title: Single-cell transcriptomic analysis of zebrafish fli1b mutants

status: Public on Mar 26 2025

city: Tampa

first: Saulius

last: Sumanas

address: 560 Channelside Dr

department: Department of Pathology and Cell Biology

institute: University of South Florida

summary: To characterize fli1b role in zebrafish hematopoietic development, we obtained samples from zebrafish fli1b mutant and control wild-type embryos at 23-24 hpf stage. Whole embryo single-cell RNA-seq demonstrated reduction in the expression of hematopoietic markers associated with erythroid and myeloid development. A subset of vascular endothelial markers was also reduced in fli1b mutants. These results demonstrate fli1b role in hematopoietic and vascular development.

overall_design: Approximately 40 homozygous fli1b-/- and wild-type zebrafish embryos at 23-24 hpf stage were obtained from incrosses of  fli1b-/- or sibling wild-type parents, respectively. All cells were disaggregated and subjected to single-cell RNA-seq using 10x Genomics Chromium system.

first: Valentina

last: Laverde

first: Luiza

last: Loges


title: Single-cell RNAseq of Basal Cell Carcinoma (BCC) mouse models

status: Public on Mar 18 2025

city: Bethesda

first: Ramiro

last: Iglesias-Bartolome

address: 37 Convent Drive

department: LCMB

country: USA

institute: NIH/NCI

type: Expression profiling by high throughput sequencing

summary: Single-cell RNA sequencing of mouse tail epidermis from Basal Cell Carcinoma mouse models

overall_design: Gene expression analysis was performed from isolated tail epidermis from BCC mouse models (BCC, SmoM2, Gnas-eKO or PKI mice) with respective control mice (no tumor epidermis, Con)

first: Sarah

last: Krantz


title: Dynamic modelling of EWS::FLI1 fluctuations reveals molecular determinants of phenotypic tumor plasticity and prognosis in Ewing sarcoma

status: Public on Apr 10 2025

city: Vienna

first: Florian

last: Halbritter

address: Zimmermannplatz 10

country: Austria

institute: St. Anna Children's Cancer Research Institute (CCRI)

type: Genome binding/occupancy profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: APOE determines cell type-specific pathological landscape of Alzheimer's disease

status: Public on Mar 17 2025

city: Jacksonville

last: Zhao

state: FL

address: 4500 San Pablo Rd S

department: Neuroscience

institute: Mayo Clinic

summary: The apolipoprotein E (APOE) gene is the strongest genetic risk modifier for Alzheimer's disease (AD), with the APOE4 allele increasing risk and APOE2 decreasing it compared to the common APOE3 allele. Using single-nuclei RNA sequencing of the temporal cortex from APOE2 carriers, APOE3 homozygotes, and APOE4 carriers, we found that AD-associated transcriptomic changes were highly APOE genotype-dependent. Comparing AD with controls, APOE2 carriers showed upregulated synaptic and myelination-related pathways, preserving synapses and myelination at the protein level. Conversely, these pathways were downregulated in APOE3 homozygotes, resulting in reduced synaptic and myelination proteins. In APOE4 carriers, excitatory neurons displayed reduced synaptic pathways similar to APOE3, but oligodendrocytes showed upregulated myelination pathways like APOE2. However, their synaptic and myelination protein levels remained unchanged or increased. APOE4 carriers also showed increased pro-inflammatory signatures in microglia but reduced responses to amyloid-β pathology. These findings reveal APOE genotype-specific molecular alterations in AD across cell types.

overall_design: Single-nucleus RNA sequencing (snRNA-seq) was performed on temporal cortex samples from 60 cases. After quality control, 56 cases were retained for analysis, including 29 pathologically confirmed Alzheimer's disease (AD) cases and 27 controls. Within the AD group, 8 individuals were APOE2 carriers (E2/3), 10 were APOE3/3 homozygotes, and 11 were APOE4 carriers (E3/4 or E4/4). Among controls, 9 subjects were APOE2 carriers, 10 were APOE3/3 homozygotes, and 8 were APOE4 carriers

first: Zonghua

last: Li

first: Yuka

middle: A

last: Martens

first: Yingxue

last: Ren

first: Yunjung

last: Jin

first: Hiroaki

last: Sekiya

first: Sydney

middle: V

last: Doss

first: Naomi

last: Kouri

first: Monica

last: Castanedes-Casey

first: Trace

last: Christensen

first: Kai

last: Chen

first: Chia-Chen

last: Liu

first: Yixing

first: Alexandra

last: Soto-Beasley

first: Baayla

middle: D

last: Boon

last: A

first: Tadafumi

middle: C

last: Ikezu

first: David

last: Bennett

first: Ross

middle: R

last: Reichard

first: Ronald

last: Petersen

first: Takahisa

last: Kanekiyo

first: Owen

last: Ross

first: Melissa

middle: E

last: Murray

first: Dennis

middle: W

last: Dickson

first: Guojun

last: Bu


title: A cellular epigenetic classification system for glioblastoma

city: Philadelphia

first: Dana

last: Silverbush

state: PA

address: 421 curie Blvd

department: Cancer Biology

institute: University of Pennsylvania

type: Methylation profiling by genome tiling array

summary: The recognition of tumor heterogeneity has highlighted the necessity of examining tumor samples through the lens of single-cell genomics. In glioblastoma (GBM), a highly heterogeneous tumor, single-cell analysis is critical to assist in assessing tumor composition and in the longitudinal analysis of response to therapies. However, single-cell genomic approaches face practical challenges for broad implementation, underscoring the importance of developing deconvolution methods that may assist in the interpretation of bulk profiles and can be deployed at scale. Bulk DNA methylation data, a stable and widely used diagnostic tool in gliomas and central nervous system tumors, provides a promising substrate for deconvolution. However, the limited availability of cell state-specific references in DNA methylation, coupled with low-coverage single-cell DNA methylation data, poses significant challenges. We present a hierarchical non-negative matrix factorization approach to deconvolute bulk DNA methylation profiles, initially resolving cell types and subsequently refining cell states within a cell type. By integrating multi-omics single-cell data, we mapped DNA methylation components to their transcriptional counterparts, enabling accurate predictions of transcriptional cellular composition from bulk DNA methylation. This methodology allows the decomposition of GBM bulk DNA methylation into glial, immune, neuronal, and malignant cell types, with further distinction into malignant stem-like and malignant differentiated cell states. Our findings reveal that low cancer cell fractions can distort classification, prompting the development of an in-silico purification method to enhance diagnostic accuracy. Additionally, we provide a framework to assist in quantifying the influences of the immune micro-environment on GBM bulk classification, unmasking the underlying genetic heterogeneity and tumor subtype. Our work provides a blueprint to reconcile DNA methylation, bulk transcription-based and single-cell classifications of GBM.

overall_design: Illumina Infinium MethylationEPIC (850K)  for 11 samples of 9 GBM tumors. Out of the 11 samples 4 are FFPE and 7 Fresh, for two tumors we sequenced matched FFPE and Fresh tissue.

first: Liv

last: Jürgensen

first: Channing

middle: S

last: Pooley

first: Fabio

last: Boniolo

first: Federico

last: Gaiti

first: Mario

middle: L

last: Suvà

first: Volker

last: Hovestadt

first: Nelson

last: Freeburg


title: Modifying inhibitor specificity for homologous enzymes by machine learning

status: Public on Mar 10 2025

city: Be'er Sheva

first: Dor

middle: Shlomo

last: Gozlan

address: David Ben Gurion Blvd 1

country: Israel

institute: Ben Gurion University

summary: Selective inhibitors are essential for targeted therapeutics and for probing enzyme functions in various biological systems. The two main challenges in identifying such inhibitors lie in the extensive experimental effort required, including the generation of large libraries, and in tailoring the selectivity of inhibitors to enzymes with homologous structures. To address these challenges, machine learning (ML) is being used to improve protein design by training on targeted libraries and identifying key interface mutations that enhance affinity and specificity. However, such ML-based methods are limited by inaccurate energy calculations and difficulties in predicting the structural impacts of multiple mutations. Here, we present an ML-based method that leverages HTS data to streamline the design of selective inhibitors. To demonstrate its utility, we applied our new method to finding inhibitors of matrix metalloproteinases (MMPs), a family of homologous enzymes involved in both physiological and pathological processes. By training ML models on binding data for three MMPs (MMP-1, MMP-3, and MMP-9), we successfully designed a novel N-TIMP2 variant with a differential specificity profile, namely, high affinity for MMP-9, moderate affinity for MMP-3, and low affinity for MMP-1. Our experimental validation showed that this novel variant exhibited a significant specificity shift and enhanced selectivity compared to wild-type N-TIMP2. Through molecular modeling and energy minimization, we obtained structural insights into the variant’s enhanced selectivity. Our findings highlight the power of ML-based methods to reduce experimental workloads, facilitate the rational design of selective inhibitors, and advance the understanding of specific inhibitor-enzyme interactions in homologous enzyme systems.

overall_design: This study investigates the binding selectivity of human amyloid protein precursor inhibitor (APPI) variants toward the human serine proteases: kallikrein-6 (KLK6 ) and mesotrypsin. A yeast surface display (YSD) library containing 3.5 million APPI-3M variants, each with 0–2 amino acid mutations, was generated. These variants were subjected to pairwise selective screening using fluorescence-activated cell sorting (FACS) and labeled protease pairs to isolate variants with enhanced or diminished selectivity for specific proteases. The sorted variants were analyzed using next-generation sequencing (NGS) on the Illumina Miseq platform to identify mutations associated with differential binding selectivity.

first: Dor S.

first: Reut

last: Meiri

first: Gili

last: Shapira

first: Matt

last: Coban

first: Yaron

last: Orenstein

first: Evette

last: Radisky

first: Niv

last: Papo


title: Anti-CD3 mAb treatment reshapes infiltrating T and beta cells in the islets in autoimmune diabetes

status: Public on Mar 24 2025

city: New Haven

first: Ying

last: Wu

state: CT

address: 300 George St

institute: Yale University

summary: Anti-CD3 mAb delays or prevents type 1 diabetes (T1D) by modulating the immune mediated destruction of beta cells. Our findings described the reshaping of islet-infiltrating T cells and beta cells that lead to operational, but tenuous tolerance to autoimmune diabetes following anti-CD3 mAb treatment.

overall_design: Islets and paired pancreatic draining lymph nodes were isolated from anti-CD3 treated NOD mice in remission and processed for single-cell 5' RNA sequencing. Each scRNAseq dataset is multiplexed and contains islet cells or pancreatic draining lymph nodes of 3 mice, each individually labeled by hashtag barcode.

first: Maxwell

last: Spurrell

first: Ana

last: Lledó-Delgado

first: Songyan

last: Deng

first: Dejiang

last: Wang

first: Yang

last: Liu

first: Mahsa

last: Nouri Barkestani

first: Ana Luisa

last: Perdigoto

first: Kevan

last: Herold


title: Chronic morphine treatment leads to a global DNA hypomethylation via active and passive demethylation mechanisms in mESCs

status: Public on Mar 21 2025

city: Leioa

first: Iraia

middle: Maialen

last: Muñoa Hoyos

state: Bizkaia

address: Barriso Sarriena S/N

country: Spain

institute: Universidad del País Vasco

type: Methylation profiling by high throughput sequencing

summary: Epigenetic changes are essential for normal development and ageing, but there is still limited understanding of how environmental factors can cause epigenetic changes that leads to health problems or diseases. Morphine is known to pass through the placental barrier and impact normal embryo development by affecting the neural tube, frontal cortex and spinal cord development, and, as a consequence, delaying nervous system development. In fact, in-utero morphine exposure has shown alterations in anxiety-like behaviours, analgesic tolerance, synaptic plasticity and the neuronal structure of offspring. However, how morphine leads to abnormal neurogenesis and other physiological consequences during embryo development is still unknown. Considering that DNA methylation is a key epigenetic factor crucial for embryo development, our aim is to elucidate the role of methylation in response to morphine. Chronic morphine treatment (24h, 10μM) induces a global hypomethylation in mESC.  WGBSeq identifies 16,808 sensitive to morphine which are involved in embryo development, signalling pathways, metabolism and/or gene expression, suggesting that morphine might impact methylation levels at developmental genes. Integrative analyses between WGBSeq and RNASeq identified Tet1 as morphine-sensitive gene. Morphine increased the gene expression of Tet1, modifying the methylation levels at the promoter. On the other hand, RNASeq and qRT-PCR analyses revealed that Dnmt1 gene expression decreased after morphine treatment, without altering the methylation patter at its promoters. By MS/MS approaches confirms a decrease in DNA methylation after chronic morphine treatment, together with an increase in hydroxymethylation global levels in mESCs. In conclusion, morphine induces a global hypomethylation in mESC through different mechanisms that involves passive demethylation and a self-regulatory mechanism via active demethylation.

overall_design: Examination of methylation profiles using methylation sequencing data of control mESCs and morphine treated mESCs

first: Manu

last: Araolaza-Lasa

last: Muñoa-Hoyos

first: Itziar

last: Urizar-Arenaza

first: Ainize

last: Odriozola

first: Irune

last: Calzado

first: Mikel

last: Albizuri

first: Nerea

last: Subiran Ciudad


title: Hypercapnia induces long-term DNA methylation changes in skeletal muscle

status: Public on Mar 19 2025

city: Nutley

first: Andy

last: Madrid

state: NJ

address: 111 Ideation Way

institute: Hackensack Meridian Health

summary: Skeletal muscle dysfunction and elevated CO2 in the blood, or hypercapnia, are both associated with higher mortality in acute and chronic pulmonary diseases. Hypercapnia causes accelerated protein degradation, reduced protein synthesis, and dysfunctional myogenesis. We have recently reported that hypercapnia-induced skeletal myogenic dysfunction persists after resolution of CO2 exposure, suggesting the existence of durable cellular aberrations in previously hypercapnic cells. No data on lingering myofiber atrophy after hypercapnia resolution currently exist. Hypercapnia and age-induced skeletal muscle loss phenotypically overlap, and myogenic progenitor cells from hypercapnic mice elicit senescence-associated transcriptional programs. While aging is characterized by unique DNA methylation and other epigenetic changes, the potential association of hypercapnia and age-related differential methylation have never been investigated. In the present study, we show that hypercapnic mice elicit protracted muscle deterioration even after returning to normocapnia and demonstrate aberrant DNA methylation in comparison to animals never exposed to elevated CO2. We also show that these DNA methylation changes do not overlap with age-induced alterations.

overall_design: Whole-genome methylation sequencing of skeletal muscle tissue from mice exposed to hypercapnic conditions, six months post-hypercapnia, and mice through three aging stages


title: Description of Bacterial RNA Transcripts Detected in Mycobacterium tuberculosis – Infected Cells from Peripheral Human Granulomas using Single Cell RNA Sequencing

status: Public on Mar 04 2025

city: Salt Lake City

first: Philip

middle: J

last: Moos

state: UT

department: Pharmacology & Toxicology

institute: University of Utah

summary: Next-gen sequencing has facilitated studies of bacterial genomes and uncovered pathogen variants associated with clinically relevant phenotypes such as antibiotic resistance. However, these studies are primarily focused on bacteria cultured from patient tissues, and thus, viable but non-cultivable bacteria are not assessed. This yields an incomplete genetic state of bacteria during infection. Nevertheless, it is possible to capture bacterial transcripts in libraries designed to amplify eukaryotic mRNA. These reads are often considered spurious or nuisance and are rarely investigated. Here we demonstrate that Mycobacterium tuberculosis (Mtb) ribosomal RNA sequences are detected in host human cells obtained from peripheral lymph node aspirates from patients infected with Mtb.

overall_design: Fine needle aspirates from periferal lymphadenopathies were collected from concenting patients arriving at the Port Moresby General Hospital (POMGH) Tuberculosis Clinic, Papua New Guinea, processed to collect the single cells, and fixed and sterilized with MeOH before returning to the University of Utah for processing for single-cell RNA sequencing using the 10X Genomics system.

first: Louis

last: Barrows


title: DMS-MaPseq chemical probing of pri-miRNA and human mRNA segments

status: Public on Mar 05 2025

city: Boston

first: Silvi

last: Rouskin

state: MA

address: 77 avenue louis pasteur

institute: Harvard Medical School

summary: Understanding macromolecular structures, such as proteins and nucleic acids, is critical for discerning their functions and biological roles. Advanced techniques like crystallography, NMR, and CryoEM have facilitated the determination of over 180,000 protein structures, all cataloged in the Protein Data Bank (PDB). This comprehensive repository has been pivotal in developing deep learning algorithms for predicting protein structures directly from sequences. In contrast, RNA structure prediction has lagged, primarily due to a scarcity of RNA structural data. Here, we present the secondary structures of 1098 pre-miRNAs and 1456 human mRNA regions determined through chemical probing. We develop a novel deep learning architecture, inspired from the Evoformer model of Alphafold and traditional architectures for secondary structure prediction. This new model, called eFold, was trained on our newly created database and over 100,000 secondary structures across multiple sources. We benchmark eFold on a set of challenging RNA structures and show that both our new architecture and dataset contributes to increasing the prediction performance, and outperforming similar end-to-end methods.This result reveals that merely expanding the database size is inadequate; rather, incorporating a greater diversity and complexity of structures is crucial for enhancing performance.

overall_design: DMS MaP seq method in vitro. 1.5% DMS

first: Fede

last: Fuchs

first: Dragui

last: Salazar


title: Bulk RNAseq of Gpr161 epidermal knockout mice.

first: Ramiro

last: Iglesias-Bartolome

summary: RNA sequencing (RNAseq) of mouse tail keratinocytes from Gpr161 epidermal knockout mice (K14CRE)

overall_design: Gene expression analysis was performed from isolated tail keratinocytes from Gpr161 epidermal knockout mice or control mice.

first: Sarah

last: Krantz


title: MicroRNA signatures of CD4+ T cell subsets in healthy and multiple sclerosis subjects determined by small RNA-sequencing

city: Paris

first: Marc

last: Monot

address: 28 rue du docteur roux

country: France

institute: Institut Pasteur

type: Non-coding RNA profiling by high throughput sequencing

summary: Diverse CD4+ T cell subsets with specialized functions operate at different phases of the immune response. Among these are naïve, central memory (CM), effector memory (CM), and regulatory (Treg) cells that have been phenotypically and functionally characterized. To expand current knowledge and identify highly subset-selective markers, we have profiled miRNAs using small RNA-sequencing in FACS-sorted CD4+ T cell subsets from healthy subjects and untreated patients with relapsing-remitting multiple sclerosis (RRMS). Genomic clustering and abundance of the miRNAs were also investigated. From the 60 most differentially expressed miRNAs, broad signatures and highly selective core signatures were identified for naïve and memory CD4+ T cells at homeostasis, while miR-146a-5p was strongly upregulated in Treg cells. In line with other studies, a 5-miRNA core signature was identified for naïve cells (miR-125b-5p, miR-99a-5p, miR-365a-3p, miR-365b-3p, miR-193b-3p). Supporting the progressive memory T cell differentiation model, a number of miRNAs were upregulated in CM and EM cells and, notably, at higher level in EM cells. This was particularly the case for a miRNA core of 8 miRNAs from miR-23a~27a~24-2, miR-23b~27b~24-1, miR-221~222 clusters, and miR-22-3p, miR-181c-5p, with highly abundant miR-24-3p. In addition, we detected activation of the large ChrXq27.3 miR-506~514 cluster in EM cells. Interestingly, most of the miRNAs that were enriched in EM cells were shown to negatively regulate cell proliferation and survival. Finally, we found that the miRNA core signatures of naïve and memory CD4+ T cells were conserved in RRMS patients. Only few miRNAs were quantitatively modified, among these, miR-1248 was downregulated and may be a candidate disease marker.

overall_design: We have profiled miRNAs using small RNA-sequencing in FACS-sorted CD4+ T cell subsets from healthy subjects and untreated patients with relapsing-remitting multiple sclerosis (RRMS)

first: Frédérique

last: Michel


title: Bulk RNAseq of Basal Cell Carcinoma (BCC) mouse models

summary: RNA sequencing (RNAseq) of mouse tail keratinocytes from Basal Cell Carcinoma mouse models

overall_design: Gene expression analysis was performed from isolated tail keratinocytes from BCC mouse models (BCC, SmoM2, Gnas-eKO or PKI mice) with respective control mice (no tumor epidermis, Con)


title: Aerobicity stimulon in Escherichia coli revealed using multi-scale computational systems biology of adapted respiratory variants

city: Mumbai

first: Amitesh

last: Anand

state: Maharashtra

address: Homi Bhabha Road

department: DBS

country: India

institute: TIFR

summary: Respiratory variants of E. coli were evolved in succinate and glycerol as carbon sources, to understand their metabolic flexibility and rewiring to attain energy homeostatis.

sample_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: Growth of E. coli WT MG1655 and the respiratory variants was performed in their respective evolution media, i.e. minimal media supplemented with succinate (2g/L sodium succinate dibasic) and glycerol (0.2%) as the carbon source. The strains were grown at 37C in two biological replicates for each strain. Total RNA was extracted from the mid-log phase culture for RNA-seq library preparation.


title: Transcriptomic profiling of signet ring cell precursors of diffuse-type gastric cancer informs latent phenotype

status: Public on Feb 21 2025

city: BETHESDA

first: Cihan

last: Oguz

state: MD

address: NIHBC 29B BG RM 1NN20, 9000 ROCKVILLE PIKE

institute: NIH

summary: Germline CDH1 loss-of-function mutations are causally linked to an increased lifetime risk of diffuse gastric cancer (DGC). Early, multifocal signet ring cell (SRC) lesions are ubiquitous among CDH1 variant carriers, yet only a subset of patients will develop advanced DGC. Transcriptomic profiling was performed to establish the molecular phenotype of early SRC lesions.

overall_design: We performed single cell RNA sequencing (scRNA-Seq) of normal-appearing gastric mucosa that contained SRC lesions from three patients that contributed a total of 20 replicates (eight replicates each from patients 1 and 2, and four replicates from patient 3).

first: Amber

middle: F

last: Gallanis

first: Lauren

last: Gamble

middle: G

last: Samaranayake

first: Noemi

last: Kedei

first: Maria

middle: O

last: Hernandez

first: Madeline

last: Wong

first: Desiree

last: Tillo

first: Benjamin

last: Green

first: Paul

last: McClelland

first: Cassidy

last: Bowden

first: Irene

last: Gullo

first: Mark

last: Raffeld

first: Liqiang

last: Xi

first: Michael

last: Kelly

first: Markku

last: Miettinen

first: Martha

last: Quezado

first: Sun

last: Kim

first: Andrew

middle: M

last: Blakely

first: Justin

last: Lack

first: Theo

last: Heller

first: Jonathan

first: Jeremy

last: Davis


title: The role of structural vs cellular remodeling in arrhythmogenesis: personalized computer models of atrial fibrillation

status: Public on Mar 08 2025

city: Chicago

first: Igor

last: Efimov

state: Illinois

address: 303 E Superior St

institute: Northwestern University

summary: Atrial fibrillation (AF) is a progressive disease involving both structural and functional remodeling. To investigate the contribution of cell-scale functional remodeling to AF pathogenesis, we combined personalized 3D anatomical models with pathology-specific ionic models. The latter were developed using recordings in myocytes isolated from patients in sinus rhythm, paroxysmal, postoperative, and persistent AF. To quantify AF dynamics, we developed a novel algorithm for locating RDs by backtracking the conduction velocity field from the wavebreak regions. We demonstrate that our novel algorithm is at least 700 times faster than the traditional phase singularity analysis. The inducibility of simulated AF was not pathology-dependent, but pathological models demonstrate a more extensive arrhythmogenic substrate compared to the sinus rhythm. AF driver locations depend on electrophysiological remodeling; differences between pathology-specific models are explained by differences in wavebreak patterns. Specifically, RDs tend to dwell in the regions with the highest wavebreak probability.

overall_design: Right atrial appendages from human patients were collected and sequenced with RNA-Seq. Samples were grouped based on diagnosis: Sinus Rhythm (SR), postoperstive AF (poAF), paroxysmal AF (pAF), and persistent AF (persAF). Each diagnosis group had at least five patient samples.

first: Pikunov

last: Andrey

first: Syunyaev

last: Roman

first: Ali

last: Rheeda

first: Prakosa

last: Adityo

first: Boyle

last: Patrick

first: Steckmeister

last: Vanessa

first: Kutschka

last: Ingo

first: Gams

last: Anna

first: Rytkin

last: Eric

first: Voigt

last: Niels

first: Trayanova

last: Natalia

first: Efimov

last: Igor


title: Microglia and myeloid cell populations of the developing mouse retina

status: Public on Apr 08 2025

city: Durham

last: Kay

state: NC

institute: Duke University

summary: Microglia make important contributions to central nervous system (CNS) development, but the breadth of their distinct developmental functions remain poorly understood. The mouse retina has been a key model system for understanding fundamental mechanisms controlling assembly of the CNS. To gain insight into where and how microglia might influence retinal development, here we identified molecularly unique myeloid cell populations that are selectively present during development, and characterized their anatomical locations. Development-specific transcriptional states were identified using single-cell (sc) and single-nucleus (sn) RNA-sequencing (RNA-seq) across multiple timepoints: postnatal day (P)5, P10, and P17. Transcriptional states were validated in vivo by histological staining for key RNA and/or protein markers.

overall_design: For scRNA-seq, retinal myeloid cells were purified by flow cytometry from Cx3cr1-YFP mice at three timepoints: P5, P10, and P17. Two repilcates were collected at each timepoint, although one P5 replicate failed and is not included here. Each replicate comprised all of the pups from a single litter (10-14 retinas from 5-7 mice). One of the P10 replicates included a cocktail of transcriptional/translational inhibitors to suppress ex vivo gene activation. For snRNA-seq, myeloid cell nuclei were fluorescently labeled using Cx3cr1-CreER; lox-stop-lox-Sun1GFP mice. GFP+ nuclei from P5 and P10 retinas were purified by flow cytometry. One replicate (i.e. litter) was run for each snRNA-seq timepoint.

first: Sage

last: Martineau

first: Samantha

last: Zarnick

middle: N


title: From cyst to tapeworm: Transcriptome of Taenia solium in vitro activation and growth

status: Public on Feb 09 2025

city: Lima

first: Cristina

last: Guerra-Giraldez

state: Lima

address: Honorio Delgado 430

department: Facultad de Ciencias e Ingeniería

country: Peru

institute: Universidad Peruana Cayetana Heredia

summary: The cestode Taenia solium develops as a tapeworm in the human intestine, starting from a larva (cyst). Upon maturing, it produces hundreds of thousands of infectious eggs. When ingested by pigs and humans, the eggs develop as cysts that lodge in various tissues, including the brain, leading to neurocysticercosis. Despite advances in understanding cestode biology through genomic and transcriptomic studies, particularly in model organisms, much remains unknown about the activation of T. solium cysts in the human digestive tract and the events that drive the development into adult worms—the stage responsible for dispersing the parasite. We present a transcriptome generated by Next Generation Sequencing from T. solium cysts activated in culture and collected at three different in vitro growth stages, as defined by their morphology. We identified differentially expressed genes and biological processes relevant to growth phases that can be further explored with the dataset. The information is valuable for identifying genes that regulate the molecular, metabolic, and cellular events leading to parasite maturation or elements driving its transmission.

overall_design: Bulk RNA-seq was performed on pooled T. solium cysts under four conditions: non-cultured (NC), pre-evaginated (PRE), evaginated (EV), and post-evaginated (POST). Each condition was further analyzed with or without taurocholic acid supplementation (TA+ or TA-) to assess its role in promoting evagination.

first: David

last: Castaneda-Carpio

first: Renzo

last: Gutierrez-Loli

first: Jose

last: Maravi-Jaime

first: Valeria

last: Villar-Davila

first: Segundo W

last: Del Aguila


title: Dexamethasone drives macrophage repolarization linked to increased triple-negative breast cancer aggressiveness

city: Solna

first: Ivan

last: Nalvarte

address: Visionsgatan 4, J9:20

department: NVS

country: Sweden

institute: Karolinska Institute

summary: Glucocorticoids (GCs) are known for their anti-inflammatory potential, which includes macrophage polarization into an anti-inflammatory and tissue remodeling state. GCs are routinely co-administered to cancer patients to alleviate the side effects of chemotherapy. However, it is not well known if GCs can modulate tumor-associated macrophages (TAMs) to promote tumor progression. Here, we show that dexamethasone (DEX) induces dose-dependent differentiation of THP-1 monocyte-derived anti-tumorigenic (M1) macrophages into pro-tumorigenic (M2-like) macrophages, even in the presence of M1 cues, and that DEX can repolarize fully differentiated M1 macrophages into an M2-like state. These macrophages have a cytokine profile similar to the pro-tumorigenic (M2) macrophages and can stimulate the proliferation and invasion of triple-negative breast cancer (TNBC) cells in vitro. DEX treatment of an orthotopic mouse model of TNBC attenuated paclitaxel-mediated tumor growth inhibition, increased M2-like TAMs in primary tumors, and enhanced lung metastasis. Transcriptomic analysis of DEX-treated M1 macrophages revealed not only transcriptomic overlap to M2 macrophages, but to human breast cancer TAM transcriptomic data and further to a specific TAM signature associated with aggressive estrogen receptor-negative breast cancer. Our study illustrates a remarkable macrophage repolarization plasticity upon DEX exposure and warrants care in prescribing high doses of GCs to breast cancer patients, especially to those considered for chemotherapy.

first: Mohamed

last: Shamekh

first: Birgitta

last: Lindqvist


title: Male-biased gene cyp17a2 enhances virus resistance in fish by promoting STING expression and degrading viral protein

status: Public on Mar 01 2025

city: Wuhan

first: Shengchi

last: Shi

address: Donghu South Road #7

country: China

institute: Institute of Hydrobiology, Chinese Academy of Sciences

overall_design: In the EPC cell line, overexpress the control plasmid and the cyp17a2 plasmid, then add SVCV infection after 24 hours, and perform RNA-seq on both groups.

first: Longfeng

last: Lu


title: Paramutation-like behaviour of genic piRNA-producing loci in Drosophila virilis

status: Public on Mar 20 2025

city: Moscow

first: Alina

last: Bespalova

address: Vavilova, 32

department: Russian Academy of Sciences

country: Russia

institute: Engelhardt Molecular Biology Institute

summary: Piwi-interacting RNAs (piRNAs) play a crucial role in silencing transposable elements (TEs) in the germ cells of Metazoa by acting as sequence-specific guides. Originating from distinct genomic loci, called piRNA clusters, piRNA can trigger an epigenetic conversion of TE insertions into piRNA clusters by means of paramutation-like process. However, the variability in piRNA clusters' capacity to induce such conversion remains poorly understood. Here, we investigated two Drosophila virilis strains with differing capacities to produce piRNAs from the RhoGEF3 and Adar gene loci. We found that active piRNA generation correlates with high levels of the heterochromatic mark H3K9me3 over genomic regions that give rise to piRNAs. Importantly, maternal transmission of piRNAs drives their production in the progeny, even from homologous loci previously inactive in piRNA biogenesis. The subtelomeric RhoGEF3 locus, once epigenetically converted, maintained enhanced piRNA production in subsequent generations lacking the original allele carrying the active piRNA cluster. In contrast, piRNA expression from the converted Adar locus was lost in offspring lacking the inducer allele. Our findings highlight that the paramutation-like behavior of piRNA clusters is influenced not only by piRNAs but also by structural features of chromosomal regions, providing new insights into epigenetic regulation in Drosophila.

overall_design: The submission includes samples from chromatin immunoprecipitation sequencing (ChIP-seq), RNA-seq and small RNA-seq from ovaries of female Drosophila virilis. The research focuses on two distinct Drosophila virilis strains: strain 9, collected from Batumi, Georgia, and the laboratory strain 140 (eb, va).

first: Dina

last: Kulikova

first: Elena

last: Zelentsova

first: Alexander

last: Rezvykh

first: Julia

last: Guseva

first: Ana

last: Dorador

first: Justin

last: Blumenstiel

first: Mikhail

last: Evgen’ev

first: Sergei

last: Funikov


title: Single Cell Analysis of Peripheral TB-Associated Granulomatous Lymphadenitis

first: Philip

last: Moos

summary: We employed a single cell RNA sequencing (scRNA-seq) approach to describe the cells and communication networks characterizing granulomatous lymph nodes of TB patients. When mapping cells from individual patient samples, we uniformly identify several cell types that characterize granulomas. Whether high or low Mtb burden, overall we find the T cell cluster to be the most abundant. Other cell clusters that are uniformly detected, but that vary dramatically in abundance amongst the individual patient samples, are the B cell, plasma cell and macrophage/dendrocyte and NK cell clusters. When we combine all our scRNA-seq data from our current 23 patients (to gain greater n), we distinguish T, macrophage, dendrocyte and plasma cell subclusters. The sizes of these subclusters also varies dramatically amongst the individual patients. In comparing FNA composition we noted trends in which T cell populations and macrophage/dendrocyte populations were negatively or positively correlated with NK cell populations. In addition, we discovered that the scRNA-seq pipeline detects Mtb RNA transcripts and associates them with their host cell’s transcriptome, thus identifying individual infected cells. The number of infected cells also varies dramatically in abundance amongst the patient samples. CellChat analysis identified predominating signaling pathways amongst the cells comprising the various granulomas.

first: Louis

last: Barrows


title: Correlations of m6A Methylation-Related mRNAs with Thyroid Cancer

status: Public on Feb 28 2025

city: 福建省福州市

first: 瀛若

last: 林

address: 福建省福州市台江区福建医科大学附属第一医院

institute: 福建医科大学附属第一医院

summary: The specific role of m6A modification in the malignant progression of papillary thyroid carcinoma (PTC) without autoimmune thyroid disease (AITD)  remains unclear. We collected cancerous and para-cancerous tissues and associated clinical information form PTC patients without AITD to both identify differentially expressed genes and quantify m6A levels in the mRNA and ncRNA transcripts.

overall_design: We generated the following comparisons using Fold-change (FC) ≥ 1.5 or ≤ 0.5, and p < 0.05 were used to identify the differentially m6A methylated RNAs.The total RNAs were immunoprecipitated with anti-N6-methyadenosine (m6A) antibody. The modified RNAs were eluted from the immunoprecipitated magnetic beads as the “IP”. The unmodified RNAs were recovered from the supernatant as “Sup”. Two channel arrays were used for these samples but the data were analyzed as single-channel so that each sample represents either IP or supernatent and the sample table includes the normalized signal values.

first: Yingruo

last: Lin

first: Zhen

last: Jiang

first: Sheng

last: Luo

first: Huihao

last: Zhang

first: Ruhai

last: Yi


title: CCR7 Expression Distinguishes Functionally Distinct pTfh1 Subsets with Roles in Malaria-Specific Immunity

status: Public on Feb 27 2025

city: Brisbane

first: Zuleima

last: Pava Imitola

state: Queensland

address: 300 Herston road

country: Australia

institute: QIMR Berghofer

summary: T-follicular CD4 T (Tfh) cells play essential roles in antibody induction during infection and following vaccination. In humans, peripheral Tfh (pTfh) cells are commonly analysed based on expression of CXCR3 and CCR6, with different subsets of pTfh (pTfh1, pTfh2, pTfh17) associated with antibody induction in a context-dependent manner. In malaria, the specific roles of pTfh subsets in antibody development is not clear. Several studies in human malaria infection and vaccination have identified an important role of pTfh2 cells, which associate with antibody development while pTfh1 cells do not. However, in vitro studies and animal models highlight that pTfh1 cells are key drivers of cytophilic antibody development, which are protective. To dissect these contradictions, we mapped the heterogeneity of pTfh cells in healthy individuals and individuals with controlled human malaria infection using scRNAseq. We identified two, previously unidentified, pTfh1-like subsets with functional relevance, which can be defined based on CCR7 expression. CCR7pos pTfh1 cells have increased capacity to produce IL-21, whereas CCR7neg pTfh1 cells express markers of cytotoxicity. In controlled human malaria infection, we show that both CCR7pos and CCR7neg pTfh1 cells, along with Tfh2 cells, clonally expand, are transcriptionally and phenotypically activated, and are malaria specific. However, only CCR7pos pTfh1 and pTfh2 cells associated with antibody responses to infection. Our data advance our knowledge of Tfh cell diversity, and may inform the development of vaccines and therapeutics that target these key CD4 T cell responses.

overall_design: In brief, healthy malaria naive individuals underwent Controlled Human Malaria Infection with 2800 viable P. falciparum parasitized RBCs, and peripheral parasitaemia was measured at least daily by qPCR. Participants were treated with antimalarial drugs at day 8 of infection when parasitaemia reaches approximately 20,000 parasites/ml. Blood samples (from 5 studies across 6 independent cohorts) were collected prior to infection (day 0), at peak infection (day 8) and 14 or 15 and 27-36 days (end of study, EOS) after inoculation (in analyses these time points are grouped as 0, 8, 14/15 and EOS). PBMCs were isolated by Ficoll-Paque (Sigma, USA) density gradient centrifugation, isolated PBMCs were cryopreserved in 10% DMSO/FBS. Participants were healthy malaria naïve adults with no prior exposure to malaria or residence in malaria-endemic regions.  Samples used here were from samples opportunistically collected from volunteers who consented to donate blood for immunological studies within the parent clinical trial. As such, no sample size estimation was performed for this immunology study.

first: M

last: Soon

first: D

middle: A

last: Oyong

first: N

last: Dooley

first: R

last: Mukhiya

first: Z

last: Pava

last: Andrew

first: J

last: Loughland

last: McCarthy

last: Chan

last: Beeson

first: C

last: Engwerda

first: A

last: Haque

last: Boyle


title: Hi-C profiling in tissues reveals 3D chromatin-regulated breast tumor heterogeneity informing a looping-mediated therapeutic avenue

status: Public on Mar 03 2025

city: Milwaukee

first: Victor

last: Jin

state: WI

address: 8701 Watertown Plank Rd.

institute: Medical College of Wisconsin

summary: Current knowledge in three-dimensional (3D) chromatin regulation in normal and disease states was mostly accumulated through Hi-C profiling in in vitro cell culture system. The limitations include failing to recapitulate disease-specific physiological properties and often lacking clinically relevant disease microenvironment. In this study, we conduct tissue-specific Hi-C profiling in a pilot cohort of 12 breast tissues comprising of two normal tissues (NTs) and ten ER+ breast tumor tissues (TTs) including five primary tumors (PTs), and five tamoxifen-treated recurrent tumors (RTs). We find largely preserved compartments, highly heterogeneous topological associated domains (TADs) and intensively variable chromatin loops among breast tumors, demonstrating 3D chromatin-regulated breast tumor heterogeneity. Further cross-examination identifies RT-specific looping-mediated biological pathways and suggests CA2, an enhancer-promoter looping (EPL)-mediated target gene within the bicarbonate transport metabolism pathway, might play a role in driving the tamoxifen resistance. Remarkably, the inhibition of CA2 not only impedes tumor growth both in vitro and in vivo, but also reverses chromatin looping. Our study thus yields significant mechanistic insights into the role and clinical relevance of 3D chromatin architecture in breast cancer endocrine resistance.

overall_design: Hi-C experiment of 12 tissue sample, including 2 normal tissues, 5 primary breast tumor and 5 tamoxifen recurrent breast tumor tissues

first: Lavanya

last: Choppavarapu

first: Tianxiang

last: Liu

first: Kun

last: Fang


title: Braingeneers mouse organoids

status: Public on Feb 24 2025

city: Santa Cruz

first: Lon

last: Blauvelt

state: CA

address: 1156 High St

institute: University of California, Santa Cruz

summary: We characterized the cell composition of cortical organoids of dorsal forebrain identity derived from three independent cell lines using single-cell RNA sequencing to complement electrophysiological data from spontaneous neuronal activity.

overall_design: Single cell RNA sequencing was performed on 30 mouse cortical organoids from three separate cell lines (10 organoids per cell line).

first: Hunter

last: Schweiger

first: Sebastion

last: Hernandez

first: Gregory

last: Kaurala

first: Tal

last: Sharf

first: Mohammed

last: Mostajo-Radji


title: Transcriptional profiling of 6 month striatum and cortex from a mouse model of Huntington's disease (Q140) crossed with Msh2 conditional knockout mice [Msh2_6mon]

status: Public on Feb 10 2025

city: Princeton

first: Jeff

last: Aaronson

state: New Jersey

address: 155 Village Boulevard, Suite 200

department: Discovery & Systems Biology

institute: CHDI

summary: As part of collaboration between UCLA and CHDI, UCLA is creating knockouts (KOs) of 123 genes, implicated in Huntington's disease (HD) through various computational modeling efforts. Striatum and cortex were isolated from 6-month-old Msh2 flox/flox mice crossed with the Rgs9Cre+/- and Q140 knock-in (KI) HD mice and their respective controls. Transcriptomic analysis (RNASeq) was performed on 4 genotypes: Het (Q140) KI X Rgs9Cre+/-, Het (Q140) KI X Rgs9Cre+/- X Msh2fl/fl, Het (Q140) KI, and WT, 8 replicates per genotype.

overall_design:
RNASeq analysis was performed on samples from the striatum and cortex tissues of 6-month-old Het (Q140) KI X Rgs9Cre+/-, Het (Q140) KI X Rgs9Cre+/- X Msh2fl/fl, Het (Q140) KI, and WT mice. N = 8 per group (4F/4M).

UPDATE: [Mar-22-2026] The genotypes of the Samples were updated.

first: X

last: Yang

last: Wang

first: Jim

last: Rosinski


title: OTX2 controls  chromatin accessibility to direct somatic versus germline differentiation [ATAC-Seq]

status: Public on Feb 17 2025

city: Edinburgh

first: Elisa

last: Barbieri

address: 5 Little France Drive

country: United Kingdom

institute: University of Edinburgh

summary: The choice between somatic and germline fates is essential for species survival. This choice occurs in embryonic epiblast cells, as these cells are competent for both somatic and germline differentiation. Transcription factors (TFs) play a central role in this process by binding to specific chromatin loci to modulate gene expression and determine cell identity. The TF OTX2 regulates the choice between somatic and germline fates, as Otx2-null epiblast-like cells (EpiLCs) form primordial germ cell-like cells (PGCLCs) with enhanced efficiency.  However, the mechanisms by which OTX2 achieves this function are not fully characterized. Here we show that OTX2 controls chromatin accessibility to enable somatic differentiation. By performing CUT&RUN for OTX2 and ATAC-seq in wild-type and Otx2-null embryonic stem cells and EpiLCs, we identified regions where OTX2 binding opens chromatin. Enforced OTX2 expression maintains accessibility at these regions and induces opening of 4,000 additional somatic-associated regions in the presence of PGC-inducing cytokines. Once cells have acquired germline identity, the 4,000 additional somatic associated regions do not respond to OTX2 and remain closed. Our results indicate that OTX2 works in cells with dual competence for both somatic and germline differentiation to increase accessibility of somatic regulatory regions and induce the somatic fate at the expense of the germline.

overall_design: Chromatin accessiblity profiling by ATAC-seq of WT and OTX2-/- ESCs, EpiLCs, PGCLC or somatic cell aggregates. Chromatin profiling of OTX2-/- EpiLCs and aggregates after enforced expression of OTX2 using a tamoxifen-induced system.

first: Ian

last: Chambers


title: JAX Study 2- RNA-seq of male KOLF2.2J hiPSC-derived trophoblast cell lines homozygous null for seven different transcription factors

status: Public on Feb 03 2025

city: Hinxton

first: MorPhiC

last: Consortium

address: Wellcome Genome Campus

institute: EMBL-EBI

summary: This study aims to phenotype iPSC-derived trophoblast lines that contain homozygous null alleles for transcription factors expressed within the extra-embryonic lineage, with a focus on differentiation toward either primitive syncytium or extra-embryonic mesenchymal cells, depending on the TF being analyzed. Null alleles were generated using three distinct genetic engineering approaches: full protein coding region deletion (KO), critical exon deletion (CE), and insertion of a premature termination codon with frameshift (PTC+1). RNA-seq data was generated after 6 days of initiating differentiation from iPSCs. The study included the homozygous null alleles for the following transcription factors: MEIS1, MXD1, MEIS2, RUNX1, MEF2C, NCOA3, and BHLHE40. This analysis provided insight into the differences between the various CRISPR-Cas9-based approaches and the impact of loss of function for these transcription factors in trophoblast lineage differentiation.

overall_design: The study involved the generation of RNA-seq on 87 samples. Four of the transcription factors were analyzed in the primitive syncytium differentiation scheme and three in the extra-embryonic mesenchymal cell scheme. Three biological replicates were run for each genetic engineering approach for each gene. Additionally, three biological replicates were run of wild-type KOLF2.2J cells (the parental male cell line in which the KOs were generated) under the same differentiation conditions. This comprehensive design allowed for a robust comparison of genetic engineering methods and the effects of transcription factor manipulation on cell fate decisions within the extra-embryonic lineage.

first: Juliana

last: Diniz

first: Bill

last: Skarnes

first: Paul

last: Robson


title: OTX2 controls  chromatin accessibility to direct somatic versus germline differentiation [CUT&Run]


title: Segment specific loss of NFAT5 function in the kidneys

status: Public on Feb 05 2025

city: Halle

first: Bayram

last: Edemir

address: Ernst-Grube-Str. 20

country: Germany

institute: University Hospital Halle (Saale)

summary: Nuclear factor of activated T-cells 5 (NFAT5) is a transcription factor known for its role in osmotic stress adaptation in the renal inner medulla, due to osmotic gradient that is generated between renal cortex and renal inner medulla. However, its broader implications in kidney injury and chronic kidney disease (CKD) are less understood. Here we used two different Cre deleter mice (Ksp1.3-Cre and Aqp2-Cre) to generate tubule segment and even cell type specific NFAT5 deficient mice in the principal cells of the collecting duct (CD) and in the distal nephron starting in the TAL up to the CD and performed extensive gene expression profiling. In both Nfat5 knockout models, we observed massive changes in gene expression pattern, with heightened inflammatory responses and renal injury, culminating in renal fibrosis. Interestingly, inflammatory responses and fibrosis were much more prominent in the Aqp2Cre+/-Nfat5fl/fl mice that lack NFAT5 only in the collecting duct. By analyzing gene expression in the medullary and cortical regions of the kidney separately, we confirmed that the loss of NFAT5 results in kidney injury that extends beyond hypertonic areas. The correlation between the levels of inflammatory response, injury severity, and fibrosis indicates that cytokines are key mediators of stress signals in the kidney. Thus, NFAT5 is essential not only for adapting to osmotic stress but also for regulating cytokine signaling.

overall_design:
To explore the impact of spatial Nfat5-KO, we compared transcriptomic data from two different Nfat5-KO mouse models and distinguished between cortex and inner medulla. The first transgenic model involved a knockout driven by the Aqp2-promoter, causing Nfat5-KO in the principal cells of the collecting duct (CD). The second model used a knockout driven by the Ksp-promoter, leading to Nfat5-KO in the distal nephron starting in the TAL up to the CD.
We performed differentially expression analyses using data from RNA-seq of renal cortex (CTX) and inner medulla (IM) of control mice (4 from Aqp2-Cre+/-; 4 from  Nfat5fl/fl, 4 from WT) and 6 from the  in the distal nephron (TAL up to the CD) Nfat5-KO mice (KspCre+/-Nfat5fl/fl). The RNA-seq data from Nfat5-KO in the principal cells of the CD (Aqp2Cre+/-Nfat5fl/fl) and control (Aqp2-Cre+/-) were collected in a previous study (GSE195881).

first: Kristina

last: Engel

first: Vera

last: Kulow

first: Dmitry

last: Chernyakov

first: Edith

last: Willscher

first: Michael

last: Fähling


title: MSKCC- Mapping the Temporal Impact of Perturbations on Lineage Plasticity and Gene Regulation

summary: This study characterizes gene and enhancer knockout phenotypes of 30 genes in a human pluripotent stem cell (hPSC)-directed islet differentiation system. Samples were collected at five differentiation stages from hPSCs to SC-islet cells for scRNA-seq to examine transcriptional impact of the knockouts over time.

overall_design:
30 genes were edited using CRISPR-Cas9, including HHEX, GATA6, PDX1, GATA4, FOXA1, FOXA2, MNX1, NEUROD1, NKX2-2, RFX6, GSC, HNF4A, PAX6, PBX1, GLIS3, NEUROG3, ARX, OTUD5, BMPR1A, TLE3, BCOR, QSER1 , TET1, TET2, TET3, TADA2B, KDM2B, PROSER1 gene knockouts, and HHEX, ONECUT1 and NANOG enhancer deletions. The CRISPR-Cas9 edited lines were generated in two hPSC backgrounds (HUES8 and H1 human embryonic stem cells), with each genotype represented by multiple clones. The clonal lines were individually barcoded (using the expressed LARRY barcodes) and pooled for differentiation. Samples were collected at five differentiation stages as follows, ESC, definitive endoderm (DE, Day 3), Posterior foregut (PFG or PP1, Day 7), pancreatic progenitor (PP, Day 11), islet-like (SC-islet, Day 18) for scRNA-seq. In addition, WT cells in H1 and HUES8 backgrounds were differentiated and sequenced in parallel at 17 time points: day -1 (ESC), 3, 5, 7, 9, 10, 11, 12, 13, 14, 15, 16, 18, 21, 25, 26, 35.
Please note that processed .h5ad data was generated from both *transcriptome and *larry samples, and is linked to the corresponding *transcriptome records.

first: Danwei

last: Huangfu

first: Dingyu

last: Liu

first: Dapeng

first: Renhe

last: Luo


title: Transcriptional profiling of 12 month striatum and cortex from a mouse model of Huntington's disease (Q140) crossed with Msh2 heterozygous or homozygous conditional knockout mice [Msh2_12mon]

first: Jeff

last: Aaronson

summary: Striatum and cortex were isolated from 12-month-old Msh2 flox/flox or Msh2 +/flox mice crossed with the Rgs9Cre+/- and Q140 knock-in (KI) HD mice and their respective controls. Transcriptomic analysis (RNASeq) was performed on 9 genotypes: Msh2fl/fl, Het (Q140) KI X Msh2fl/fl, Rgs9Cre+/-, Het (Q140) KI X Rgs9Cre+/-, Het (Q140) KI X Rgs9Cre+/- X Msh2+/fl, Rgs9Cre+/- X Msh2fl/fl, Het (Q140) KI X Rgs9Cre+/- X Msh2fl/fl, Het (Q140) KI, and WT; 6-8 replicates per genotype.

overall_design:
RNASeq analysis was performed on samples from the striatum and cortex tissues of 12-month-old Msh2fl/fl, Het (Q140) KI X Msh2fl/fl, Rgs9Cre+/-, Het (Q140) KI X Rgs9Cre+/-, Het (Q140) KI X Rgs9Cre+/- X Msh2+/fl, Rgs9Cre+/- X Msh2fl/fl, Het (Q140) KI X Rgs9Cre+/- X Msh2fl/fl, Het (Q140) KI, and WT mice. N = 6-8 per group (2-5F/3-5M).

UPDATE: [Mar-22-2026] The genotypes of the Samples were updated.

first: X

last: Yang

last: Wang

first: Jim


title: NWU- Bulk RNA seq timecourse of KOLF2.2J hiPSC after Auxin-induced protein degradation of specific gene products

first: MorPhiC

summary: This study investigates the impact of 2 gene products (RBBP4, CHD1) on transcriptional programming in growing hiPSC at 0, 6 & 24 hours after acute protein depletion. Crispr editing was used to add an auxin-dependent degron to the C-termini of gene products expressed from the endogeneous loci and to insert an auxin-responsive degradation E3 ligase to the AAVS locus. Rapid (<30 min), specific target protein degradation in KOLF2.2J hiPSCs was induced by a synthetic auxin derivative (5-Ad-IAA) and bulk RNA-seq was performed at timepoints to define early (6 hr) and later (24 hr) downstream transcriptional dependencies.

overall_design: Crispr editing was used to introduce an auxin dependent E3 ligase (atOsTir1) into the AAVS locus of the parental KOLF2.2J hiPSC line.    Cells were subsequently Crispr-edited to create independent clones with an auxin-specific degron added to the C-terminus of individual target gene products expressed from endogenous loci. Rapid degradation of target proteins was induced by treatment with the synthetic auxin derivative, 5-Ad-IAA, and protein loss was confirmed by Western blotting. Control cells were treated with DMSO. Bulk RNA-seq was performed in duplicate at 0 hr, 6hr, and 24 hr to evaluate transcriptional responses.

first: Mazhar

last: Adli

first: Tao

last: Bai

first: De

last: Xing

first: Ozlem

last: Neyisci

first: Hengqiang

last: Zhao

first: Xintong

last: Chen


title: Spatial transcriptomic profiling of signet ring cell precursors of diffuse-type gastric cancer

first: Cihan

last: Oguz

summary: Germline CDH1 loss-of-function mutations are causally linked to an increased lifetime risk of diffuse gastric cancer (DGC). Early, multifocal signet ring cell (SRC) lesions are ubiquitous among CDH1 variant carriers, yet only a subset of patients will develop advanced DGC. Transcriptomic profiling was performed to establish the molecular phenotype of early SRC lesions.

overall_design: We performed NanoString GeoMx digital spatial profiling (DSP) using the whole transcriptome atlas (WTA) panel to characterize gastric tissue regions of interest (ROIs). Early tumor stage (T1a) gastric epithelial (T1a_NEP) and SRC lesion (T1a_SRC) ROIs were captured from total gastrectomy samples of four patients with germline CDH1 P/LP variants. In four patients with advanced DGC found on gastrectomy explants, ROIs were selected from gastric epithelial (GC_NEP), superficial lamina propria (GC_sup), as well as from submucosa and deeper (GC_deep).

first: Amber

last: Gallanis

first: Lauren

last: Gamble

first: Sarah

last: Samaranayake

first: Noemi

last: Kedei

first: Maria

last: Hernandez

first: Madeline

last: Wong

first: Desiree

last: Tillo

first: Benjamin

last: Green

first: Paul

last: McClelland

first: Cassidy

last: Bowden

first: Irene

last: Gullo

first: Mark

last: Raffeld

first: Liqiang

last: Xi

last: Kelly

first: Markku

last: Miettinen

first: Martha

last: Quezado

first: Sun

last: Kim

first: Andrew

last: Blakely

first: Justin

last: Lack

first: Theo

last: Heller

first: Jonathan

first: Jeremy

last: Davis


title: UCSF- Bulk RNAseq of differentiation timecourse of embryoid bodies

first: MorPhiC

last: Consortium

summary: Bulk RNAseq of differentiation timecourse of embryoid bodies from KOLF2.2J AAVS1 inducible CRISPRi lines

overall_design: Aggregates were generated using StemCell Tech aggrewell media on day 0 and then aggregates were moved into suspension dishes on day 1 using DMEM+20% KSR media. EBs remained in DMEM+20% KSR throughout experiment.

first: Aaron

last: Huebner

first: Ashir

last: Borah

first: Luke

last: Gilbert


title: JAX Study 4- RNA-seq of male KOLF2.2J hiPSC-derived cortical brain organoids and extra-embryonic lineages homozygous null for six different transcription factors

summary: This study aimed to phenotype iPSC-derived cortical brain organoids that contain homozygous null alleles for transcription factors DMRTA1 and TBR1 and to phenotype iPSC-derived extra-embryonic cell lineages that contain homozygous null alleles for transcription factors OVOL1, PITX1, LMO2, and ZBTB7C. Null alleles were generated by the insertion of a premature termination codon with frameshift (PTC+1). This analysis provided insight into the impact of loss of function for these transcription factors in the respective cell lineages.

overall_design: The study involved the generation of RNA-seq on 28 samples, comprising 10 differentiated into cortical brain organoids (with dorsal forebrain patterning) and 18 differentiated into the trophoblast lineage, promoting the formation of a primitive syncytium. Three biological replicates were run for each gene KO line. Additionally, three biological replicates were run of wild-type KOLF2.2J cells (the parental male cell line in which the KOs were generated) under the same differentiation conditions.

first: Juliana

last: Diniz

first: Bill

last: Skarnes

first: Paul

last: Robson


title: TDP-43 directly inhibits RNA accumulation in neurites through modulation of RNA stability

status: Public on Feb 01 2025

city: Aurora

first: Matthew

last: Taliaferro

state: CO

department: Biochemistry and Molecular Genetics

institute: University of Colorado Anschutz Medical Campus

summary: The subcellular localization of hundreds of RNAs to neuronal projections allows neurons to efficiently and rapidly react to spatially restricted external cues. However, for the vast majority of these RNAs, the mechanisms that govern their localization are unknown. Here we demonstrate that the ALS-associated RNA binding protein TDP-43 primarily acts to keep RNAs out of neuronal projections. Using subcellular fractionation and single molecule FISH we find that TDP-43 loss results in the increased neurite accumulation of hundreds of RNAs. These RNAs were highly enriched for known TDP-43 binding sites, suggesting that TDP-43 directly binds them to regulate their localization. We then identified precise regions within them that mediate their TDP-43-dependent localization and interaction with TDP-43 using high-throughput functional assays in cells and high-throughput binding assays in vitro. We found that these regions also mediated TDP-43-dependent RNA instability, identifying the mechanism by which TDP-43 regulates RNA localization. ALS-associated mutations in TDP-43 resulted in similar RNA mislocalization phenotypes as did TDP-43 loss in human iPS-derived motor neurons. These findings establish TDP-43 as a direct negative regulator of RNA abundance in neurites and suggest that mislocalization of specific transcripts may occur in ALS patients.

overall_design: We profiled subcellular transcriptomes of neuronal cells with and without TDP-43 and used high-throughput techniques to identify cis-elements within transcripts that mediate their TDP-43-dependent RNA localization patterns.


title: JAX Study 6- RNA-seq of female WIBJ2 hiPSC-derived cortical brain organoids and extra-embryonic lineages homozygous null for five different transcription factors

summary: This study aimed to phenotype iPSC-derived cortical brain organoids that contain homozygous null alleles for transcription factor PAX6 and to phenotype iPSC-derived extra-embryonic cell lineages that contain homozygous null alleles for transcription factors PPARG, EPAS1, GCM1, and GRHL1. Null alleles were generated by the insertion of a premature termination codon with frameshift (PTC+1). This analysis provided insight into the impact of loss of function for these transcription factors in the respective cell lineages.

overall_design: The study involved the generation of RNA-seq on 27 samples, comprising cells differentiated into cortical brain organoids (with dorsal forebrain patterning) and differentiated into the trophoblast lineage, promoting the formation of a primitive syncytium. Three biological replicates were run for each gene KO line. The hypoxia-inducible factor (EPAS1) was differentiated under both 20% oxygen and 3% (~ level present at peri-implantation) oxygen concentrations. Additionally, three biological replicates were run of wild-type WIBJ2 cells (the parental female cell line in which the KOs were generated) under the same differentiation conditions.


title: JAX Study 7- RNA-seq of male KOLF2.2J hiPSC-derived differentiated cells for lines in which the original mutation to create null alleles was reverted back to wildtype

summary: This study aimed to phenotype iPSC-derived cortical brain organoids that contained corrections (reversions to wt) for homozygous null alleles for transcription factor PAX6 and to phenotype iPSC-derived extra-embryonic cell lineages that had contained homozygous null alleles for transcription factors PPARG, EPAS1, GCM1, and GRHL1. The original homozygous mutant lines were created by introducing a premature termination codon and frameshift (PTC+1) approach. Here in these clones, these mutations were reverted back to wt and restored gene function. The homozygous null alleles for the five genes studied here showed pronounced transcriptomic changes compared to the parental line, the data provided here indicate, upon reversion of the PTC+1 mutations back to wt, the normal wt transcriptomic response was largely restored indicating specificity of the engineered alleles in manipulating the respective gene’s function.

overall_design: The study involved the generation of RNA-seq on 30 samples, comprising cells differentiated into cortical brain organoids (with dorsal forebrain patterning) and differentiated into the trophoblast lineage, promoting the formation of a primitive syncytium. Three biological replicates were run for each gene KO line. The hypoxia-inducible factor (EPAS1) was differentiated under both 20% oxygen and 3% (~ level present at peri-implantation) oxygen concentrations. Additionally, three biological replicates were run of wild-type KOLF2.2J cells (the parental male cell line in which the KOs were generated) under the same differentiation conditions.


title: JAX Study 1 and Study 2 - RNA-seq of male KOLF2.2J hiPSC-derived trophoblast cell lines homozygous null for 14 different transcription factors

summary:
This SuperSeries is composed of the SubSeries listed below.
This study aims to phenotype iPSC-derived trophoblast lines that contain homozygous null alleles for transcription factors expressed within the extra-embryonic lineage, with a focus on differentiation toward either primitive syncytium or extra-embryonic mesenchymal cells, depending on the TF being analyzed. Null alleles were generated using three distinct genetic engineering approaches: full protein coding region deletion (KO), critical exon deletion (CE), and insertion of a premature termination codon with frameshift (PTC+1). RNA-seq data was generated after 6 days of initiating differentiation from iPSCs. The study included the homozygous null alleles for the following transcription factors: GRHL1, POU2F3, EPAS1, FOSB, GCM1, PPARG, and ISL1, MEIS1, MXD1, MEIS2, RUNX1, MEF2C, NCOA3, and BHLHE40. Notably, the hypoxia-inducible factor EPAS1 was evaluated under both 20% oxygen and 3% (~ level present at peri-implantation) oxygen concentrations. This analysis provided insight into the differences between the various CRISPR-Cas9-based approaches and the impact of loss of function for these transcription factors in trophoblast lineage differentiation.

overall_design: Refer to individual Series


title: JAX Study 4 and Study 5: 17 genes knocked out in extra-embryonic and cortical brain organoids

summary: This study investigates the effects of knocking out 17 genes - DMRTA1, TBR1, OVOL1, PITX1, LMO2, ZBTB7C, HEY1, MBNL2, DLX4, DLX5, DLX6, HOPX, MAFF, TWIST2, MBNL3, SIX3 and NR2F1, by inserting premature termination codons (PTCs) in primitive syncytium and cortical brain organoid model systems derived from the KOLF2.2J iPSC line. Bulk RNA-seq analysis was performed to examine the transcriptional consequences of these gene knockouts, providing insights into their roles in brain development and cellular differentiation in organoid models


title: JAX Study 5- RNA-seq of male KOLF2.2J hiPSC-derived cortical brain organoids and extra-embryonic lineages homozygous null for nine different transcription factors and one RNA splicing factor

summary: This study aimed to phenotype iPSC-derived cortical brain organoids that contain homozygous null alleles for transcription factors SIX3 and NR2F1 and to phenotype iPSC-derived extra-embryonic cell lineages that contain homozygous null alleles for transcription factors HEY1, HOPX, MAFF, TWIST2, DLX4, DLX5, and DLX6 and the splicing factor MBNL2. Null alleles were generated by the insertion of a premature termination codon with frameshift (PTC+1). This analysis provided insight into the impact of loss of function for these transcription factors in the respective cell lineages.

overall_design: The study involved the generation of RNA-seq on 48 samples, comprising 12 differentiated into cortical brain organoids (with dorsal forebrain patterning) and 36 differentiated into the trophoblast lineage, promoting the formation of a primitive syncytium. Three biological replicates were run for each gene KO line. Additionally, three biological replicates were run of wild-type KOLF2.2J cells (the parental male cell line in which the KOs were generated) under the same differentiation conditions.


title: Preoperative exercise induces anti-tumor Kupffer cells to prevent surgical stress-promoted colorectal cancer liver metastasis

city: CHARLOTTESVILLE

first: Chengli

last: Shen

state: Virginia

address: 345 Crispell Dr.

institute: University of Virginia

summary: We have employed single-cell RNA sequencing using the 10x Genomics platform to investigate the immune cell distribution in the liver of Sedentary (Sed) and preoperative exercise (PEx) mice following surgical stress-promoted colorectal cancer liver metastasis model. The liver immune microenvironment plays a critical role in tumor progression and response to PEx. This study provides a comprehensive immune landscape at single-cell resolution, offering novel insights into how PEx modulates hepatic immune cell populations to mitigate surgical stress-induced tumor progression.

overall_design: Following surgical stress-promoted colorectal cancer liver metastasis, hepatic immune cells from the livers of Sed and PEx mice were isolated using CD45+ MACS beads and analyzed through single-cell RNA sequencing to characterize immune cell distribution and functional heterogeneity.

first: Zhang

last: Yunwei

first: Shen

last: Chengli

last: Hongji


title: RNA sequencing data of mouse intestinal epithelial cells following different dietary treatments and Trichuris muris infection

city: Frederiksberg C

last: Williams

address: Dyrlægevej 100

country: Denmark

institute: University of Copenhagen

summary: Mice were fed either a synthetic control diet or an inulin-enriched diet for 14 days prior to infection. Half of the mice in each dietary group were infected with 300 T. muris eggs, and euthanised 72 hours post-infection.

overall_design: Mice (n=5 per group) were rnadomly allocated to groups. Diets were fed ad libitum. IEC were isolated from colon tissue using EDTA digestion.


title: Functional Dynamics of Dendritic Cells in Response to Cutibacterium acnes Strains Associated with Healthy and Acne-prone Skin

city: Los Angeles

first: Giorgia

last: Del Vecchio

address: 610 Charles E Young Dr S, Room 5146

institute: University of California Los Angeles

summary: The pathogenesis of acne vulgaris is multifactorial and is influenced by the presence of Cutibacterium acnes. This study investigated how different strains of C. acnes, associated with either healthy skin (CH) or acne-prone skin (CA), selectively modulate the function of three dendritic cell (DC) subtypes: Langerhans cell-derived (LCDC), monocyte-derived (moDCs), and myeloid (mDCs). Our findings revealed that all DC subtypes secreted varying levels of TGF-β, IL-1β, IL-6, IL-12p70, and IL-23 in response to C. acnes stimulation. Notably, moDCs exhibited the highest cytokine secretion in response to CA-ribotypes, indicating their potent immune responsiveness. Conversely, LCDCs were the least responsive, regardless of the ribotypes. Principal component (PCA) and gene ontology (GO) analysis of bulk RNA sequencing data of DCs exposed to C. acnes identified extensive transcriptional changes, particularly in immune response pathways. moDCs showed the most pronounced transcriptional changes, aligning with their high cytokine secretion levels. Additionally, we identified specific genes upregulated by CA-ribotypes, with notable differences among the DC types, highlighting the unique contributions of each DC subtype to the skin's immune defense mechanisms against C. acnes. Overall, our study underscores the complexity of DC responses in the skin's immune environment and provides valuable insights into their roles in acne.

overall_design: LCDC moDC and mDC cells have been stimulated with selected C. acne strains for 24 hours. The total RNA has been extracted to perform bulk RNA-sequencing and define the immune signatures unique to each DC cell type.

first: NamPhuong

last: Nguyen

first: Kelsey

last: Ouyang

first: Matteo

last: Pellegrini

first: George

last: Agak


title: cited4a limits CM dedifferentiation and proliferation during zebrafish heart regeneration [Bulk RNA-seq]

status: Public on Jan 31 2025

city: Pittsburgh

first: Michael

last: Tsang

state: PA

address: 3501 Fifth Ave. BST3-5062

department: Cell Biology

institute: University of Pittsburgh

summary: Cardiac regeneration involves interplay of complex interactions between many different cell types, including cardiomyocytes. The exact mechanism that enables cardiomyocytes to undergo dedifferentiation and proliferation to replace lost cells has been under intense study. Here we report single nuclear RNA sequencing profile of the injured zebrafish heart and identified distinct cardiomyocyte populations in the injured heart. These cardiomyocyte populations indicate diverse functions that includes stress-response, myofibril assembly, proliferation and contraction. The contracting cardiomyocyte population also involves activation of maturation pathways as an early response to injury. This intriguing finding suggests that constant maintenance of distinctive terminally differentiated cardiomyocyte population is important for cardiac function during regeneration. To test this, we determined that cited4a, a p300/CBP transcriptional co-activator, is induced after injury in mature cardiomyocyte population. Moreover, loss-of-cited4a mutants showed increased dedifferentiation, proliferation and accelerated heart regeneration. Thus, suppressing cardiomyocyte maturation pathway activity in injured hearts could be an approach to promote heart regeneration.

overall_design: Wildtype and cited4a mutant adult heart regeneration study injured by ventricular amputation. Hearts were collected at 3 days post amputation (dpa). Total RNA was extracted from ventricles was used for RNA-seq experiments.

first: Rachel

last: Forman-Rubinksky


title: Single-nucleus transcriptional profiling of the placenta reveals the syncytiotrophoblast stress response to COVID-19

city: Houston

last: Keuls

email: Rachel.Keuls@bcm.edu, rkeuls@gmail.com

state: TX

address: One Baylor Plaza

department: MCB

institute: Baylor College of Medicine

summary: It is poorly understood how maternal covid-19 results in severe placental disorders. Single nuclei sequencing (sci-rna-seq3) was used to profle transcriptomic changes in the human placenta in response to maternal COVID-19.

overall_design: sci-rna-seq3 was performed on 4 placentas from healthy control patients, and 4 placentas from patients with symtpomatic COVID-19 during the third trimester, and 1 placenta from a patient with symptomatic COVID-19 in the second trimester that was harvested at term post recovery


title: Control, MGO-treated, and Paclitaxel-treated RNAseq data with matched ribosome profiling

status: Public on Feb 02 2025

city: Madison

first: Zachary

middle: T

last: Campbell

address: 1111 Highland Avenue

department: Anesthesiology

institute: University of Wisconsin - Madison

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


title: JAX Study 3- RNA-seq of male KOLF2.2J hiPSC-derived cortical brain organoids homozygous null for six different transcription factors

first: MorPhiC

last: Consortium

summary: This study aims to phenotype iPSC-derived cortical brain organoids that contain homozygous null alleles for transcription factors expressed in the early neuroectoderm. Null alleles were generated using three distinct genetic engineering approaches: full coding region deletion (KO), critical exon deletion (CE), and insertion of a premature termination codon with frameshift (PTC+1). RNA-seq data was generated at the indicated day post initiation of differentiation from hiPSCs. The study included the homozygous null alleles for the following transcription factors: LHX9, RFX4, PAX6, OTX1, LHX5, and FEZF2. This analysis provided insight into the differences between the various CRISPR-Cas9-based approaches and the impact of loss of function for these transcription factors in cortical brain organoid differentiation.

overall_design: The study involved the generation of RNA-seq on 179 samples at different time points of differentiation to cortical brain organoids. At least three different time points were selected for each line and the chosen time points were selected based on the known initiation of transcription of each respective transcription factor in wildtype cells within this differentiation scheme. Three biological replicates were run for each genetic engineering approach for each gene. Additionally, three biological replicates were run of wild-type KOLF2.2J cells (the parental male cell line in which the KOs were generated) under the same differentiation conditions. This comprehensive design allowed for a robust comparison of genetic engineering methods and the effects of transcription factor manipulation on brain organoid formation.

last: Diniz

last: Skarnes


title: Non-CG DNA methylation and MeCP2 stabilize repeated tuning of long genes that distinguish closely related neuron types [MERFISH]

city: St. Louis

first: Mati

middle: Tesfaye

last: Nemera

state: Missouri

address: 660 S Euclid Ave

institute: Washington University in St. Louis School of Medicine

summary: The extraordinary diversity of neuron types in the mammalian brain is delineated at the highest resolution by subtle gene expression differences that may require specialized molecular mechanisms to be maintained. Neurons uniquely express the longest genes in the genome and utilize neuron-enriched non-CG DNA methylation (mCA) together with the Rett syndrome protein, MeCP2, to control gene expression, but the function of these unique gene structures and machinery in regulating finely resolved neuron type-specific gene programs has not been explored. Here, we employ epigenomic and spatial transcriptomic analyses to discover a major role for mCA and MeCP2 in maintaining neuron type-specific gene programs that define high resolution cell types. We uncover differential susceptibility to MeCP2 loss in neuronal populations depending on global mCA levels and dissect methylation patterns and intragenic enhancer repression that drive overlapping and distinct gene regulation between neuron types. Strikingly, we show that mCA and MeCP2 regulate genes that are repeatedly tuned to differentiate neuron types at high cellular resolution, including spatially resolved, vision-dependent gene programs in the visual cortex. These repeatedly tuned genes display genomic characteristics, including long length, numerous intragenic enhancers, and enrichment for mCA, that predispose them to regulation by MeCP2. Thus, long gene regulation by the MeCP2 pathway maintains differential gene expression between closely-related neurons to facilitate the exceptional cellular diversity in the brain.

overall_design: We used MERFISH to profile RNA transcripts in MeCP2 KO/+ and WT mouse brains.

first: James

last: Moore

first: Rinaldo

middle: D

last: D'Souza

first: Nicole

last: Hamagami

first: Adam

last: Clemens

first: Diana

last: Beard

first: Alaina

last: Urman

first: Victoria

last: Rodriguez Mendoza

first: Yasmin

last: Razia

first: Travis

last: Law

first: John

last: Edwards

first: Harrison

last: Gabel


title: cited4a limits CM dedifferentiation and proliferation during zebrafish heart regeneration [snRNA-seq]

last: Tsang

overall_design: Wildtype and cited4a mutant adult heart regeneration study injured by ventricular amputation. Hearts were collected at 3 days post amputation (dpa) and uninjured controls (UI). Extracted hearts were frozen in liquid Nitrogen and nuclei were isolated as described in methods section.

last: Forman-Rubinsky

first: Brent

last: Schlegel

first: Dhivyaa

last: Rajasundaram


title: Dynamic control of transcriptional regulator BACH2 overcomes T cell dysfunction driven by tonic CAR signaling

status: Public on Jan 21 2025

first: Nathan

last: Singh

institute: Washington University School of Medicine in St Louis

summary: Chimeric antigen receptors (CARs) integrate multiple lymphocyte-derived functional domains to enable selective antigen binding and robust T cell activation in a single synthetic protein. Through various biochemical mechanisms, nearly all CARs activate intracellular signaling in the absence of antigen, referred to as “tonic signaling”. Previous work has shown that tonic signaling of CARs containing the CD28 costimulatory domain drives T cell exhaustion; in contrast, we have shown that tonic signaling of 41BB-containing CARs enhances T cell function. Using a model of tonically signaling CARs targeting the B cell antigen CD22, we undertook studies to determine the molecular determinants of the divergent impact of tonic CAR signaling on T cell fitness. We identified that tonic 41BB signaling induces activation of BACH2, a transcriptional regulator that directs stem and memory programs to maintain T cell homeostasis. Transgenic expression of BACH2 prevented the development of exhaustion and promoted memory-like programs in tonically signaling CD28-containing CAR T cells. This enhanced acute cytotoxic function but impaired durable anti-tumor function. We linked transgenic BACH2 to a degradation domain, enabling precise control of BACH2 expression and found that low-level expression of BACH2 enabled robust and persistent tumor control in multiple cancer models.

overall_design: At the conclusion of CAR T cell manufacturing, T cells were purified to enrich CAR+ cells using a truncated CD34 selection marker and BACH2+ cells using a truncated EGFR selection marker. From these purified populations, we isolated and sequenced RNA. Studies were performed in technical triplicate using cells manufactured from three independent donors.

first: Tien-ching

last: Chang

last: Warrington


title: Analysis of B cell neighborhoods via single cell spatial transcriptomics in head and neck cancer patients

status: Public on Jan 18 2025

first: Zhentao

last: Liu

address: 5051 CENTRE AVE, HCC SUITE

institute: UMPC Hillman Cancer Center

summary: Head and neck squamous cell carcinoma tissue microarrays were stained via CD20 single plex IHC and then FOVs were selected for B cell neighborhoods (including tertiary lymphoid structures) in confirmed tumor areas via H&E evaluation by a pathologist.

overall_design: In this study, 1000 plex CoSMX via Nanostring was performed on 11 patients with multiple FOVs for B cell neighborhoods

first: Tullia

middle: C

last: Bruno

first: Raja

last: Seethala


title: JAX Study 1- RNA-seq of male KOLF2.2J hiPSC-derived trophoblast cell lines homozygous null for seven different transcription factors

status: Public on Jan 30 2025

first: MorPhiC

last: Consortium

summary: This study aims to phenotype iPSC-derived trophoblast lines that contain homozygous null alleles for transcription factors expressed within the extra-embryonic lineage, with a focus on differentiation toward either primitive syncytium or extra-embryonic mesenchymal cells, depending on the TF being analyzed. Null alleles were generated using three distinct genetic engineering approaches: full protein coding region deletion (KO), critical exon deletion (CE), and insertion of a premature termination codon with frameshift (PTC+1). RNA-seq data was generated after 6 days of initiating differentiation from iPSCs. The study included the homozygous null alleles for the following transcription factors: GRHL1, POU2F3, EPAS1, FOSB, GCM1, PPARG, and ISL1. Notably, the hypoxia-inducible factor EPAS1 was evaluated under both 20% oxygen and 3% (~ level present at peri-implantation) oxygen concentrations. This analysis provided insight into the differences between the various CRISPR-Cas9-based approaches and the impact of loss of function for these transcription factors in trophoblast lineage differentiation.

overall_design: The study involved the generation of RNA-seq on 95 samples. Five of the transcription factors were analyzed in the primitive syncytium differentiation scheme and two in the extra-embryonic mesenchymal cell scheme. Three biological replicates were run for each genetic engineering approach for each gene. Additionally, three biological replicates were run of wild-type KOLF2.2J cells (the parental male cell line in which the KOs were generated) under the same differentiation conditions and under both 20% and, in the case of EPAS1, 3% oxygen concentrations. This comprehensive design allowed for a robust comparison of genetic engineering methods and the effects of transcription factor manipulation on cell fate decisions within the extra-embryonic lineage.

first: Juliana

last: Diniz

first: Bill

last: Skarnes

first: Paul

last: Robson


title: Research on the occurrence and progression of colorectal cancer.

status: Public on Jan 23 2025

city: chongqing

first: Ruyi

last: HANG

address: Yuzhong District

institute: Army Medical Center of PLA

summary: APX3330 suppresses the occurrence and advancement of colorectal cancer by reducing infiltration of PMN-MDSCs

overall_design: In the AOM/DSS-induced CAC model of C57BL/6 mice, RNA-seq profiling of mice treated with APX3330/saline.

first: Tianyi

last: Chen

last: Hang


title: Non-CG DNA methylation and MeCP2 stabilize repeated tuning of long genes that distinguish closely related neuron types.

first: Mati

last: Nemera

platform_organism:
Mus musculus
synthetic construct


title: Non-invasive diagnosis of non-melanoma skin cancer subtypes predicted by microRNAs recovered from adhesive  tape discs

status: Public on Jan 20 2025

city: Salt Lake City

first: Doug

last: Grossman

state: Utah

address: Huntsman Cancer Institute, 2000 Circle of Hope

department: Dermatology

institute: Huntsman Cancer Institute

summary: We investigated whether a tape-stripping approach could be used for analysis of microRNA (miR) expression as a non-invasive method for prediction of non-melanoma skin cancer subtypes. From direct miR sequencing of RNA recovered from adhesive tape discs applied to skin lesions prior to biopsy, we determined rank lists of miRs to predict superficial or invasive BCC or SCC. Expression of these miRs was then assessed by digital PCR of disc-derived miR from independent validation cohorts of BCC (n=24 Sup, n=38 Inv) and SCC (n=45 SCC-in-situ, n=14 Inv). Expression ratios or products of >2 miRs were superior to individual miRs in discriminating Sup from Inv BCC and SCC subtypes. Moreover, several miR combinations distinguished Sup BCC and Inv SCC from a cohort of benign keratoses that can clinically resemble these cancers.

overall_design:
Cohort identification and RNA preparation for sequencing
Prior to biopsy of lesions, an adhesive disc (D-Squame D101, Clinical and Derm) was firmly applied to the lesion multiple times with rotation to ensure the entire adhesive surface had made physical contact. Discs were immediately stored at -20 oC and then at -80 oC until processing. Whole RNA was isolated from the discs using a miRNeasy Micro Kit (217084, Qiagen) and the RNA eluate was stored at -80 oC until sequencing.
Analysis of miR sequencing
The human GRCh38 genome and gene annotation files were downloaded from Ensembl release 106 and a reference database was created using STAR version 2.7.9a (Dobin et al., 2013). A custom Perl script (smallRNA_pe_umi_extractor.pl from https://github.com/HuntsmanCancerInstitute/UMIScripts) was used to trim adapters and extract the Unique Molecular Index (UMI) from the second read of a paired-end Qiagen small RNA library with 76x51 base pair reads. The trimmed reads were aligned to the reference database using STAR options optimized for shorter miRNA reads (>=16 bases matched to the genome, <= 5% mismatches over mapped length, splicing switched off). A second Perl script (bam_umi_dedup.pl) removed PCR duplicates from the aligned BAM file and the mapped reads were assigned to annotated genes in Ensembl and to miRNAs in miRbase release 22.1 (Kozomara et al., 2019) using featureCounts version 1.6.3 (Liao et al., 2014). Differentially expressed miRNA in miRbase were identified using a 5% false discovery rate with DESeq2 version 1.36.0 (Love et al., 2014). Principal components analysis was run using the regularized log values from the top 500 variable miRNAs and ellipses were added representing the 90% confidence interval from a multivariate t-distribution.
Statistical Method for Feature Selection
Features (i.e. miRs) were identified as described previously (Fastner et al., 2024). Briefly, binomial lasso regression, as implemented in the R (version 4.3.2) package “glmnet” was used to select a group of features that separated subtypes (e.g. superficial from invasive BCC). Ten-fold cross validation was used to select the penalty parameter for the lasso regression. Low expression miRs with an adjusted mean count of less than 10 were removed from consideration.
qRT-PCR assays
RNA isolated from discs was converted to cDNA using a TaqMan™ Advanced miRNA cDNA Synthesis Kit (Thermo Fisher Scientific, A28007) and stored at -80 oC until used. Samples were read using QuantStudio™ Absolute Q™ Digital PCR System (Thermo Fisher Scientific) and analyzed using QuantStudio™ Absolute Q™ Digital PCR Software (version 6.3, Applied Biosystems) according to the manufacturer’s recommendations. Samples were diluted 1:100 in 0.1X TE buffer (Ambion, AM9849) and digital PCR was performed using a MAP16 Digital PCR kit (Ambion, A52688) according to the manufacturer’s instructions. Briefly, each reaction consisted of 2 L Absolute Q DNA Digital PCR 5X mix, 0.5 L 20X TaqMan Advanced miRNA probe (Applied Biosystems, A25576) and 7.5 L cDNA, with 15ul of Isolation Buffer added over each sample prior to sealing the wells with the provided gasket strips. Data was analyzed using Quant Studio Absolute Q digital PCR software, version 6. Threshold bars were set using the manual option and results reported as number of copies per L. Normalized expression values were obtained copy number for an individual miR by that for the invariant miR-16-5p. Combined expression scores were obtained from the products and/or quotients (based on whether miRs were “up” or “down,” respectively) of normalized expression scores of multiple miRs. Pearson correlations of normalized digital PCR values with normalized regularized log sequencing values was determined using R version 4.3.2. ROC curves and other analyses were generated using Prism software.

first: Hafeez

last: Rahman

first: David

last: Mwelwa

first: Scott

middle: R

last: Florell

first: Abigail

first: Carly

last: Becker

first: Chris

last: Stubben

first: Kenneth

last: Boucher

first: Dekker

last: Deacon

first: Robert

last: Judson-Torres

first: Douglas


title: Combinatorial phosphorylation on CTD of RNA polymerase II selectively controls transcription and export of protein-coding mRNAs

status: Public on Jan 16 2025

city: Austin

first: Qian

last: Zhang

address: 2515 Speedway

institute: The Unibersity of Texas at Austin

summary: C-terminal domain (CTD) of RNA polymerase II is crucial for recruiting transcription regulators via specific post-translational modifications (PTM), especially phosphorylation. The hypothesis of combination of PTMs, or ‘CTD code’, that can allow precise and dynamic recruitment of transcription machinery is highly attractive, yet the experimental evidence to support this hypothesis has been scarce. Here, despite lacking specific antibodies for combinatorial CTD phosphorylation, we developed an innovative approach that detects double phosphorylation patterns on the CTD in a whole-genomic fashion by leveraging the antibody masking effect with selectively removing the flanking interference. Using this method, we detected pT4pS5 double phosphosites occurring exclusively during the transcription of protein-coding genes. Furthermore, we showed that pT4pS5 marks recruit the Transcription and Export complex (TREX), which specifically facilitates mRNA processing and nucleocytoplasmic export of protein-coding mRNAs. The recruitment of TREX by pT4pS5 phosphosites is particularly important for the processing of lengthy neurogenesis-related genes. Our results provide experimental support for the notion that CTD coding system can function combinatorially and in a gene-specific manner, which encodes an exact information about the transcription of specific gene clusters. This method can be broadly applied to map all combinatorial PTM patterns on RNA polymerase II, paving the way for a deeper understanding of gene-specific transcription regulation at the molecular level.

overall_design: ChIP-Sequencing of pThr4-CTD upon treatment with active Ssu72 phosphatase or inactive enzyme (Mutant/Mut), two biological replicates per sample (HEK293 cells).

first: Svetlana

middle: B

last: Panina

first: Seema

last: Irani

first: Haley

middle: H

last: Hardtke

first: Renee

last: Stephenson

first: Brendan

last: Floyd

first: Rosamaria

middle: Y

last: Moreno

first: Edward

last: Marcotte

first: Jessie


summary:
C-terminal domain (CTD) of RNA polymerase II is crucial for recruiting transcription regulators via specific post-translational modifications (PTM), especially phosphorylation. The hypothesis of combination of PTMs, or ‘CTD code’, that can allow precise and dynamic recruitment of transcription machinery is highly attractive, yet the experimental evidence to support this hypothesis has been scarce. Here, despite lacking specific antibodies for combinatorial CTD phosphorylation, we developed an innovative approach that detects double phosphorylation patterns on the CTD in a whole-genomic fashion by leveraging the antibody masking effect with selectively removing the flanking interference. Using this method, we detected pT4pS5 double phosphosites occurring exclusively during the transcription of protein-coding genes. Furthermore, we showed that pT4pS5 marks recruit the Transcription and Export complex (TREX), which specifically facilitates mRNA processing and nucleocytoplasmic export of protein-coding mRNAs. The recruitment of TREX by pT4pS5 phosphosites is particularly important for the processing of lengthy neurogenesis-related genes. Our results provide experimental support for the notion that CTD coding system can function combinatorially and in a gene-specific manner, which encodes an exact information about the transcription of specific gene clusters.
This method can be broadly applied to map all combinatorial PTM patterns on RNA polymerase II, paving the way for a deeper understanding of gene-specific transcription regulation at the molecular level.

overall_design: Small RNA-Sequencing in T4A-CTD vs wild-type (WT) cells, two biological replicates per sample (HEK293T cells).


title: EHMT2 represses totipotency- and differentiation-associated genes in pluripotent stem cells through distinct regulatory modes [ChIP-exo]

status: Public on Jan 15 2025

city: New York

first: Christopher

middle: Mitsuo

last: Uyehara

state: NY

address: 413 E 69th St

department: Meyer Cancer Center

institute: Weill Cornell Medicine

summary: Mouse embryonic stem cells (mESCs) and other naïve pluripotent stem cells, under specific conditions, can reverse typical developmental trajectories and de-differentiate into 2-cell-like cells (2CLCs) that resemble the mammalian embryo during zygotic genome activation (ZGA). We implemented a multipurpose allele for acute protein depletion and efficient immunoprecipitation to dissect the molecular functions of EHMT2 in mESCs. This approach allowed us to define different types of EHMT2 target genes and distinct modes of chromatin engagement and repression. Most notably, EHMT2 directly represses large clusters of co-regulated gene loci that comprise a significant fraction of the 2CLC-specific transcriptome by initiating H3K9me2 spreading from distal LINE-1 elements. Conversely, the loss of EHMT2 allows the recruitment of the activator DPPA2/4 to promoter-proximal ERVs in 2CLCs, thus facilitating the mESC-to-2CLC transition. Repression of germ layer-associated transcripts by EHMT2 occurs outside of gene clusters in collaboration with ZFP462 and entails binding to non-repeat enhancers. Our data suggests that EHMT2 functions as an attenuator of the bidirectional differentiation potential of mouse naïve pluripotent stem cells and define molecular models for the gene-specific repressive activity of this essential histone methyltransferase.

overall_design: EHMT-dTAG lines were treated with DMSO for 24 hours. HA tag was used to probe for genomic occupancy of EHMT by CHIP-exo.

first: Kaushiki

last: Chatterjee

first: Kritika

last: Kasliwal

first: Effie

last: Apostolou

first: Matthias

last: Stadtfeld


title: EHMT2 represses totipotency- and differentiation-associated genes in pluripotent stem cells through distinct regulatory modes [ATAC-Seq]

overall_design: EHMT-dTAG lines were treated with DMSO  for 24 hours. Cells were harvested using trypsin. Open chromatin regions were mapped by ATAC-seq on three technical replicates per sample.


title: EHMT2 represses totipotency- and differentiation-associated genes in pluripotent stem cells through distinct regulatory modes

overall_design: Refer to individual Series


title: EHMT2 represses totipotency- and differentiation-associated genes in pluripotent stem cells through distinct regulatory modes [ULI-NChIP]

overall_design: Bulk ChIP - 100,000 cells for EHMT2-HA-dTAG targeted lines were counted and flash frozen. Sorted ChIP -  72hr DMSO and dTAG treated EHMT2-dTAG-mCherry MERVL-GFP sorted for (~100,000) mESC and 2CLCs. ULI-NChIP was conducted according to the protocol published by Brind’Amour et al. (PMID 25607992). 1ug H3K9me2 antibody ab1220 was used.