 20 2024

city: Boston

first: Scott

last: Armstrong

address: 450 Brookline Ave

department: pediatric oncology

institute: Dana Farber Cancer Institute

summary: To compare gene expression changes following treatment with two CELMoDs (cereblon E3 ligase modulators) in a Lysine Methyltransferase 2A-rearranged (KMT2A-r) acute myeloid leukemia cell line, we compared gene expression changes after 72 hour of treatment with Iberdomide/CC-220 and Mezigdomide/CC-92480.

overall_design: MOLM13 cells were treated for 72 hours with DMSO control, Iberdomide (CC-220) 1 uM, or Mezigdomide (CC-92480) 50 nM with triplicates for each condition.

first: Wallace

last: Bourgeois

first: Charlie

last: Hatton


title: Single-nucleus RNA sequencing demonstrates an autosomal dominant Alzheimer's disease profile and possible mechanisms of disease protection

status: Public on Feb 28 2024

city: Santa Barbara

first: Kenneth

last: Kosik

state: CA

address: 5165 Biology II

department: Molecular, Cellular, and Developmental Biology

institute: University of California Santa Barbara

overall_design: Refer to individual Series


title: Single-nucleus RNA sequencing demonstrates an autosomal dominant Alzheimer's disease profile and possible mechanisms of disease protection [spatial transcriptomics]

summary: Highly penetrant autosomal dominant Alzheimer's disease (ADAD) comprises a distinct disease entity as compared to the far more prevalent form of AD in which common variants collectively contribute to risk. The downstream pathways that distinguish these AD forms in specific cell types have not been deeply explored. We compared single-nucleus transcriptomes among a set of 27 cases divided among PSEN1-E280A ADAD carriers, sporadic AD, and controls. Autophagy genes and chaperones clearly defined the PSEN1-E280A cases compared to sporadic AD. Spatial transcriptomics validated the activation of chaperone-mediated autophagy genes in PSEN1-E280A. The PSEN1-E280A case in which much of the brain was spared neurofibrillary pathology and harbored a homozygous APOE3-Christchurch variant revealed possible explanations for protection from AD pathology including overexpression of LRP1 in astrocytes, increased expression of FKBP1B, and decreased PSEN1 expression in neurons. The unique cellular responses in ADAD and sporadic AD require consideration when designing clinical trials.

overall_design: We performed comparative gene expression profiling analysis using spatial transcriptomics data obtained from two different brain regions (frontal cortex and hippocampus) of post mortem brains of two patient carriers of the autosomal dominant PSEN-1E280A mutation and one healthy older control.

first: Maria

last: Almeida

first: Sarah

middle: J

last: Eger

first: Caroline

last: He

first: Morgane

last: Audouard

first: Arina

last: Nikitina

first: Stella

last: Glasauer

first: Dasol

last: Han

first: Barbara

last: Cupajita

first: Juliana

last: Acosta-Uribe

first: Nelson

last: Villalba-Moreno

first: Jessica

last: Littau

first: Megan

last: Elcheikhali

first: Erica

middle: K

last: Rivera

first: Daniel

last: Carrettiero

first: Carlos

last: Villegas-Lanau

first: Diego

last: Sepulveda-Falla

first: Francisco

last: Lopera


title: Non-canonical functions of UHRF1 maintain DNA methylation homeostasis in cancer cells [RNA-seq]

status: Public on Feb 12 2024

city: Paris

first: Pierre-Antoine

last: Defossez

address: 35 rue Hélène Brion

institute: Epigenetics and Cell Fate Unit

summary: DNA methylation is an essential epigenetic mark in mammals. It controls gene expression and genome stability. Global DNA methylation pattern is abnormal in cancers. Ubiquitin like with PHD and RING finger domains 1 (UHRF1) is a key epigenetic regulator that recruits and activates DNA methyltransferase 1 (DNMT1), the methylation maintenance enzyme. UHRF1 is a proven oncogene and its overexpression transforms cells in vitro and causes cancer in animal models. Therefore, UHRF1 provides a unique entry point into the links between epigenetics and cancer. However, it is still not fully clear how UHRF1 works in cancer cells.  To understand UHRF1 functions in cancer, we employed experimental strategy to use an advanced chemical/genetic system, the auxin-inducible degron (AID) technology, whereby the degron-fused protein can be totally and rapidly degraded upon the addition of a small molecule, auxin. We chose the human CRC cell line HCT116 as our model and successfully generated UHRF1-AID and DNMT1-AID. Through this study, we made the significant discovery that UHRF1 not only regulates DNMT1, but also influences the activities of de novo methyltransferases DNMT3A and DNMT3B, as well as the active demethylase TET2.

overall_design: To elucidate the roles of UHRF1 and DNMT1 in the HCT116 cell line, we generated four distinct cell lines: wild-type (WT), UHRF1-AID, DNMT1-AID, and UHRF1-AID/DNMT1-AID. To analyze the transcriptome change, we treated these cell lines with auxin for 0,  4 days.

first: Kosuke

last: YAMAGUCHI

first: Fumihito

last: MIURA

first: Pavel

last: Bashtrykov

first: Olivier

last: KIRSH

first: Albert

last: Jeltsch

first: Takashi

last: Ito


first: Kenneth

last: Kosik

overall_design: We used droplet-based single nucleus RNA sequencing to perform comparative gene expression profiling analysis using data obtained from frontal cortex of post mortem brains of 8 control patients, 8 carriers of the autosomal dominant PSEN-1 E280A mutation and 8 patients that developed sporadic Alzheimer's disease.

first: Maria

last: Almeida

first: Sarah

last: Eger

first: Caroline

last: He

first: Morgane

last: Audouard

first: Arina

last: Nikitina

first: Stella

middle: M

last: Glasauer

first: Dasol

last: Han

first: Barbara

last: Cupajita

last: Villegas Lanau


title: Single Nucleotide Polymorphism (SNP) and Antibody-based Cell Sorting (SNACS): A tool for demultiplexing single-cell DNA sequencing data

status: Public on Feb 09 2024

city: San Francisco

first: Catherine

last: Smith

state: California

address: 513 Parnassus Avenue

institute: University of California, San Francisco

summary:
Motivation
Recently, single-cell DNA sequencing (scDNA-seq) and multi-modal profiling with the addition of cell-surface antibodies (scDAb-seq) have provided key insights into cancer heterogeneity. Scaling these technologies across large patient cohorts, however, is frequently cost and time prohibitive. Multiplexing, in which cells from unique patients and pooled into a single experiment, offers a possible solution. While multiplexing methods are available for scRNAseq, accurate demultiplexing in scDNAseq remains an unmet need.
Results
Here, we introduce SNACS: Single-Nucleotide Polymorphism (SNP) and Antibody-based Cell Sorting. SNACS relies on a combination of patient-level cell-surface identifiers and natural variation in genetic polymorphisms to demultiplex scDNAseq data.  We demonstrated the performance of SNACS on a dataset consisting of multi-sample experiments from patients with leukemia where we knew truth from single-sample experiments from the same patients. Relative to demultiplexing methods derived from the scRNAseq literature, SNACS offered superior accuracy.
Availability Implementation
SNACS is available at https://github.com/olshena/SNACS.

overall_design:
This study includes samples from 8 de-identified adult patients with acute myeloid leukemia. The samples underwent single-cell DNA sequencing with the inclusion of cell-surface antibody-conjugated oligonucleotides to serve as sample-level identifiers. The conditions included combining the samples in multiple combinations to develop and test a novel demultiplexing algorithm as such based on the following schematic: Sample 1: Patient A; Sample 2: Patient B; Sample 3: Patient C; Sample 4: Patient D; Sample 5: Patient A and B; Sample 6: Patient B, C, and D; Sample 7: Patient A, B, C, and D; Sample 8: Patient E and F; Sample 9: Patient E, F, G, and H; Sample 10: Patient A, B, C, D, E, F, G, and H; Sample 11: Patient A, B, C, D, E, F, G, and H. Aside from Sample 11, which is a biologic replicate for Sample 10, no other replicates were included. Samples 1-4 served as controls for Samples 5-7.
The hdf5 files were directly outputted from and open-source cell calling and alignment pipeline for single cell DNA sequencing data (https://github.com/AbateLab/DAb-seq). Data have not been normalized or filtered unless specified below. The hdf5 files contain the following matrices:
AB_DESCRIPTIONS: Names of cell-surface antibodies used
ABS: Antibody counts using various calling methods; rows are single cells and columns are antibodies. Note for this analysis and manuscript, we used the CLR (center log ratio transformed) antibody counts. Other methods are described in https://github.com/AbateLab/DAb-seq/umi_tools/Documentation.py.
AMPLICONS: This includes 2 matrices: (1) read counts for all DNA amplicons; rows are single cells, columns are DNA amplicons and (2) names of all DNA amplicons used in the experiment.
AD: Alternate allele depth; rows are single cells columns are genetic variants (SNPs)
CELL_BARCODES: Names of all called single-cell barcodes in the experiment
DP: Total read depth; rows are single cells columns are genetic variants (SNPs)
GQ: Quality score, ranging from 0 (worst quality) to 100 (best quality); rows are single cells columns are genetic variants (SNPs)
GT: Mutation call where 0 = not mutated, 1 or 2 = mutated, and 3 = NA; rows are single cells columns are genetic variants (SNPs)
RD: Reference allele depth; rows are single cells columns are genetic variants (SNPs)
VARIANTS: Names of all variants (SNPs) in the experiment

This study includes samples from 4 de-identified adult patients with acute myeloid leukemia. The samples underwent single-cell DNA sequencing with the inclusion of cell-surface antibody-conjugated oligonucleotides to serve as sample-level identifiers. The conditions included combining the samples in multiple combinations to develop and test a novel demultiplexing algorithm as such based on the following schematic: Sample 1: Patient A; Sample 2: Patient B; Sample 3: Patient C; Sample 4: Patient D; Sample 5: Patient A and B; Sample 6: Patient B, C, and D; Sample 7: Patient A, B, C, and D. No replicates were included and Samples 1-4 served as controls for Samples 5-7.

first: Vanessa

middle: E

last: Kennedy

first: Ritu

last: Roy

first: Cheryl

last: Peretz

first: Andrew

last: Koh

first: Elaine

last: Tran

first: Adam

last: Olshen


title: Effect of high temperature on Arabidopsis thaliana root tips

status: Public on Feb 07 2024

city: Nijmegen

first: Ivo

last: Rieu

address: Heyendaalseweg 135

department: Radboud Institute for Biological and Environmental Sciences

country: Netherlands

institute: Radboud University

summary: Plant roots located in the upper soil layers are prone to experience high temperatures. To gain insight into the effect of high temperature on root development and functioning, we exposed five-day-old Arabidopsis thaliana seedlings grown on agar plates to 30 °C for 48 hours, and compared the gene expression profile in the root tip with that from seedlings that remained at 22 °C.

overall_design: Seedlings were grown on agar plates for 5 days at 22 °C before start of the treatment. Control treatment was 48 hours at 22 °C, high temperature treatment was 48 hours at 30 °C. At the end of the treatment, 1-mm long tips of the roots were collected. 4 biological replicates were used from each treatment. A replicate consisted of pooled tissue from 50 seedlings (grown on 5 plates, 10 seedlings/plate).

first: Gaigai

last: Du

first: Xin

last: Tian

first: Jian

last: Xu

first: Eric

last: Visser


title: Transcriptome of Malassezia globosa in Inflammatory Bowel Disease

status: Public on Feb 01 2024

city: Seoul

first: Yong-Joon

last: Cho

address: 1 Gwanak-ro, Gwanak-gu

country: South Korea

institute: Seoul National University

summary: The human gut is inhabited by a complex ecosystem of microorganisms, encompassing bacteria, viruses, protozoa, and fungi. Recent research has illuminated the significance of the gut fungal microbiota (mycobiota) in shaping host immunity and influencing the onset and progression of various human diseases. While most investigations into gut microbiota have centered on bacteria, accumulating evidence has underscored the role of mycobiota in the development of inflammatory bowel diseases (IBD), including both ulcerative colitis (UC) and Crohn's disease (CD). In this study, we present the isolation of the live Malassezia globosa strains from the intestinal mucosa of UC patients for the first time. We provide a comprehensive analysis of the characteristics and virulence of this fungus. Malassezia, primarily known to inhabit human skin, prompted us to compare the genomes, transcriptomes, and virulence of M. globosa gut isolates with those of M. globosa strains isolated from the skin. This comparative analysis aimed to discern potential niche-specific adaptations of the fungus. Our findings reveal a striking disparity in the pathogenicity of M. globosa isolated from the gut compared to its skin counterpart. In a mouse model, gut-isolated M. globosa exhibited a more pronounced exacerbation of DSS-induced colitis and elevated production of inflammatory cytokines. Additionally, transcriptome analysis indicated that gut isolates of M. globosa display heightened sensitivity to normoxia compared to their skin-isolated counterparts, suggesting adaptation to the hypoxic conditions prevalent in the intestinal mucosal environment

overall_design: Transcriptome of skin derived strains vs. Transcriptome of gut derived strains in different oxygen circumstance

first: Seung

middle: Y

last: Shin

first: Juan

last: Yang

first: Hyo

last: Kim

first: Piyapat

last: Rintarhat

first: Minji

last: Park

first: Kate

last: Lagree

first: M

last: Underhill

first: Chang

middle: H

last: Choi

first: Chul-Su

first: Won

last: ung


title: Viral reprogramming of host transcription initiation [CAGE-seq]

city: New Orleans

first: Erik

last: Flemington

state: LA

address: 1430 Tulane Ave., SL79

institute: Tulane Health Sciences Center

summary: Viruses are master remodelers of the host cell environment in support of infection and virus production. For example, viruses typically regulate cell gene expression through modulating canonical cell promoter activity. Here, we show that Epstein Barr virus (EBV) replication causes “de novo” transcription initiation at 29,674 new transcription start sites throughout the cell genome. De novo transcription initiation is facilitated in part by the unique properties of the viral pre-initiation complex (vPIC) that binds a simple TATT[T/A]AA, TATA box-like sequence and activates transcription with minimal support by additional transcription factors. Other de novo promoters are driven by the viral transcription factors, Zta and Rta and are influenced by directional proximity to existing canonical cell promoters, a configuration that fosters transcription through existing promoters and transcriptional interference. These studies reveal a new way that viruses interact with the host transcriptome and shed light on primal features driving promoter function.

overall_design: CAGE-Seq of B cell lines, Mutu (EBV+), Akata (EBV+), during latency and reactivation conditions

first: Nate

last: Ungerleider


title: Multimodal profiling of chordoma immunity reveals distinct immune contextures

status: Public on Jan 26 2024

city: Leiden, The Netherlands

first: Siddh

last: van Oost

address: Albinusdreef 2

institute: Leiden University Medical Center

summary: Chordomas are cancers from the axial skeleton presenting immunological hallmarks of unknown significance. In recent years, some clinical trials demonstrated that chordomas can respond to immunotherapy. We present a comprehensive characterisation of immunological features of 76 chordomas through application of a multimodal approach comprising transcriptional profiling, multidimensional immunophenotyping and TCR profiling. Chordomas generally presented an immune “hot” microenvironment in comparison to other sarcomas, as indicated by the immunologic constant of rejection transcriptional signature. We identified two distinct groups of chordomas based on T cell infiltration. The highly infiltrated group was further characterised by high dendritic cell infiltration and the presence of multicellular immune aggregates in tumours, whereas low T cell infiltration was associated with lower overall cell densities of immune and stromal cells. Interestingly, patients with higher T cell infiltration displayed a more pronounced clonal enrichment of the T cell receptor repertoire compared to those with low T cell counts. Furthermore, we observed that the majority of chordomas maintained HLA class I expression. Our findings shed light on the natural immunity against chordomas. Understanding their immune landscape could guide the development and application of immunotherapies in a tailored manner, ultimately leading to an improved clinical outcome for chordoma patients.

overall_design: 20 samples

first: Judith

last: Bovee

first: Noel

middle: F

last: de Miranda


title: Congestion enriches intra-hepatic macrophages through reverse zonation of CXCL9 in liver sinusoidal endothelial cells

status: Public on Jan 27 2024

city: Rochester

first: Mrunal

middle: Kishor

last: Dehankar

state: MN

address: 200 First St. SW

department: Computational genomics

institute: Mayo Clinic

summary:
Background & Aims
Congestion alters the microenvironment of the liver sinusoid along the portal-central axis.  We studied spatial changes in immune cells in the sinusoid which contribute to congestive fibrosis and portal hypertension (PHTN).
Methods
To visualize the distribution of immune cells in congestive hepatopathy (CH), we performed imaging mass cytometry (IMC) on liver tissue from patients with CH, Fontan-associated liver disease (FALD), and controls. We performed partial ligation of the inferior vena cava (pIVCL) to simulate CH in mice and isolated primary liver cells for single cell RNA-sequencing (scRNA-seq) to study zonation of liver sinusoidal endothelial cells (LSECs).  After pIVCL, mice were treated with intraperitoneal injections of AMG487, an inhibitor of the CXCL9 receptor.
Results
Peri-central macrophages are enriched in CH and FALD.  Given the role of CXCL9 in macrophage patterning in the liver, we performed RNA in situ hybridization (RNAish) in CH and determined that CXCL9 was highly expressed in LSECs in FALD, suggesting that LSECs recruit macrophages in CH. After pIVCL, treatment with AMG487 attenuated portal pressures, fibrosis, and intra-hepatic macrophages. To study changes in LSECs which promote macrophage chemotaxis, we performed scRNA-seq after pIVCL and sham procedures. Analysis revealed 3 LSEC subpopulations according to sinusoidal location. RNANish identified peri-central LSECs as the predominant source of CXCL9 in FALD. In vitro analyses revealed that -catenin and hypoxia inducible factor-1α regulate CXCL9 transcription in peri-central LSECs.
Conclusions
CXCL9 derived from peri-central LSECs enriches intra-hepatic macrophages in CH and FALD, contributing to congestive fibrosis and PHTN.  Strategies to target LSEC-derived CXCL9 may prevent the progression of CH and FALD.

overall_design: Isolated primary cells from mice livers that underwent 6 weeks of partial Inferior Vena Cava ligation to induce portal hypertension and were subjected to Single Cell RNA sequencing.

first: Tejasav

middle: S

last: Sehrawat

first: Shawna

last: Cooper

first: Sheng

last: Cao

first: Vijay

last: Shah


title: Viral reprogramming of host transcription initiation [RNA-seq]

summary: Viruses are master remodelers of the host cell environment in support of infection and virus production. For example, viruses typically regulate cell gene expression through modulating canonical cell promoter activity. Here, we show that Epstein Barr virus (EBV) replication causes “de novo” transcription initiation at 29,674 new transcription start sites throughout the cell genome. De novo transcription initiation is facilitated in part by the unique properties of the viral pre-initiation complex (vPIC) that binds a simple TATT[T/A]AA, TATA box-like sequence and activates transcription with minimal support by additional transcription factors. Other de novo promoters are driven by the viral transcription factors, Zta and Rta and are influenced by directional proximity to existing canonical cell promoters, a configuration that fosters transcription through existing promoters and transcriptional interference. These studies reveal a new way that viruses interact with the host transcriptome and shed light on primal features driving promoter function

overall_design: RNA-Sequencing of B cell lines, Mutu (EBV+), Akata (EBV+), during latency and reactivation conditions and DG75 (EBV-) following Zta/Rta transfection.