title: Sequencing of chondrocytes under dermal fibroblast co-culture

status: Public on Jan 13 2025

city: Shanghai

first: Lei

last: Shi

state: 请选择

address: 115 Jinzun Road, Pudong New Area, Shanghai

institute: Shanghai Jiaotong university

summary: Knee osteoarthritis (KOA), as a degenerative multifactorial disease, affects the quality of life and mental health of patients, and also brings a huge socioeconomic burden. Treating synovitis have shown promise as anti-inflammatory therapeutics in mitigating OA symptoms and disease progression. Here, by analysing synovial single-cell sequencing (scRNA-seq) data from KOA, we found that synovial fibroblasts (FLS) in OA synovium showed a distinct pro-inflammatory phenotype. We collected synovial tissue from patients with clinical OA as well as from healthy donors, and histological examination was consistent with findings in scRNA-seq. Inspired by recent cross-tissue fibroblast lineage studies, we identified by sequencing that healthy FLS in synovial tissues share transcriptome-level similarities with dermal fibroblasts (DFb). Subsequently, we revealed the local as well as systemic distribution of intra-articular injected DFbs by constructing/extracting two types of rat fibroblasts (luciferase DFbs as well as GFP DFbs). The results demonstrate that DFbs can be locally retained in the synovium for up to three weeks following targeted engrafting on it. And intra-articular injection does not result in DFbs migration to vital organs or the occurrence of histological changes in these organs. A rat model of KOA was constructed by anterior cruciate ligament transection (ACLT) in order to study the therapeutic effect of DFbs on KOA. After injection, the rats showed improvement in painful gait. In addition, histological as well as imaging results showed reduced synovitis and improvement in articular cartilage. Finally we verified the protective effect of DFbs on cytokine-stimulated chondrocytes in a co-culture system.

overall_design: In order to achieve this objective, a co-culture experiment was conducted in which extracted primary chondrocytes were combined with primary DFbs in transwell chambers. IL-1β (10ng/ml) was added 24 h after cell attachment. After 36 h and 72 h of IL-1β addition, the chondrocytes from the lower chamber were obtained for bulk RNA sequencing

first: Kewei

last: Rong

first: Kexin

last: Liu

first: Jie

last: Zhao


title: Sequencing of dermal fibroblasts and synovial fibroblasts cultured in vitro

country: China

type: Expression profiling by high throughput sequencing

overall_design: Three main groups of cells were involved in this experiment. Group D refers to dermal fibroblasts cultured in vitro. Group HF refers to synovial fibroblasts of healthy donor origin cultured in vitro. Group OAFILB refers to synovial fibroblasts of OA patient origin cultured in vitro with IL-1β stimulation.


title: LncRNA CARINH regulates expression and function of innate transcription factor IRF1 in macrophages [RNA-Seq]

status: Public on Jan 03 2025

city: New York

first: Coen

last: van Solingen

state: NY

address: 435 E30th street SB 704

department: Medicine/Cardiology

country: USA

institute: NYU Grossman School of Medicine

summary: The antisense lncRNA CARINH acts as an additional regulatory layer of antiviral immunity through coordinated regulation of the transcription factor IRF1 and downstream interferon-stimulated genes.

overall_design: We used three replicate samples of BMDM cells from WT mice and Carinh-KO mice. Either group treated with IFN for 8 hours or vehicle.

first: Yannick

last: Cyr

first: Florencia

last: Schlamp

first: Kathryn

middle: J

last: Moore


title: LncRNA CARINH regulates expression and function of innate transcription factor IRF1 in macrophages [RNA-Seq 2]

overall_design: Mice were inoculated with influenza strain PR8 and were euthanized 4 days later. Lungs were harvested and snapfrozen. Four mice of each groups were used.


title: RNAseq of CD4 T cells from 83 individuals with musculoskeletal compains

status: Public on Jan 02 2025

city: Gothenburg

first: Maria

middle: I

last: Bokarewa

state: VG

address: Guldhedagatan 10A

department: Rheumatology and Inflammation Research

country: Sweden

institute: Gothenburg University

summary: RNAseq of CD4 cells from 83 individuals with musculoskeletal complains (age 51y, range 19-86. male 22%) were collected at the Rheumatology Clinic during 2022-2023 . Cells were activated with anti-CD3 for 48 hours.

overall_design: CD4 cells were isolated by positive selection on Dynabeads (cat.11331D) from human PBMC. Cells were stimulated with anti-CD3 (OKT3, Sigma-Aldrich)  for 48h

first: Karin

middle: M

last: Andersson

first: Malin

middle: C

last: Erlandsson

first: Eric

last: Malmhäll-Bah


title: Autism mutations rewire protein interaction networks to drive neurodevelopmental pathology

status: Public on Dec 25 2024

city: San Francisco

first: Kelsey

last: Hennick

state: CA

address: 1651 4th St room 521A

institute: UCSF

summary: Systematic mapping of protein-protein interaction (PPI) networks and determining how causal mutations rewire them in autism spectrum disorder (ASD) provide a powerful framework for uncovering disease mechanisms and therapeutic opportunities. Using affinity purification-mass spectroscopy (AP-MS), we systematically mapped PPIs for 100 high-confidence ASD genes, uncovering over 1,800 interactions. By assessing the impact of pathogenic missense mutations, leveraging AlphaFold, and validating key findings in human-derived model systems, we identify marked convergence onto shared protein complexes in the wild-type state and convergent PPI rewiring driven by independent mutations. For example, distinct patient-derived variants in FOXP1 disrupt its interactions with FOXP4, leading to changes in cortical neurogenesis and neural activity in brain organoids. Overall, these findings link genetic variation to protein networks and convergent neurodevelopmental dysfunction in ASD.

overall_design: FOXP1 R513H: Forebrain organoids were dissociated at D111 and split into two technical replicates for each sample (WT and R513H) for scRNA-seq. FOXP1 L327P: Two biological replicates for WT/WT and L327P/WT forebrain organoids were dissociated at D111 for scRNA-seq.

first: Tomasz

last: Nowakowski


title: Single cell transcriptomic profiling of human CD19+ B cells enriched for CD5+ Innate-like B cells

status: Public on Dec 04 2024

city: East Lansing

first: Rance

last: Nault

state: United States

address: 1355 Bogue St. B403

department: Department of Pharmacology and Toxicology

institute: Michigan State University

summary: Aryl hydrocarbon receptor is a ligand-activated transcription factor whose activation can regulate B cell-mediated immune responses. Given the diverse nature of human B cell subsets, scRNA-seq transcriptomic profiling was used to ascertain the heterogeneity of AHR expression in discrete B cell subpopulations. We identified a unique cell cluster enriched in expression of AHR and CD9. Further, these cells were diminished in their expression of canonical B cell-associated genes and enriched with a myeloid cell-specified gene signature. These results were confirmed in vitro as CD14 expressing B cells were readily detectable in human donor PBMC and expressed significantly more AHR protein. Following activation in vitro, CD9+ B cells secreted IgM, IL-10, TGFβ1, and several myeloid-associated pro-inflammatory mediators, specifically CXCL8. These results demonstrate CD14 expression by B cells is readily detectable in healthy human donors and their release of CXCL8 could implicate these cells in inflammatory diseases and autoimmunity.

overall_design: PBMC were isolated by ficoll density gradient from 2 anonymous platelet donor leukapheresis packs naïve B cells were isolated by magnetic assisted cell sorting via negative selection. CD5 expressing B cells were enriched by positive selection for CD5 protein and then added back into a 1:1 ratio with CD5 negative B cells. RNA was extracted and then libraries were prepared using the 10X Genomics Chromium Single Cell 3’ v3 kit and Chromium i7 Multiplex Kit.

first: Lance

middle: K

last: Blevins

first: Norb

middle: E

last: Kaminski


title: Structure-guided development of high-performance antibodies targeting site-specific post-translational modifications

status: Public on Dec 02 2024

city: Chicago

first: Rohan

middle: Nishant

last: Shah

state: IL

address: CLSC 856, 920 E. 58th Street

institute: University of Chicago

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Post-translational modifications (PTMs) of proteins play critical roles in regulating many cellular events. Antibodies targeting site-specific PTMs are essential tools for detecting and enriching PTMs at sites of interest. However, fundamental difficulties in molecular recognition of both PTM and surrounding peptide sequence have hindered the efficient generation of highly specific anti-PTM antibodies. Furthermore, the widespread use of potentially inconsistent, non-renewable and molecularly undefined antibodies presents experimental challenges thought to contribute to the reproducibility crisis in biomedical research. In this study, we describe the structure-guided development of a platform that efficiently generates potent and selective recombinant antibodies to PTMs that are molecularly-defined and infinitely renewable. Our platform is built on our previous discovery of an unconventional binding mode of anti-PTM antibodies, antigen clasping, where two antigen binding sites cooperatively clasp  a single antigen, creating extensive interactions with the antigen and leading to high selectivity and potency. We designed the platform that generates clasping antibodies with two distinct binding units, resulting in efficient generation of antibodies to a set of trimethylated histone H3  with high levels of specificity and affinity. Performance comparison in chromatin immunoprecipitation (ChIP), a common application in epigenomics, revealed that an anti-H3K27me3 clasping antibody exhibited superior specificity to a widely-used conventional antibody and captured symmetric and asymmetric nucleosomes in a less biased manner. We further generated clasping antibodies to phosphotyrosine antigens by using the same principle. These results suggest the broad applicability of our platform to generating high-performance clasping antibodies to diverse PTMs.

overall_design: Internally calibrated chromatin immunoprecipitation (ICeChIP) against H3K27me3 using commercially available antibody (Cell Signaling Technologies C36B11) or 3C2-H2 clasping Fab.

first: Mariapia

last: Riso

middle: N

first: Akiko

last: Koide

first: Alexander

last: Ruthenburg

first: Shohei

first: Takamitsu

last: Hattori


title: Single cell gene expression profile of cells isolated from human aortic valve leaflets.

status: Public on Dec 01 2024

city: La Jolla

first: Talia

last: Baddour

state: California

address: 9500 Gilman Dr.

department: Bioengineering

institute: University of California San Diego

summary: Aortic valve cell  heterogeneity increases during Aortic Valve Stenosis (AVS) progression. We used single cell RNA sequencing (scRNA-seq) to characterize valve cell phenotype in AVS patients.

overall_design: Whole cells were isolated from aortic valve leaflets of age and disease-matched AVS patients. We chose one male and one female patient.

first: T

first: B

last: Aguado


title: Paired CRISPR screens to map gene regulation in cis and trans

status: Public on Nov 28 2024

city: NEW YORK

first: Xinhe

last: Xue

state: New York

address: 101 Avenue of the Americas

institute: New York Genome Center/New York University

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

overall_design: Refer to individual Series


title: Alpha-ketoglutarate dehydrogenase is a therapeutic vulnerability in acute myeloid leukemia [RN2_shOGDH]

status: Public on Nov 27 2024

first: Yu-Jui

last: Ho

address: 1275 York Avenue

institute: Memorial Sloan Kettering Cancer Center

summary: Perturbations in intermediary metabolism contribute to the pathogenesis of acute myeloid leukemia (AML) and can produce therapeutically actionable dependencies. Here, we probed whether alpha-ketoglutarate (aKG) metabolism represents a specific vulnerability in AML. Using functional genomics, metabolomics, and mouse models, we identified the aKG dehydrogenase complex, which catalyzes the conversion of aKG to succinyl CoA, as a molecular dependency across multiple models of adverse-risk AML. Inhibition of 2-oxoglutarate dehydrogenase (OGDH), the E1 subunit of the aKG dehydrogenase complex, impaired AML progression and drove differentiation. Mechanistically, hindrance of aKG flux through the tricarboxylic acid (TCA) cycle resulted in rapid exhaustion of aspartate pools and blockade of de novo nucleotide biosynthesis, while cellular bioenergetics was largely preserved. Additionally, increased aKG levels following OGDH inhibition impacted the biosynthesis of other critical amino acids. Thus, this work has identified a previously undescribed, functional link between certain TCA cycle components and nucleotide biosynthesis enzymes across AML. This metabolic node may serve as a cancer-specific vulnerability amenable to therapeutic targeting in AML and perhaps in other cancers with similar metabolic wiring.

overall_design: RNA sequencing of murine MLL-fusion AML cell lines was perfored at days 0 and 4 after induction of the indicated shRNAs with doxycycline. Two clonal isolates of two independent shRNAs targeting Ogdh (shOgdh346 and shOgdh2081) and a control shRNA (shRen713) were used.

first: Scott

last: Millman

first: Almudena

last: Chaves-Perez

first: Sudha

last: Janaki-Raman

first: John

middle: P

last: Morris IV

first: Varun

last: Narendra

first: Chi-Chao

last: Chen

first: Benjamin

middle: T

last: Jackson

first: Jossie

last: Yashinskie

first: Riccardo

last: Mezzadra

first: Tessa

last: Devine

first: Valentin J

middle: A

last: Barthet

first: Michelle

last: Saoi

first: Timour

last: Baslan

first: Sha

last: Tian

first: Zohar

last: Sachs

first: Lydia W

middle: S

last: Finley

first: Justin

middle: R

last: Cross

middle: W

last: Lowe


title: Paired CRISPR screens to map gene regulation in cis and trans (ATAC-seq)

first: Xinhe

last: Xue

summary: In this study, we develop a scalable approach that pairs discovery of cis- and trans-acting elements to systematically decipher the gene regulatory network. We explore the regulation of the immune checkpoint PD-L1 under both basal and IFNγ-stimulated contexts. PD-L1 expression on tumor cells serves as a clinically-actionable diagnostic for immune checkpoint blockade therapy across multiple cancer types.

overall_design:
Assay for transposase-accessible chromatin with sequencing  (ATAC-seq) for untreated and IFNγ-treated BxPC-3 human pancreatic adenocarcinoma cells
Assay for transposase-accessible chromatin with sequencing  (ATAC-seq) for BxPC-3 human pancreatic adenocarcinoma cells

first: Xiao

last: Wang

first: Zoran

last: Gajic

first: Christina

last: Caragine

first: Mateusz

last: Legut

first: Mari

last: Bell

first: Wen-Wei

last: Liang

first: Conor

last: Walker

first: James

last: Kim

first: Rachel

last: Yan

first: Gina

last: Hong

first: Hans-Hermann

last: Wessels

first: Congyi

last: Lu

first: Neil

last: Bapodra

first: Gamze

last: Gursoy

first: Neville

last: Sanjana


title: Paired CRISPR screens to map gene regulation in cis and trans (HiChIP)

overall_design: H3K27ac HiChIP for untreated and IFNγ-treated BxPC-3 human pancreatic adenocarcinoma cells


title: Paired CRISPR screens to map gene regulation in cis and trans (CUT&RUN)

overall_design:
Cleavage Under Targets & Release Using Nuclease (CUT&RUN) for SRF, BPTF, and H3K4me3 in BxPC3 human pancreatic adenocarcinoma cells
Cleavage Under Targets & Release Using Nuclease (CUT&RUN) for H3K27ac, H3K4me3 and IgG (control) in BxPC3 human pancreatic adenocarcinoma cells


title: Phenotypic plasticity and increased tissue infiltration of TREM1+ mono-macrophages following radiotherapy in rectal cancer. [scRNA-Seq]

status: Public on Nov 20 2024

city: wuhan

first: Mingjie

last: Li

address: union hospital

institute: Huazhong University of Science and Technology

summary: Our previously reported phase II and phase III trials have demonstrated that short-course radiotherapy (SCRT) combined with neoadjuvant immunochemotherapy (SIC) has led to clinical benefits in locally advanced rectal cancer (LARC). Characterization of the molecular mechanisms underlying responses to SIC may lead to improved treatment strategies. Here, we prospectively collected and applied multi-omic analyses to paired pre- and post-treatment LARC specimens undergoing SIC. The TREM1+ mono-macrophages subsets that display a pro-inflammatory phenotype are identified and correlate with complete response to SIC. Mechanically, ionizing radiation (IR) induces peripheral TREM1+ mono-macrophages expansion in tumors. Following IR, loss of TREM1 in mono-macrophages undermines antitumor immunity by altering mono-macrophages differentiation and inhibiting CD8+ T cell infiltration and activation. The TREM1+ mono-macrophages response may rely on activation of key inflammatory pathways, including NF-κB signaling and Toll-like receptor pathway. Pharmacological inhibition of TREM1 abolishes IR-induced immunoactivation and reduces combined IR and/or anti-PD-1 treatment. Thus, we establish a crucial role of a mono-macrophages state in mediating effective cancer therapy.

overall_design:
To investigate changes in peripheral blood mononuclear/macrophage system in LARC after neoadjuvant therapy with LC or SIC, we employed a large-scale transcriptomic analysis of peripheral blood mononuclear cells (PBMCs). Specimens were collected from 16 consecutive patients. To investigate the cellular and molecular contexture of the tumor immune microenvironment (TIME) following SIC treatment, we characterized CD45+ immune cells isolated from tumor tissue specimens before and after SIC by high-throughput single-cell RNA sequencing (scRNA-seq).

***************************************************************
no raw data due to patient privacy
***************************************************************


title: Analysis of transcriptomes of germinal center (GC) B cells derived from miR-155-sufficient and -deficient mice

status: Public on Nov 01 2024

city: London

first: Probir

last: Chakravarty

address: 1Midland Road

country: United Kingdom

institute: Francis Crick

summary: To investigate the impact caused by miR-155 ablation, positively selected light zone (LZ) GC B cells were sorted by flow cytometry and used to analyse transcriptomic profiles.

overall_design: Splenic B cells derived from miR-155-sufficient or -deficient mice expressing HEL-specific B cell receptor together with endogenous cMyc-GFP reporter cassette were adoptively transferred into congenic mice, followed by immunization with HEL3x mutant conjugated with sheep red blood cells (SRBCs). After five days, cMyc-GFP+ HEL+ LZ GC-B cells were sorted and used for the mRNA sequencing library.

first: Rinako

last: Nakagawa


title: Transcriptomic atlas of the morphologic development of the fungal pathogen Coccidioides reveals key phase-enriched transcripts [ChIP-Seq]

status: Public on Oct 21 2024

middle: M.

last: Homer

department: Biochemistry & Biophysics

summary: Coccidioides spp. are highly understudied but significant dimorphic fungal pathogens that can infect both immunocompetent and immunocompromised people. In the environment, they grow as multicellular filaments (hyphae) that produce vegetative spores called arthroconidia. Upon inhalation by mammals, arthroconidia undergo a process called spherulation. They enlarge and undergo numerous nuclear divisions to form a spherical structure, and then internally segment until the spherule is filled with multiple cells called endospores. Mature spherules rupture and release endospores, each of which can form another spherule, in a process thought to facilitate dissemination. Spherulation is unique to Coccidioides and its molecular determinants remain largely unknown. Here, we report the first high-density transcriptomic analyses of Coccidioides development, defining morphology-dependent transcripts and those whose expression is regulated by Ryp1, a major regulator required for spherulation and virulence. Of approximately 9000 predicted transcripts, we discovered 273 transcripts with consistent spherule-associated expression, 82 of which are RYP1-dependent, a set likely to be critical for Coccidioides virulence. ChIP-Seq revealed 2 distinct regulons of Ryp1, one shared between hyphae and spherules and the other unique to spherules. Spherulation regulation was elaborate, with the majority of 227 predicted transcription factors in Coccidioides displaying spherule-enriched expression. We identified provocative targets, including 20 transcripts whose expression is endospore-enriched and 14 putative secreted effectors whose expression is spherule-enriched, of which 6 are secreted proteases. To highlight the utility of these data, we selected a cluster of RYP1-regulated, arthroconidia-associated transcripts and found that they play a role in arthroconidia cell wall biology, demonstrating the power of this resource in illuminating Coccidioides biology and virulence.

overall_design: We generated arthroconidia and grew them as spherules (39˚C, 10% CO2) or hyphae (room temp, no additional CO2) in the specified media. At each timepoint, RNA was harvested and the transcriptome was analyzed by RNA-Seq. We have included 3 timecourses of WT spherules, 1 full and 1 limited timecourse of the ryp1∆ mutant which cannot form the spherule morphology but was subjected to spherulation or hyphal growth conditions. For samples labeled as originating from Figure S4, we also performed ChIP-Seq on the same biological samples using a polyclonal anti-Ryp1 antibody.

first: Mark

last: Voorhies

first: Keith

last: Walcott

first: Elena

last: Ochoa

first: Anita

last: Sil


title: Transcriptomic atlas of the morphologic development of the fungal pathogen Coccidioides reveals key phase-enriched transcripts [RNA-seq]

first: Christina


title: IRE1a silences double stranded RNA through RIDD

status: Public on Oct 16 2024

city: Houston

first: FEI

last: YUAN

state: TX

address: 1 Baylor Plaza

department: Pathology & Immunology

institute: Baylor College of Medicine

summary: Chemotherapy is often combined with immune checkpoint inhibitor (ICI) to enhance immunotherapy responses. Despite the approval of chemo-immunotherapy in multiple human cancers including triple-negative breast cancer (TNBC), immunologically cold tumors remain largely unresponsive due to weak chemotherapy-stimulated immune responses. The mechanisms determining the immunogenicity of chemotherapy in immunologically cold tumors remain largely unknown. Here, we identify the endoplasmic reticulum (ER) stress sensor IRE1α as a critical checkpoint that restricts the immunogenicity of taxane chemotherapy and prevents the innate immune recognition of immunologically cold TNBC. IRE1α RNase silences chemotherapy-induced double-stranded RNA (dsRNA) through RIDD (Regulated IRE1-Dependent Decay) to prevent NLRP3 inflammasome–dependent pyroptosis. Inhibition of IRE1α RNase activity with a selective RNase inhibitor ORIN1001, currently in Phase I clinical trial, allows taxane chemotherapy to induce extensive dsRNA accumulation and activate NLRP3 inflammasome–dependent gasdermin D (GSDMD) cleavage. This triggers pyroptosis and a highly effective immune response in immunologically cold TNBC.

overall_design: J2-RIP and paired INPUT controls were performed in biological triplicates.

first: Longyong

last: Xu

first: Fei

last: Yuan

first: Xi

last: Chen


title: Genome-wide in vivo dynamics of cohesin-mediated loop extrusion and its role in transcription activation

status: Public on Oct 07 2024

first: Jesse

last: Dixon

address: 10010 N. Torrey Pines Rd.

institute: Salk Institute for Biological Studies


title: Ursolic acid-induced DECR1 degradation via NF-κB/NLRP3 pathway inihibits vascular calcification

status: Public on Sep 13 2024

city: Guangzhou

first: Hongwei

last: Yang

address: Nanshitou Street

institute: Southern Medical University

summary: In this study, we repoort the protective effect of ursolic acid (UA) on vascular calcification in chronic kidney disease. To elucidate the molecular mechanism underlying the anti-vascular calcification effect of UA, we performed RNA-seq to identify the gene expresion under UA treatmment.

overall_design: Human vasculr smooth muscle cells were cultured in calcifying medium in the presence or absence of UA for 48 hours. RNA-seq was performed to determine the downstream target of UA. The gene expression level was calculated using the Fragment Per Kilobase of Million mapped reads (FPKM) methods. Differential expression  was assessed using the DEGseq packages.

first: Jianyun

last: Yan


title: Single nucleus multiome (RNAseq and ATACseq) of female mouse pituitary

status: Public on Oct 01 2024

last: Zaslavsky

address: 1 Gustave L. Levy Pl

institute: Icahn School of Medicine at Mount Sinai

summary: We report the single nucleus multiome (RNAseq+ATACseq) of a female mouse pituitary sample. This dataset was generated for supporting the development of a data-driven batch inference method and transforms often heterogeneous data matrices obtained from different samples into a uniformly cell-type annotated and integrated dataset.

overall_design: Multiomic (RNAseq+ATACseq) sequencing of pituitary tissue from 14 female samples.

first: Frederique

last: Ruf-Zamojski


title: Deep mutational scanning and mobility-based selection for structured RNAs

status: Public on Sep 10 2024

city: Shenzhen

first: Zhe

last: Zhang

state: Guangdong

address: Gaoke Innovation Center

department: Institute of Physical and Systems Biology

institute: Shenzhen Bay Laboratory

summary: Deciphering RNA three-dimensional structures remains a central challenge in molecular biology. Deep mutational profiling has captured evolutionary constraints. However, its current application has been limited to a few RNA functions that are compatible with high-throughput selection. Here, we introduce RNA-MobiSeq, where deep-mutational variants with similar structures to the wild type were enriched by mobility-selected native gel bands and analyzed for covariation by an unsupervised, RNA-specific method and thermodynamic modeling. This de novo folding platform enabled accurate prediction of base-pairing (F1-score ≥ 90%) and improved performance on tertiary structure prediction over existing template-free modeling techniques for nine RNAs in diverse structures and functions. Validated for robustness by replicate experiments, the integrated platform provides a probe-free and broadly applicable framework for elucidating structural landscapes for RNAs that can be synthesized and assayed under in vitro native gel electrophoresis conditions, thereby bridging sequence information and functional understanding.

overall_design: The RNA-MobiSeq pipeline is a method to predict the 3D structure of a target RNA from its sequence by obtaining artificial homologous sequences and their native-mobility-fitness scores from high-throughput sequencing of mobility-selected RNA mutants, followed by a new RNA-specific covariation analysis as restraints for energy-based structure prediction. Error-prone PCRs (or oligo pools) were first employed to obtain randomly mutated sequences, which were assembled into plasmids and then electro-transformed into DH5α for library constructions. The DNA libraries were further in vitro transcribed into RNAs that were subjected to native PAGE separation according to mobility. The band with the same mobility as the wild-type RNA sequence was excised for reverse transcription. Both pre- and post-selection samples were sent for high-throughput sequencing. The obtained sequences along with their fitness scores estimated from mutants’ fractions in the excised band were then analyzed for co-variation to infer the base pairing structures by an RNA-specific method CODA2 and MC simulated annealing. The resulting base pairing structures were employed as restraints for energy minimization to model the 3D structures of the RNA by using the BRiQ statistical energy function and a folding-tree-based folding algorithm.

first: Jinle

last: Tang

first: Yazhou

last: Shi

first: Jian

last: Zhan

first: Yaoqi

last: Zhou


title: Most adult gonadotrophs originate from postnatal pituitary stem cells during minipuberty.

first: Probir

last: Chakravarty

summary: In the first three weeks postnatally, the pituitary gland undergoes a period of rapid growth, due to both cell proliferation and endocrine cell size increase. SCs are initially the most proliferative, and lineage tracing experiments previously showed that neonatal SCs can give rise to all endocrine cell types. However, because the Cre drivers used for lineage tracing were relatively inefficient and relied on tamoxifen, a selective estrogen receptor modulator which perturbs normal physiology, we could not decipher SC contribution to organ growth. We have thus performed lineage tracing using a new, more efficient and physiologically neutral Sox2rtTA allele. When we traced early postnatal SCs using Sox2rtTA, we observed, similarly in both sexes, that most adult gonadotrophs derive from this neonatal SC population. These observations were confirmed and complemented by a single cell RNAseq dataset from postnatal day 3 (PND3) SOX9iresGFP XX and XY positive cells. From SOX9iresGFP cells, which comprise SCs and their immediate progeny, cell trajectories were inferred, and relevant gene regulatory networks predicted. Along with lineage tracing experiments, this dataset confirmed that SCs mostly differentiate into gonadotrophs postnatally. As animals develop, SC-derived gonadotrophs invade the gland, while the minor embryonic population remains confined ventrally. The discovery of a dual origin for gonadotrophs may help understand aspects of gonadotrophin regulation and mechanisms of diseases affecting puberty and fertility.

overall_design: FACsorted Sox9iresGFP positive cells from post-natal day 3 (PND3) males and females.

first: Daniel

last: Sheridan

first: Karine

last: Rizzoti


title: Type 2 Diabetes Specific Spatial Differential Gene Expression in Pancreas Visium Data

status: Public on Aug 31 2024

city: Bethesda

first: Michael

last: Erdos

address: 50 South Dr

institute: NIH

summary: Understanding the spatial distribution of gene expression in the pancreas could provide new insights into its function and islet biology. Visium spatial gene expression is a powerful tool for quantifying gene expression of cells within their spatial context. We describe the generation and analysis of spatial transcriptomics data from six pancreas samples from healthy donors. Our analysis revealed high expression of INS within islets. However, we also identified a bleed of highly abundant islet genes (e.g., INS, GCG) into surrounding tissue areas. Through computational analyses, we successfully corrected for this bleed. We then identify genes which are spatially differentially expressed in the Type 2 Diabetes samples.  We provide these data for researchers to study the organization of the pancreas and islets.

overall_design:
human pancreatic tissue flash frozen with isopentane; tissue sections prepared and processed with the 10X Genomics Visium Spatial Gene Expression Reagent kit
>>>Raw data were deposited in dbGaP under phs001188.v3.p1 due to patient privacy concerns.<<<<

first: Lori

middle: L

last: Bonnycastle

first: Mike

first: Caleb

last: Grenko

first: Zoe

last: Weiss

first: Cassie

last: Robertson

first: Narisu

last: Narisu

first: Leland

last: Taylor

first: Francis

last: Collins


title: IL-9 secreted by leukemia stem cells induces Th1-skewed CD4+ T cells, which promote their expansion

city: Bern

first: Ramin

last: Radpour

address: Murtenstrasse 35

department: Medical Oncology

country: Switzerland

institute: Bern University Hospital

summary: We performed a comprehensive transcriptomic profiling of BM-infiltrating CD4+ T cells of AML patients from different AML patients and controls. This analysis revealed that BM-infiltrating CD4+ T cells are activated and skewed towards Th1 polarization.

overall_design: We characterized the molecular signature of BM-infiltrating CD4+ T cells in patients with acute myeloid leukemia (AML) compared to control subjects.

first: Adrian

middle: F

last: Ochsenbein


title: Hexokinase 3 enhances myeloid cell survival via non-glycolytic functions

summary:
Purpose: The goal was to compare transcriptome profiles of HL60 HK3-null and Cas9 expressing control cells treated with either ATRA or DMSO using RNA sequencing. In addition, to find accessible DNA regions by probing open chromatin with hyperactive mutant Tn5 Transposase using ATAC sequencing.
Methods: HL60 HK3-null and Cas9 expressing control cells were treated with 1µM of ATRA or DMSO (0.01%) for 36h. Treated cells were then harvested for RNA-Seq and ATAC-Seq analysis.
Results: We mapped sequence reads per sample to the human genome (GRCh38 - hg38) and identified differentially expressed transcripts as well as open chromatin peaks in AML HL60 HK3-null and Cas9 expressing cells upon treatment with ATRA or DMSO as control. Differentially expressed genes, were identified with a fold change ≥1.5 and FDR p-value <0.05.
Conclusions: Our results show that HK3 knockout (HK3-null) AML cells had an accumulation of reactive oxygen species (ROS) as well as DNA damage during ATRA-induced differentiation. RNA sequencing analysis confirmed pathway enrichment for programmed cell death, oxidative stress and DNA damage response in HK3-null AML cells. These signatures were confirmed in ATAC sequencing, showing that loss of HK3 leads to changes in chromatin configuration and increases accessibility of genes involved in apoptosis and stress response.

overall_design: HL60 HK3-null and Cas9 expressing control cells were treated with ATRA or DMSO. Next, RNA-Seq and ATAC-Seq analyses was perfomred and differences between control and HK3-null as well as ATRA and control treated HL60 cells were assessed.

first: Kristina

last: Seiler

first: Mario

last: Tschan

first: Bruce

last: Torbett


title: Chinmo maintains adult stem cell sex identity by directly regulating Doublesex and Insulin signaling

status: Public on Aug 30 2024

first: Qing

last: Ma

address: 1068 Xueyuan Avenue, Shenzhen University Town, Shenzhen, P.R.China

institute: Shenzhen Institutes of Advanced Technology


title: MYC and p53 cooperate through VEGF signaling to repress cytotoxic T cell and immunotherapy responses in prostate cancer

status: Public on Aug 26 2024

first: Yu-Jui

last: Ho

summary: To study the genetic factors that influence the immune landscape of castration-resistant prostate cancer (CRPC), we utilized a flexible, electroporation-based system to introduce genetic alterations relevant to human disease directly into the prostate glands of mice. These electroporation-based genetically engineered mouse models (EPO-GEMM) recapitulate features of traditional models, and allow us to investigate distinct genetic subtypes of prostate cancer within an intact tumor-immune microenvironment. We observed differences between genetic subtypes of CRPC, with MYC-driven subtypes exhibiting a "cold" immune landscape compared to others. Interestingly, we found that the compound loss of different tumor suppressors such as Pten or p53 with MYC further impacts the inflammatory profile of prostate cancer, with MYC and p53  (MP) alterations cooperating to drive VEGF expression and immune suppression. VEGF signaling blockade resulted in re-activation of cytotoxic T cell anti-tumor immunity and restored sensitivity to immunotherapy in MP CRPC. Thus, by leveraging the power of EPO-GEMMs to generate distinct subtypes of CRPC, our studies reveal a functional role for VEGF signaling in driving prostate cancer immune evasion and validate the VEGF pathway as an actionable therapeutic target in MYC and p53 altered prostate cancer.

overall_design: For RNA-seq analysis of the transcriptional profiles of PtPRb EPO-GEMM prostate tumors, total RNA was extracted from bulk tissue using the RNeasy Mini Kit (Qiagen). Purified polyA mRNA was subsequently fragmented, and first and second strand cDNA synthesis performed using standard Illumina mRNA TruSeq library preparation protocols. Double stranded cDNA was subsequently processed for TruSeq dual-index Illumina library generation.

first: Katherine

last: Murphy

first: Kelly

middle: D

last: DeMarco

first: Lin

last: Zhou

first: Yvette

last: Lopez

first: Yu-jui

first: Junhui

last: Li

first: Shi

last: Bai

first: Karl

last: Simin

first: Lihua

last: Zhu

first: Arthur

last: Mercurio

first: Marcus

last: Ruscetti


title: Effect of PLX4720 on melanoma-associated fibroblasts co-cultured with melanoma cells

status: Public on Aug 09 2024

first: Probir

last: Chakravarty

summary: The experiment was designed to assess the effect of PLX4720 on the gene expression in human primary melanoma-associated fibroblast co-cultired with 5555 mouse melanoma cells

overall_design:
Total RNA from DMSO or PLX4720-treated melanoma-associated fibroblast (MAF2-mCherry) co-cultured with melanoma cells (5555-mEGFP), FACS sorted. 4 samples previoulsy uploaded under GSE63160
idat files not availabe for this submission.

first: E

last: Hirata

first: G

last: Giangreco

last: Sahai


title: Effect of PLX4720 on melanoma cells co-cultured with melanoma-associated fibroblasts

status: Public on Jul 29 2024

summary: The experiment was designed to assess the effect of PLX4720 on the gene expression in 5555 mouse melanona cells co-cultured with human primary melanoma-associated fibroblast

overall_design: Total RNA from DMSO or PLX4720-treated melanoma cells (5555-mEGFP) co-cultured with melanoma-associated fibroblast (MAF2-mCherry), FACS sorted

first: Erik

first: Eishu


title: Harmonized cross-species cell atlases of trigeminal and dorsal root ganglia

city: Boston

first: Shamsuddin

last: Bhuiyan

state: MA

address: 60 Fenwood Road

institute: Brigham and Women's Hospital

summary: Peripheral sensory neurons in the dorsal root ganglion (DRG) and trigeminal ganglion (TG) are specialized to detect and transduce diverse environmental stimuli including touch, temperature, and pain to the central nervous system. Recent advances in single-cell RNA-sequencing (scRNA-seq) have provided new insights into the diversity of sensory ganglia cell types in rodents, non-human primates, and humans, but it remains difficult to compare transcriptomically defined cell types across studies and species. Here, we built cross-species harmonized atlases of DRG and TG cell types that describe 18 neuronal and 11 non-neuronal cell types across 6 species and 31 studies. We then demonstrate the utility of this harmonized reference atlas by using it to annotate newly profiled DRG nuclei/cells from both human and the highly regenerative axolotl. We observe that the transcriptomic profiles of sensory neuron subtypes are broadly similar across vertebrates, but the expression of functionally important neuropeptides and channels can vary notably. The new resources and data presented here can guide future studies in comparative transcriptomics, simplify cell type nomenclature differences across studies, and help prioritize targets for future analgesic development.

overall_design: Beta-III tubulin:GAP43-EGFP transgenic axolotls (125) 7-8cm in length (snout to tail) were anesthetized in 0.1% tricaine. For DRG dissociation at single-cell level, 10X Genomics “Dissociation of Mouse Embryonic Neural Tissue for Single Cell RNA Sequencing” protocol was adapted to axolotl tissue. DRGs of brachial plexus nerves located at C3, C4 and C5 were collected from both left and right sides and combined (total of 6 DRGs) in a single tube containing chilled 0.7X HBSS buffer and kept on ice. For dissociation, 400ul papain (Worthington Biochemical, Cat: #LK003178) dissolved in 0.7X PBS was pre-warmed in 37˚C water bath for 10 minutes to activate the enzyme. HBSS was removed and activated papain was added onto DRGs. The DRG in papain was placed in 37˚C water bath for 20 minutes. After the incubation, DRGs were settled in the bottom of the tube by a brief spin at 200g for 1min and papain was removed. 500ul DRG media consisting of Neurobasal Plus Medium at 60% (v/v), (Thermo Scientific, cat: A3582901), 50X B-27 Plus Supplement stock at 1X (Thermo Scientific, cat: A3582801), 100X insulin-transferrin-selenium stock at 1% (v/v), (Peprotech, cat: 41400045), 250μg/mL amphotericin B stock at 1% (v/v), (Sigma-Aldrich, cat: A2942), gentamicin at 50mg/ml (Sigma-Aldrich, cat:  G1264), 50ug/ml recombinant human β-NGF stock at 1:1000, (Peprotech, cat: 450-01) mixed in Ringer’s solution (115mM NaCl, 2.5mM KCl, 2mM CaCl2, 10mM HEPES pH7.4, 0.5 mM EDTA dissolved in water and ph adjusted to 7.4 with NaOH) was added. DRGs were triturated in media using 1000 μl pipette. Trituration was performed slowly and gently until most of the tissue was dissociated. After dissociation, sample was centrifuged at 200g for 3min. The supernatant was removed, and the pellet was resuspended in 100 μl of0.7XPBS+%0.04 BSA. Cell count and single-cell sequencing were performed by Harvard University Bauer Sequencing core. Luna-FL Dual Fluorescence Cell Counter was used to assess number of viable cells. Cell count was repeated 4 times and averaged to determine the final cell count, viability, and cell size. Total of 405 cells/μl with 100% cell viability were present in the final cell suspension in first sample and 486 cells/μl with 97% viability in the second set. scRNA-seq libraries were prepared with the Chromium™ Single Cell 3′ NextGEM, Library and Gel Bead Kit v3.1.

first: Mengyi

last: Xu

first: Parth

last: Bhatia

first: William

last: Renthal


title: Metabolic enzymes moonlight as selective autophagy receptors to protect plants against viral induced cellular damage

status: Public on Jul 23 2024

city: Vienna

first: Yasin

last: Dagdas

address: Dr. Bohr-Gasse 3

country: Austria

institute: Gregor Mendel Institute

summary: RNA viruses co-opt host endomembranes to form replication complexes, often triggering cellular stress and immune responses. Here, we show that Arabidopsis thaliana activates selective autophagy to respond to viruses targeting mitochondria, chloroplasts, and the endoplasmic reticulum. Rather than degrading viral components, autophagy selectively removes the immune regulator Enhanced Disease Susceptibility 1 (EDS1) to prevent cell death. This targeted mechanism is mediated by oligomeric metabolic enzymes that moonlight as selective autophagy receptors, linking organelle stress to immune homeostasis. Our findings establish selective autophagy as an essential immune rheostat that fine-tunes defense responses and safeguards cellular integrity to promote host survival during viral infections.

overall_design: For RNAseq analysis, the same samples as the ones used for TMT proteomics (PRIDE ID #PXD051596) were used to generate 3’TAG seq libraries

first: Marion

last: Clavel


title: snRNA-seq dataset for "Reelin marks cocaine-activated striatal ensembles, promotes neuronal excitability, and regulates cocaine reward"

status: Public on Jul 10 2024

city: Birmingham

first: Jeremy

last: Day

state: Alabama

address: 1825 University Blvd SHEL 910

department: Neurobiology

institute: University of Alabama at Birmingham

summary: This dataset contains single-nucleus RNA-seq results from rat adult nucleus accumbens tissue treated with CRISPRi machinery (hSyn-driven dCas9-KRAB-MeCP2 fusion) with a CRISPR sgRNA targeting lacZ (a bacterial gene not found in the mammalian genome; used as a non-targeting control) or Reln.

overall_design: This experiment contains 8 biological samples, which represent individual rats (4 male, 4 female). Rats underwent stereotaxic surgery to transduce nucleus accumbens tissue with lentiviruses expressing CRISPRi machinery targeting lacZ (lacZ control; sample names beginning with lacZ), or Reln (sample names beginning with Reln). Two weeks following surgery, animals (4 lacZ, 4 Reln 2M/2F per group) were euthanized prior to rapid decapitation. Brains were blocked in 1mm sections, and NAc punches were obtained, flash frozen, and stored at -80°C prior to nuclei isolation. Following FACS-assisted nuclei isolation and quantification, nuclei from one rat were loaded into one GEM (gel beads in emulsion) well to generate barcoded nuclei (10× Genomics, PN-1000123, PN-1000122) on a Chromium iX instrument. Libraries were prepared according to the 10X Genomics manufacturer's recommendations, and sequenced on a NovaSeq 6000 platform to a depth of 300,000 reads per nuclei. Forward and reverse paired-end reads are denoted as "R1" or "R2" in raw file names.

first: Kasey

last: Brida

first: Catherine

last: Newman

first: Lara

last: Ianov

middle: J


title: Metabolic adaptations direct cell fate during tissue regeneration

status: Public on Apr 02 2025

city: New York

first: Yu-Jui

last: Ho

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

overall_design: Refer to individual Series


title: RNA-seq dataset for "Reelin marks cocaine-activated striatal ensembles, promotes neuronal excitability, and regulates cocaine reward"

summary: This dataset contains RNA-seq results from rat embryonic striatal neuronal cultures treated with CRISPRi machinery (dCas9-KRAB-MeCP2 fusion) with a CRISPR sgRNA targeting lacZ (a bacterial gene not found in the mammalian genome; used as a non-targeting control) or Reln.

overall_design: This experiment contains 8 biological samples, each of which underwent RNA-seq on an Illumina NovaSeq6000 instrument. Samples were treated with CRISPRi machinery targeting lacZ (lacZ control; samples X1i, X2i, X3i, X4i), or Reln (samples X5i, X6i, X7i, X8i) . Forward and reverse paired-end reads are denoted as "R1" or "R2" in raw file names.


title: Single-cell RNA-Sequencing of Wild-type Perinatal Mice Thymi

status: Public on Mar 19 2025

city: Nashville

first: Qi

last: Liu

state: Tennessee

address: 2525 WEST END AVE

institute: Vanderbilt University Medical Center

summary: We prepared enzymatically digested thymus single cell suspensions for scRNA-Seq. Cells were extracted from perinatal mice, and TEC cells were enriched via FACS using anti-EpCAM and Anti-CD45 to isolate EpCAM+ CD45- TECs.

overall_design: FACS purified TECs were obtained from P0, P3, P7, P10, P15 and P28 C57Bl6/J male and female thymuses. Supplicate experiments were performed for each time point except for P15.

first: Ellen

last: Richie

first: Anusha

last: Vasudev

first: Nandini

last: Singarapu

first: Ken

last: Lau

first: Bryan

last: Helm


title: TULIPs decorate the three-dimensional genome of PFA ependymoma

status: Public on Jul 05 2024

first: Marco

last: Gallo

address: 1102 Bates Avenue

department: Pediatrics

summary: Hi-C and RNA-seq for a large cohort of pediatric brain tumors including ependymoma (PFA, PFB, Ste, spinal), medulloblastoma (G3, G4, SHH), high grade glioma (H3K27 and H3-WT), pilocytic astrocytoma, and more.

overall_design:
Assessing 3D genome conformation by Hi-C for many types of pediatric brain tumors.

>>>Submitter states that raw data have been deposited at EGA under accession EGAS00001005476<<<

first: Michael

last: Taylor

first: Nada

last: Jabado


title: CITCO is a non-genotoxic hepatocarcinogen in humanised constitutive androstane receptor mice but not in wild-type animals

status: Public on Feb 17 2025

first: Probir

last: Chakravarty

summary: A large number of drugs and compounds produced by the chemical and agrochemical industry score as non-genotoxic carcinogens (NGC) in rodent models. It is unclear whether these compounds are also carcinogens for humans. The most extensively investigated compounds have been the anti-convulsive drugs, phenobarbital (PB) and phenytoin. Liver tumours induced by PB are dependent upon the activation of the constitutive androstane receptor (CAR).  However, marked species differences in CAR activation by exogenous chemicals exist with some being much more potent activators of human CAR e.g. 6-(4-chlorophenyl)imidazo[2,1-b][1,3]thiazole-5-carbaldehyde-O-(3,4-dichlorobenzyl)oxime (CITCO). We have compared CITCO-induced tumour formation in the livers of mice in which human CAR has been replaced its murine counterpart. Our findings reveal that CITCO-dependent liver tumours are only formed in mice expressing human CAR and not in wild-type animals. In addition, contrary to one of the proposed mechanisms of NGC carcinogenicity, we show that CITCO did not induce a hyperplastic response in the liver of the humanised mice. These data raise some key questions about the mechanism of action of NGCs and identifies limitations of current rodent carcinogenicity test systems in relation to risk assessment

overall_design: Adult male wild-type (WT) mice,  mice humanised for CAR and PXR (hCAR/hPXR) or mice nulled for Car and PXR and humanised for PXR (Car KO/hPXR), n=3

first: C

last: Henderson


title: Transcriptomic profiling of Schlemm’s canal cells reveals a lymphatic-biased identity and three major cell states

status: Public on Jul 04 2024

city: Baltimore

first: Taibo

last: Li

state: MD

address: 1830 E Monument Street, Suite 2-300

institute: Johns Hopkins School of Medicine

summary: Schlemm’s canal (SC) is central in intraocular pressure regulation but requires much characterization. It has distinct inner and outer walls, each composed of Schlemm’s canal endothelial cells (SECs) with different morphologies and functions. Recent transcriptomic studies of the anterior segment added important knowledge, but were limited in power by SEC numbers or did not focus on SC. To gain a more comprehensive understanding of SC biology, we performed bulk RNA sequencing on C57BL/6J SC, blood vessel and lymphatic endothelial cells from limbal tissue (∼4500 SECs). We also analyzed mouse limbal tissues by single-cell and single-nucleus RNA sequencing (C57BL/6J and 129/Sj strains), successfully sequencing 903 individual SECs. Together, these datasets confirm that SC has molecular characteristics of both blood and lymphatic endothelia with a lymphatic phenotype predominating. SECs are enriched in pathways that regulate cell-cell junction formation pointing to the importance of junctions in determining SC fluid permeability. Importantly, and for the first time, our analyses characterize 3 molecular classes of SECs, molecularly distinguishing inner wall from outer wall SECs and discovering two inner wall cell states that likely result from local environmental differences. Further, and based on ligand and receptor expression patterns, we document key interactions between SECs and cells of the adjacent trabecular meshwork (TM) drainage tissue. Also, we present cell type expression for a collection of human glaucoma genes. These data provide a new molecular foundation that will enable the functional dissection of key homeostatic processes mediated by SECs as well as the development of new glaucoma therapeutics.

overall_design: The anterior segment of the eye including the limbus was dissected from 3-month-old Prox1-GFP mice.  A strip of limbal tissue 1-2 mm wide consisting of the aqueous humor outflow tissues SC and TM, ciliary body, iris, and cornea was dissected from each anterior segment in ice-cold Dulbecco’s modified Eagle’s medium DMEM. To enrich for aqueous humor outflow tissues, limbal strips from 6 C57Bl/6J eyes (4 females, 2 males) were pooled for a single run of C57BL/6J scRNA-seq. C57BL/6J scRNA-seq was performed on two separate pools of eyes for a total of 12 eyes (8 females, 4 males). For 129/Sj scRNA-seq, 6 eyes (3 females, 3 males) were pooled for a single run.

first: Revathi

last: Balasubramanian

first: Krishnakumar

last: Kizhatil

first: Jiang

last: Qian

first: Simon

last: John


title: Keratinocytes Drive the Epithelial Hyperplasia Key to Sea Lice Resistance in Coho Salmon

status: Public on Jun 24 2024

city: Midlothian

first: Sarah

last: Salisbury

address: Easter Bush Campus

institute: The Roslin Institute, University of Edinburgh

summary: Background:  Salmonid species have followed markedly divergent evolutionary trajectories in their interactions with sea lice. While sea lice parasitism poses significant economic, environmental, and animal welfare challenges for Atlantic salmon (Salmo salar) aquaculture, coho salmon (Oncorhynchus kisutch) exhibit near-complete resistance to sea lice, achieved through a potent epithelial hyperplasia response leading to rapid louse detachment. The molecular mechanisms underlying these divergent responses to sea lice are unknown.  Results: We characterised the cellular and molecular responses of Atlantic salmon and coho salmon to sea lice using single-nuclei RNA sequencing. Juvenile fish were exposed to copepodid sea lice (Lepeophtheirus salmonis), and lice-attached pelvic fin and skin samples were collected 12h, 24h, 36h, 48h, and 60h after exposure, along with control samples. Comparative analysis of control and treatment samples revealed an immune and wound-healing response that was common to both species, but attenuated in Atlantic salmon, potentially reflecting greater sea louse immunomodulation. Our results revealed unique but complementary roles of three layers of keratinocytes in the epithelial hyperplasia response leading to rapid sea lice rejection in coho salmon. Our results suggest that basal keratinocytes direct the expansion and mobility of intermediate and, especially, superficial keratinocytes, which eventually encapsulate the parasite.  Conclusions: Our results highlight the key role of keratinocytes in coho salmon’s sea lice resistance, and the diverged biological response of the two salmonid host species when interacting with this parasite. This study has identified key pathways and candidate genes that could be manipulated using various biotechnological solutions to improve Atlantic salmon sea lice resistance.

sample_organism:
Oncorhynchus kisutch
Salmo salar

platform_organism:
Oncorhynchus kisutch
Salmo salar

overall_design: Control and sea lice (Lepeophtheirus salmonis)-attached fin and skin tissue samples of Atlantic salmon (Salmo salar) and coho salmon (Oncorhynchus kisutch) were analyzed using snRNAseq.

first: Rose

last: Daniels

first: Sean

last: Monaghan

first: James

last: Bron

first: Paula

last: Villamayor

first: Ophelie

last: Gervais

first: Mark

last: Fast

first: Lene

last: Sveen

first: Ross

last: Houston

first: Nicholas

last: Robinson

first: Diego

last: Robledo


title: Mesenchymal stromal cell reprogramming and aberrant inflammatory signalling in myelofibrosis: insights from single-cell RNA sequencing

status: Public on Jun 01 2024

city: Manchester

first: Naseer

middle: Jr

last: Basma

address: Cancer Research UK Manchester Institute, Wilmslow Road

institute: Cancer Research UK Manchester Institute

summary: Myelofibrosis (MF) is a myeloproliferative neoplasm driven by mutations that impact the JAK/STAT signaling in blood cells, eventually causing fibrosis, splenomegaly, and reduced lifespan. To better understand MF, we performed scRNAseq profiling of collagenase-digested bone marrow biopsy samples to identify disrupted cellular pathways for potential treatment. Comparative analysis of 9 MF samples vs. 7 normal controls vs. 4 MF patients after allogeneic transplant showed striking differences in cell type composition and transcriptional profiles. Of note, Compositional comparisons identified the non-hematopoietic stromal cell fraction as harboring the biggest differences with, surprisingly, a proportional relative depletion of mesenchymal stromal cells (MSCs) and enrichment of endothelial cells (ECs) in MF vs. normal controls. Concomitant transcriptional differences were observed between MSCs recovered from MF vs. normal control sample, characterised by reduced expression of hematopoietic support factor genes (eg, KITLG, CXCL12, ANGPTL2, NCAM2, CSF1) and increased expression of extracellular matrix (ECM) genes (eg, FN1, COL1A2, COL3A1, DCN, SPARC), features which suggest an explanation for the increased frequency of graft failure following allograft in MF versus other hematologic malignancies.  Single-Cell rEgulatory Network Inference and Clustering (SCENIC) indicated transcription factors with either increased (eg, NR2F2, RUNX2, FOXP2) or decreased (eg, CEBPA, FOXC1, and PPARG) transcriptional activity as candidate master transcriptional regulators of the observed MSC reprogramming in MF.  Within the hematopoietic fraction, and agnostic of JAK inhibitor treatment status, comparison of MF vs. control cells unexpectedly revealed significant upregulation of interferon response genes, especially in CD14+ monocytes, but also in neutrophils and their progenitors, centred on expression of interferon-stimulated gene factor 3 (ISGF3) genes IRF9 and STAT1/2. Comparison of MF samples according to ruxolitinib treatment status indicated that ruxolitinib induced downregulation of an NFκB-centered inflammatory program, particularly in neutrophils.  Finally, ligand-receptor analysis using NicheNet identified TGFB1, IL1B, and TNF as top predicted ligands for induction of the MF signature in MSCs. While myeloid cells were top producers of these factors, their expression did not vary between MF vs. controls, suggesting other mechanisms, such as prolonged retention of ligand in the ECM, are important.  In summary, our comparative scRNA analysis of BM cells from MF patients and controls – the largest reported to date – reveals (i) transcriptional reprogramming of MSCs leading to reduced expression of hematopoietic support factor and increased expression of ECM genes; and (ii) increased expression of interferon and NFkB inflammatory pathways in myeloid cells.

overall_design: Bone marrow cells from digested human trephine biopsy tissue were subjected to Fluorescence-activated cell sorting (FACS) to isolate the haematpoeitic and microenvironmental fraction of cells, then analyzed using scRNAseq. Comparative analysis was then done to compare cells from myelofibrosis patients and healthy controls.

first: Alexia

last: Strickson

first: Kristian

last: Gurashi

first: Maria

last: Kleppe

first: John

last: Weightman

first: Hugh

middle: Y

last: Rienhoff Jr

first: Joanna

last: Storer

first: Tim

last: Somervaille


title: Esophageal adenocarcinoma

status: Public on May 25 2024

city: Amsterdam

first: Richard

last: Volckmann

state: Noord-Holland

address: van der Boechorststraat 6a

department: Laboratory of Experimental Oncology and Radiobiology

country: Netherlands

institute: msterdam UMC and University of Amsterdam


title: Multimodal single-cell profiling reveals the enhanced antitumor activity of FGFR4/CD276 BiCisCAR T cells in rhabdomyosarcoma

status: Public on May 23 2024

first: Javed

last: Khan

address: 37 Convent Dr.

department: Genetics Branch

institute: NCI, NIH

summary: We have generated a clinical grade CAR T targeting FGFR4 with a CD8 hinge and transmembrane domain (HTM) and a 4-1BB co-stimulatory domain (CSD) that is currently being developed for a clinical trial at the NCI. This standard FGFR4-CAR demonstrated potency in low burden RMS disease but has a more modest effect for bulky disease in RMS orthotopic model. Thus, we developed optimized FGFR4-CARs by modification of HTM or CSD to improve its efficacy. However, these modified FGFR4-CARs, even FGFR4.28HTM.28z CAR were unable to eradicate solid tumors in an more aggressive RMS559 model. CAR T cells targeting another cell surface protein CD276, a direct target of the RMS core-regulatory transcription factor MYOD1 is potent and effective, also could not eradicate the tumor. Therefore, we engineered a bicistronic CAR (BiCisCAR) to target both FGFR4 and CD276, with CD28 and 4-1BB CSDs, respectively, which showed remarkable and rapid tumor eradication compared with either single CAR alone. Here, we utilized a multimodal approach including simultaneous protein and transcriptome measurement at single cell level to characterize each FGFR4 or CD276 CAR T-cell infiltrating in RMS orthotopic xenografts. This assay further determined the molecular basis of superior antitumor effect of FGFR4.28HTM.28z-CD276.8HTM.BBz BiCisCAR T cells compared with other CARs.

overall_design: To explore the molecular basis of superior antitumor effect of FGFR4.28HTM.28z-CD276.8HTM.BBz BiCisCAR T cells compared with other FGFR4 or CD276 CARs infiltrating in RMS orthotopic tumor, we designed a multimodal single cell profiling approach(surface protein staining (Biolegend TotalSeq-C  panel) and mRNA) using 10x Genomics Chromium system. This assay simultaneously identify 14 protein markers (plus 3 isotype control anitibodies) and transcriptome measurement at the single cell level (CITE-seq).

first: Meijie

last: Tian

first: Xinyu

last: Wen

first: Jun

last: Wei


title: irCLIP-RNP reveals patterns of dynamic protein associations on RNA [CLIP-seq]

status: Public on Jan 28 2025

city: Stanford

first: Luca

last: Ducoli

address: 269 Campus Drive CCSR 2150

institute: Stanford University

summary: RNA binding proteins (RBPs) control varied processes, including RNA splicing, stability, transport, and translation. Dysfunctional RNA-RBP interactions contribute to the pathogenesis of human disease, however, characterizing the nature and dynamics of multiprotein assemblies on RNA has been challenging. To address this, non-isotopic ligation-based ultraviolet crosslinking immunoprecipitation was combined with mass spectrometry (irCLIP-RNP) to identify RNA-dependent associated proteins (RDAPs) co-bound to RNA with any RBP of interest. irCLIP-RNP defined landscapes of multimeric protein assemblies on RNA, uncovering previously unknown patterns of RBP-RNA associations, including cell-type-selective combinatorial relationships between RDAPs and primary RBPs. irCLIP-RNP also defined dynamic RDAP remodeling in response to epidermal growth factor (EGF), uncovering EGF-induced recruitment of UPF1 adjacent to HNRNPC to effect splicing surveillance of cell proliferation mRNAs. To identify the RNAs simultaneously co-bound by multiple studied RBPs, a sequential immunoprecipitation irCLIP (Re-CLIP) method was also developed. Re-CLIP confirmed binding relationships seen in irCLIP-RNP and identified HNRNPC and UPF1 RBP co-binding on RND3 and DDX3X mRNAs. irCLIP-RNP and Re-CLIP provide a framework to identify and characterize dynamic RNA-protein assemblies in living cells.

overall_design: infrared crosslinking immunoprecipiation followed by RNA sequencing (irCLIPv2) and sequential immunoprecipitation irCLIP (re-CLIP) followed by RNA sequencing for several RBPs in HEK293T and EGF-stimulated A431 cells.

first: Brian

last: Zarnegar

first: Robin

middle: M

last: Meyers

first: Douglas

last: Porter


title: Genome-wide absolute quantification of chromatin looping

status: Public on Jan 13 2025

city: Cambridge

first: James

middle: Martin

last: Jusuf

address: Building 56 Room 787B, 32 Vassar Street

department: Biological Engineering

institute: Massachusetts Institute of Technology

summary: We developed a method to estimate the 3D interaction probabilities of chromatin loops across the genome on an absolute scale from Micro-C maps. To calibrate the method, we performed Micro-C on two engineered mouse embryonic stem cell (mESC) lines, each containing a fluorescently labeled chromatin loop that was quantified in previous live imaging studies. One loop is an endogenous loop containing the Fbn2 gene, and the other is a synthetic loop near the Npr3 gene. We performed two replicates of Micro-C per cell line. Using our absolute quantification method, we find that loops generally form with low probabilities. We also provide an ultra-deep merged Micro-C map for mESCs that combines all existing mESC Micro-C datasets to date, containing a total of 15.6 billion unique interactions.

overall_design: Genome-wide Micro-C was performed on two replicates of the engineered cell line containing the fluorescently tagged Fbn2 loop ("Fbn2 cell line"), as well as two replicates of the engineered cell line containing the fluorescently tagged synthetic Npr3 loop ("Npr3 cell line"); all cells were in untreated condition.

first: Simon

last: Grosse-Holz

first: Michele

last: Gabriele

first: Pia

last: Mach

first: Ilya

last: Flyamer

first: Christoph

last: Zechner

last: Giorgetti

first: Leonid

last: Mirny

first: Anders

last: Hansen


title: Optimizing in vitro spherulation cues in the fungal pathogenCoccidioides

status: Public on Jun 20 2024

city: San Francisco

first: Christina

last: Homer

institute: UCSF

summary: Coccidioides spp.are part of a group ofthermallydimorphic fungal pathogens, which grow asfilamentous cells (hyphae)in the soil and transform to a different morphology upon inhalation into the host.TheCoccidioideshost form, the spherule, is unique and highly under characterized due to both technical and biocontainment challenges.Each spherule arises from an environmental spore (arthroconidium), matures, and develops hundreds of internal endospores, which are released from the spherule upon rupture. Each endospore can then go on to form another spherule in a cycle called spherulation. One of the foremosttechnicalchallenges has been reliably growing spherules in culture and consistently inducing endospore release withoutthe generation ofhyphae. Here, wepresentoptimization ofin vitro spherule growth and endospore release, by closely controlling starting cell density in the culture, usingfreshly-harvestedarthroconidia, and decreasing the concentration of multiple salts in spherulation media. We developed a minimal media to test spherule growth on various carbon and nitrogen sources andwent on to define a critical role forTamolin both early spherule formation and dispersal of the outer spherule wall, which accumulatesinto a dramatic large film inTamol’sabsence. Finally, we examined how the conditions under which arthroconidia are generated influence their transcriptome and subsequent development into spherules, demonstrating that this is an important variable to control when designing spherulation experiments. Together, our data reveal multiple strategies to optimize in vitro spherulation growth, enabling characterization ofthis virulence-relevant morphology.

overall_design: We generated spores (arthroconidia) and then aged them for 0-7 weeks in PBS at 4 degrees C and characterized their transcriptome after storage in these conditions. Additionally, we generated spores (arthroconidia) by growing hyphal cells for 4 weeks at either 25 degree C, 30 degrees C, or 37 degrees C and then determined their transcriptome by RNA-seq (3 replicates from each spore stock, each spore stock was generated 2 separate times).

first: Homer

last: Christina

first: Ochoa

last: Elena

first: Voorhies

last: Mark

first: Sil

last: Anita


title: Enfortumab Vedotin Induces Immunogenic Cell Death and Shows Enhanced Preclinical Antitumor Activity When Combined With a PD-1 Inhibitor

status: Public on Apr 19 2024

city: Bothell

first: John

last: Gosink

email: john@gosink.org, john.gosink@pfizer.com

state: WA

department: Computational Biology

institute: Pfizer

summary: Enfortumab vedotin is a Nectin-4-directed antibody–drug conjugate designed to deliver the microtubule-disrupting agent monomethyl auristatin E (MMAE) in tumor cells. Using preclinical models of urothelial cancer (UC), we expand the understanding of the multifaceted mechanism of action for enfortumab vedotin that includes direct cytotoxicity on Nectin-4–positive tumor cells, indirect bystander effect on neighboring Nectin-4–negative tumor cells, and MMAE-mediated induction of immunogenic cell death (ICD) and associated increase in activated immune cells in the tumor microenvironment. Importantly, vaccination with enfortumab vedotin-treated tumor cells drives antitumor immunity and provides protection against rechallenge in mice. MMAE-mediated ICD induction modulates the tumor microenvironment in a complementary manner to immune checkpoint inhibition. Accordingly, enfortumab vedotin plus PD-1 inhibitor shows enhanced antitumor activity in vivo. These preclinical results demonstrate the underlying mechanisms consistent with the significantly improved clinical outcomes observed for enfortumab vedotin plus pembrolizumab relative to chemotherapy in the first-line treatment of la/mUC.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: T24-hNectin-4 cells were cultured as described and resuspended in 50% Matrigel before subcutaneous injection of 5×106 T24-hNectin-4 xenograft tumor cells (0.1 mL volume) in the flanks of athymic nude female mice (Envigo, BALB/cAnNHsd). Tumor pieces were passaged via trocar needle two times to establish consistent engraftment. On Day 15 post implantation, tumor volumes reached ~200 mm3 (calculated using 0.5 × [length] × [width]2), and the mice were treated with a single intraperitoneal (IP) dose of 3 mg/kg enfortumab vedotin. As a control, mice were treated with 3 mg/kg IgG-MMAE. Tumors were harvested 5 days after treatment and processed for RNA-seq (Illumina HiSeq platform),

first: Devra

last: Olson

first: Patrick

last: Younan

first: Bernard

last: Liu

first: Gabriele

last: Blahnik-Fagan

first: Katie

last: Snead

first: Elena

last: Tenn

first: Kelly

last: Hensley

first: Disha

last: Saheytya

first: Albina

last: Nestrerova

first: Margo

last: Zaval

first: Anthony

last: Cao

first: Christine

last: O'Day

first: Ryan

last: Heiser

first: Timothy

last: Lewis

first: Shyra

last: Gardai

first: Taisuke

last: Nakazawa

first: Masashi

last: Shimazaki

first: Christopher

last: Carosina

first: Sharsti

last: Sandall


title: Plasma Sphingosine-1-Phosphate Orchestrates the Reverse Transendothelial Migration of Aortic Intimal Myeloid Cells

status: Public on Feb 22 2024

city: Toronto

first: Stylianos

last: Prattas

address: 101 College Street

country: Canada

institute: Toronto General Hospital Research Institute

summary: Background: Myeloid cells (MCs) reside in the aortic intima at regions predisposed to atherosclerosis. Systemic inflammation triggers reverse transendothelial migration (RTM) of intimal MCs into arterial blood, which orchestrates a protective immune response that clears intracellular pathogens from the arterial intima. Molecular pathways that regulate RTM remain poorly understood. Sphingosine-1-phosphate (S1P) is a lipid mediator that regulates immune cell trafficking by signaling via five G protein-coupled receptors (S1PRs). We investigated the role of S1P in the RTM of aortic intimal MCs.  Methods: Intravenous injection of lipopolysaccharide (LPS) was used to model a systemic inflammatory stimulus that triggers RTM. CD11c+ intimal MCs in the lesser curvature of the ascending aortic arch were enumerated by en face confocal microscopy. Local gene expression was evaluated by transcriptomic analysis of microdissected intimal cells.  Results: In wild-type C57BL/6 mice, LPS induced intimal cell expression of S1pr1, S1pr3, and sphingosine kinase 1 (Sphk1, a kinase responsible for S1P production). Pharmacological modulation of multiple S1PRs blocked LPS-induced RTM and modulation of S1PR1 and S1PR3 reduced RTM in an additive manner. Cre-mediated deletion of S1pr1 in MCs blocked LPS-induced RTM, confirming a role for myeloid-specific S1PR1 signaling. Global or hematopoietic deficiency of Sphk1 reduced plasma S1P levels, the abundance of CD11c+ MCs in the aortic intima and blunted LPS-induced RTM. In contrast, plasma S1P levels, the abundance of intimal MCs, and LPS-induced RTM were rescued in Sphk1-/- mice transplanted with Sphk1+/+ or mixed Sphk1 +/+ and -/- bone marrow. Stimulation with LPS increased endothelial permeability and intimal MC exposure to circulating factors such as S1P.  Conclusions: Functional and expression studies support a novel role for S1P signaling in the regulation of LPS-induced RTM as well as the homeostatic maintenance of aortic intimal MCs. Our data provides insight into how circulating plasma mediators help orchestrate intimal MC dynamics.

overall_design: Cells were isolated by micro-dissection of the lesser curvature aortic arch intima of C57BL/6J wildtype mice (control or injected with 30 ug ultrapure LPS i.v. 18 hours prior to harvest). Cells from 5-10 mice were pooled for each sample, and were subjected to scRNAseq.

first: Chanele

middle: K

last: Polenz

first: Corey

last: Scipione

first: Sharon

last: Hyduk

first: Myron

middle: I

last: Cybulsky


title: Human transcriptomic response to periprosthetic joint infection

status: Public on Feb 19 2024

city: Rochester

first: Aditya

last: Bhagwate

state: MN

address: 200 1st Street SW

institute: Mayo Clinic

summary: Periprosthetic joint infection (PJI), a devastating complication of total joint replacement, is of incompletely understood pathogenesis and may sometimes be challenging to clinically distinguish from other causes of arthroplasty failure. We characterized human gene expression in 93 specimens derived from surfaces of resected arthroplasties, comparing transcriptomes of subjects with infection- versus non-infection-associated arthroplasty failure. Differential gene expression analysis confirmed 28 previously reported potential biomarkers of PJI, including bactericidal/permeability increasing protein (BPI), cathelicidin antimicrobial peptide (CAMP), C-C-motif chemokine ligand 3 (CCL3), 4(CCL4), and C-X-C-motif chemokine ligand 2 (CXCL2), colony stimulating factor 2 receptor beta (CSF2RB), colony stimulating factor 3 (CSF3), alpha-defensin (DEFA4), Fc fragment of IgG receptor 1B (CD64B), intercellular adhesion molecule 1 (ICAM1), interferon gamma (IFNG), interleukin 13 receptor subunit alpha 2 (IL13RA2), interleukin 17D (IL17D), interleukin 1 (IL1A, IL1B, IL1RN), interleukin 2 receptors (IL2RA, IL2RG), interleukin 5 receptor (IL5RA), interleukin 6 (IL6), interleukin 8 (IL8), lipopolysaccharide binding protein (LBP), lipocalin (LCN2), lactate dehydrogenase C (LDHC), lactotransferrin (LTF), matrix metallopeptidase 3 (MMP3), peptidase inhibitor 3 (PI3), and vascular endothelial growth factor A (VEGFA), and identified three novel molecules of potential diagnostic use for detection of PJI, namely C-C-motif chemokine ligand CCL20, coagulation factor VII (F7), and B cell receptor FCRL4. Comparative analysis of infections caused by staphylococci versus bacteria other than staphylococci and Staphylococcus aureus versus Staphylococcus epidermidis showed elevated expression of interleukin 13 (IL13), IL17D, and MMP3 in staphylococcal infections, and of IL1B, IL8, and platelet factor PF4V1 in S. aureus compared to S. epidermidis infections. Pathway analysis of over-represented genes suggested activation of host immune response and cellular maintenance and repair functions in response to invasion of infectious agents. The data presented provides new potential targets for diagnosis of PJI and for differentiation of PJI caused by different infectious agents.

overall_design: Human host RNA from sonicate fluids collected from resected implants of arthroplasty failure individuals were analyzed for gene expression.

first: Thao

last: Masters

middle: V

first: Mrunal

last: Dehankar

first: Kerryl

middle: E

last: Greenwood-Quaintance

first: Matthew

middle: P

last: Abdel

first: Jay

middle: N

last: Mandrekar

first: Robin

last: Patel


title: Single-cell analysis of tobacco-associated lung adenocarcinoma development

status: Public on Feb 16 2024

city: Houston

first: Humam

last: Kadara

address: 2130 W. Holcombe Blvd

institute: University of Texas MD Anderson Cancer Center

summary: Understanding cellular processes underlying early lung adenocarcinoma (LUAD) development is needed to devise intervention strategies. Here, we performed single-cell RNA sequencing (scRNA-seq) of mouse lungs from Gprc5a-/- mice during lung tumor development. We coupled scRNA-seq analysis with spatial transcriptomics of tumor-bearing lungs.

overall_design: scRNA-seq profiling was performed on lungs from Gprc5a-/- mice treated with nicotine-derived nitrosamine ketone (NNK) or saline at the end of exposure (EOE, timepoint at which lungs are still clear of lesions), at 3 months and at 7 months post-exposure, the time point of Kras-mutant LUAD onset in all mice (n = 4 mice per group and time point). We also performed scRNA-seq of GFP+ and GFP- fractions of lungs from Gprc5a-/- mice carrying alveolar type II reporter (Gprc5a-/-; SftpcCreER/+; RosaSun1GFP/+) at three months post-exposure to saline of NNK (n = 2 mice per group). scRNA-seq was performed using the 10X Genomics platform. Spatial transcriptomics of tumor-bearing lungs from Gprc5a-/- mice at 7 months post-exposure to NNK (n = 3) was performed using the Visium platform from 10X Genomics to further characterize and validate scRNA-seq findings in a spatial context.

first: Guangchun

last: Han

first: Fuduan

last: Peng


title: Analyses of barcode repertoire in hematopoietic cells

status: Public on Feb 13 2024

city: Chiba

first: Ichiro

last: Manabe

address: 1-8-1 Inohana, Chuo

country: Japan

institute: Chiba University

summary: We compared the barcode repertoire in circulating leukocytes, and cardiac and renal macrophages.

overall_design: Amplicon sequencing.

first: Manabe

last: Ichiro

first: Nakayama

last: Yukiteru

first: Fujiu

last: Katsuhito


title: Transcriptomic and epigenetic analyses of hematopoietic stem cells

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

overall_design: Refer to individual Series


title: Expression analyses of circulating monocytes and cardiac macrophages in hematopoietic stem cell (HSC)-transplanted mice

summary: We analyzed transcriptomes of blood and cardiac macrophages in the mice transplanted with the mixture of HSCs from control and four-weeks post-transverse aortic constriction (TAC) mice.

overall_design: RNA-seq profiling of sorted monocytes and cardiac macrophages.


title: Epigenetic analyses of hematopoietic stem cells

summary: We analyzed genome-wide chromatin accessibility in hematopoietic stem cells (HSCs) using ATAC-seq in mice after sham operation and transverse aortic constriction (TAC).

overall_design: ATAC-seq of HSCs.


title: Multiple Generation Distinct Toxicant Exposures Induce Epigenetic-Transgenerational-Inheritance of Enhanced Pathology and Obesity

status: Public on Dec 02 2024

city: Pullman

first: Michael

last: Skinner

address: Abelson 507

department: SBS

institute: WSU

type: Methylation profiling by high throughput sequencing

summary: This study examines the actions of the agricultural fungicide vinclozolin on the F0 generation, followed by jet fuel hydrocarbon mixture exposure of the F1 generation, and then pesticide DDT (dichlorodiphenyltrichloroethane) on the F2 generation gestating females. The subsequent F3 and F4 generations and F5 transgenerational-generation were examined for male sperm epigenetic alterations and pathology in males and females. Significant impacts on the male sperm differential DNA methylation regions (DMRs) were observed. The F3-F5 generations were similar in approximately 50% of the DMRs. The pathology of each generation was assessed in the testis, ovary, kidney, prostate, obesity, and tumors. The pathology used a newly developed Deep Learning (DL), artificial intelligence (AI), based histopathology analysis. Observations demonstrated compounded disease impacts in obesity and metabolic parameters, but other pathologies plateaued with smaller increases at the F5 transgenerational generation. Therefore, the multiple generational exposures, which occurs in human populations, may increase epigenetic impacts and disease susceptibility.

overall_design: The experimental design used three successive generations of gestating female exposures during fetal gonadal sex determination, followed by three generations of no exposure to obtain the F5 generation to reveal the ultimate transgenerational phenotype for pathology and disease. The F0 generation gestating female was exposed to the agricultural fungicide vinclozolin. The second successive exposure of the F1 generation gestating female involved jet fuel (JP8). The third successive exposure of the F2 generation gestating female involved DDT (dichlorodiphenyltrichloroethane). The subsequent F3 and F4 generations and F5 transgenerational-generation were examined for male sperm epigenetic alterations and pathology in males and females. Methylation differences between treatment groups was done using an MeDIP-seq procedure. The control lineage was only taken to the F3 generation. This F3 generation control was then compared with the F3, F4, and F5 treatment lineage sample groups.

first: Eric

last: Nilsson

first: Margaux

last: McBirney

first: Sarah

last: De Santos

first: Stephanie

last: King

first: Daniel

last: Beck

first: Colin

last: Greeley

first: Lawrence

middle: B

last: Holder


title: Transcriptomic analyses of hematopoietic stem cells

summary: We analyzed transcriptomes of hematopoietic stem cells (HSCs) using RNA-seq in mice after sham operation and transverse aortic constriction (TAC).

overall_design: RNA-seq profiling of HSCs.


title: Granulocytic myeloid-derived suppressor cell activity during biofilm infection is regulated by a glycolysis-HIF-1α axis II

status: Public on Feb 08 2024

city: Omaha

first: Sankarasubramanian

last: Jagadesan

state: NE

address: 42end and Emile st

department: Department of Genetics, Cell Biology & Anatomy

institute: University of Nebraska Medical Center

summary: Staphylococcus aureus is a leading cause of biofilm-associated prosthetic joint infection (PJI). A primary contributor to infection chronicity is an expansion of granulocytic myeloid-derived suppressor cells (G-MDSCs) that are critical for orchestrating the anti-inflammatory biofilm milieu. Single-cell sequencing and bioinformatic metabolic algorithms were used to explore the link between G-MDSC metabolism and S. aureus PJI outcome. Glycolysis and the hypoxic response through hypoxia-inducible factor-1 alpha (HIF-1α) were significantly enriched in G-MDSCs. Interfering with both pathways in vivo, using a 2-deoxyglucose nanopreparation and granulocyte-targeted HIF-1α conditional knockout mice, respectively, attenuated G-MDSC-mediated immunosuppression and reduced bacterial burden in a mouse model of S. aureus PJI. In addition, scRNA-seq analysis of granulocytes from PJI patients also showed an enrichment in glycolysis and hypoxic response genes. These findings support the importance of a glycolysis/HIF-1α axis in promoting G-MDSC anti-inflammatory activity and biofilm persistence during PJI.

overall_design: Cells were collected from the blood and periprosthetic soft tissue of human subjects with prosthetic joint infection by FACS and prepared for single-cell sequencing using the 10X Genomics platform.

first: Chris

last: Horn

first: Prabhakar

last: Arumugam

first: Cortney

last: Heim

first: Tammy

last: Kielian


title: Targeted proteomics analysis of Cutaneous Lupus Erythematosus patient interstitial skin fluid and plasma (Inflammation panel data)

status: Public on Jan 31 2024

city: North Grafton

first: Jillian

last: Richmond

state: Massachusetts

address: 200 Westboro Road

department: Comparative Pathobiology

institute: Tufts Cummings School of Veterinary Medicine

summary: We performed a targeted proteomics analysis of Cutaneous Lupus Erythematosus (CLE) patient blister biopsies and plasma using Olink Proteomics panels to validate the blister biopsy technique using previously identified biomarkers, to confirm protein-level expression of biomarkers previously identified at the RNA level, and to identify new biomarkers of disease.

overall_design: Protein biomarkers in lesional and nonlesional interstitial skin fluid and blood plasma from 4 CLE patients and 3 healthy controls were measured using the Olink Proteomics Target 96 Inflammation panel v.3022.

first: Mehdi

last: Rashighi


title: Granulocytic myeloid-derived suppressor cell activity during biofilm infection is regulated by a glycolysis-HIF-1α axis I

overall_design: Live CD45+ cells were collected from the soft tissue adjacent to the knee joint of Mrp8CreHif1afl/fl and WT littermates (Mrp8NullHif1afl/fl) at days 3 and 14 post-infection by FACS and prepared for single-cell sequencing using the 10X Genomics platform.


title: Single-cell sequencing Unveils Age-Dependent Differential Impacts of Mxra7 Knockout on Mouse Bone Marrow Immune Cell Development and Function

status: Public on Feb 04 2024

city: Suzhou

first: Yiqiang

last: Wang

state: Jiangsu

address: 111 Renai Road

institute: Xi'an Jiaotong-Liverpool University

summary: Matrix remodeling associated 7 (Mxra7) belongs to the matrix remodeling-related gene family and is implicated in inflammatory neovascularization, liver injury, and autoimmune skin diseases. To elucidate the composition and transcriptional changes of bone marrow cells in Mxra7 knockout mice, we conducted single-cell RNA sequencing (scRNA-seq) in two-month-old and two-year-old mice. Our investigation revealed that Mxra7 knockout influences early B cell development and normal cell cycle functioning, impacting processes such as chromosome segregation and cell mitosis. Additionally, bone marrow B cells from knockout mice exhibited heightened differentiation potential compared to wild-type counterparts in aged mice. Mxra7 knockout initially suppressed basophil development at two months of age, only to yield an increased basophil proportation in two-year-old mice. Moreover, knockout mice exhibited elevated basophil numbers within bone marrow granulocytes. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis indicated alterations in neurological disease-related functions in Mxra7 knockout bone marrow granulocytes. Cell communication analysis highlighted heightened quantity and quality of cellular interactions within bone marrow cells of knockout mice, notably enriching three inflammation-regulating signaling pathways. These findings underscore the pivotal role of Mxra7 in bone marrow immune cell development, with age-dependent effects on immune and inflammatory responses.

overall_design: Comprehensive analysis of single-cell RNA-seq data revealed differences between two-month-old and two-year-old wild-type (WT) and Mxra7 knockout (KO) mice.

first: Yihan

last: Chen

first: Yudan

last: Zheng


title: Granulocytic myeloid-derived suppressor cell activity during biofilm infection is regulated by a glycolysis-HIF-1α axis

status: Public on Jan 23 2024


title: Impact of Vitamin-D3 on the gene expression profile of mouse colon

status: Public on Nov 29 2024

city: London

first: Probir

last: Chakravarty

address: 1Midland Road

institute: Francis Crick

summary:
Vitamin-D3 (VitD3) exhibits pleiotropic effects in host’s physiology by acting on several organs. To mechanistically understand how VitD3 influences the colon physiology we interrogated the gene expression profile of mouse colon in response to high or low VitD3 bioavailability. VitD3 tissue bioavailability was modulated in mice with the use of diets supplemented with different concentrations of VitD3 (0, 2 or 10 IU/g) and the use of Gc knockout mouse strain in where VitD3 bioavailability is enhanced.
Vitamin-D3 (VitD3) exhibits pleiotropic effects in host’s physiology by acting on several organs. To mechanistically understand how VitD3 influences the colon physiology we interrogated the gene expression profile of mouse colon in response to high or low VitD3 bioavailability. VitD3 tissue bioavailability was modulated in mice with the use of diets supplemented with different concentrations of VitD3 (0, 2 or 10 IU/g) and the use of Gc knockout mouse strain in where VitD3 bioavailability is enhanced.

overall_design: RNA will be extracted from 5 biological replicates (5 individual mice) per condition (20 samples in total). Male mice (8 ½ weeks old) were fed with the following diets 3 ½ weeks :1)C57BL/6J/WT mice that were fed with standard VitD3 diet (2 IU/g) for 3 ½ weeks (n=5), 2) C57BL/6J/WT mice that were fed with high VitD3 diet (10 IU/g) for 3 ½ weeks (n=5), 3) Gc KO mice that were fed with standard VitD3 diet (2 IU/g) for 3 ½ weeks (n=5), 4) Gc KO mice that were fed with VitD3 deficient diet (10 IU/g) for 3 ½ weeks (n=5)

first: Evangelos

last: Giampazolias

first: Caetano

last: Reis e Sousa


title: Prophilactic and long-lasting efficacy of senolytic CAR T cells against age-related metabolic dysfunction.

status: Public on Dec 08 2023

city: New York

first: Yu-Jui

last: Ho

address: 1275 York Avenue

institute: Memorial Sloan Kettering Cancer Center

summary: Senescent cells accumulate in organisms over time as a result of tissue damage and impaired immune surveillance and are thought to contribute to age-related tissue decline1,2. In agreement, genetic ablation studies reveal that elimination of senescent cells from aged tissues can ameliorate various age-related pathologies, including metabolic dysfunction and decreased physical fitness3-7. While small-molecule drugs capable of eliminating senescent cells (known as ‘senolytics’) partially replicate these phenotypes, many have uncertain mechanisms of action and all require continuous administration to be effective. As an alternative approach, we previously developed a cell-based senolytic therapy based on chimeric antigen receptor (CAR) T cells targeting uPAR, a cell-surface protein upregulated on senescent cells, and showed these can safely and efficiently eliminate senescent cells in young animals and reverse liver fibrosis8. We now show that uPAR-positive senescent cells accumulate during physiological aging and that they can be safely targeted with senolytic CAR T cells in aged animals. Treatment with anti uPAR CAR T cells ameliorates metabolic dysfunction by improving glucose tolerance and exercise capacity in physiological aging as well as in a model of metabolic syndrome. Importantly, the beneficial effects of senolytic CAR T cells are long lasting; single administration of a low dose is sufficient to achieve long-term therapeutic and preventive effects.

overall_design: Liver, gonadal adipose tissue and pancreas of 4 aged mice (20 months old; 2 females and 2 males) was dissociated into single cell suspension and subsequently stained for surface uPAR expression. Cells were FACS sorted into uPAR+ and uPAR- populations for each of the organs. After cell sorting the two females and the two males of each organ and population were pooled together so that each organ would have a replicate consisting of 2 pooled male samples and 2 pooled female samples. Single-cell RNAseq was then carried out using 10X Genomics 3' Gene Expression chemistry.

first: Corina

last: Amor

first: Jonathan

last: Preall


title: Granulocytic myeloid-derived suppressor cell activity during biofilm infection is regulated by a glycolysis-HIF-1α axis (scRNA-Seq)

overall_design: Live CD45+ cells were collected from the soft tissue adjacent to the knee joint of C57BL/6J mice at days 3, 7, and 14 post-infection by FACS and prepared for single-cell sequencing using the 10X Genomics platform.


title: Targeted proteomics analysis of Cutaneous Lupus Erythematosus patient interstitial skin fluid and plasma (Neuro Exploratory panel data)

overall_design: Protein biomarkers in lesional and nonlesional interstitial skin fluid and blood plasma from 4 CLE patients and 3 healthy controls were measured using the Olink Proteomics Target 96 Neuro Exploratory panel v.3911.


title: Gene expression profile at single cell level of pre-cDCs from the bone marrow, spleen and lung of mice

status: Public on Nov 29 2023

summary: Type 2 pre-cDCs are considered as a homogeneous population in the bone marrow. We used single cell RNA sequencing to show that  the bone marrow contains two subsets of pre-cDC2s that are commited  to give rise to different different cDC2 subsets in peripheral tissues.

overall_design: pre-cDCs from the bone marrow, spleen and lung of WT C57BL6 mice were isolated by Fluorescence-activated cell sorting (FACS) and analyzed using scRNAseq.

first: Carlos

last: Minutti


title: Alpha-ketoglutarate dehydrogenase is a therapeutic vulnerability in acute myeloid leukemia [KL974_shOGDH]

status: Public on Nov 27 2024

summary: Perturbations in intermediary metabolism contribute to the pathogenesis of acute myeloid leukemia (AML) and can produce therapeutically actionable dependencies. Here, we probed whether alpha-ketoglutarate (aKG) metabolism represents a specific vulnerability in AML. Using functional genomics, metabolomics, and mouse models, we identified the aKG dehydrogenase complex, which catalyzes the conversion of aKG to succinyl CoA, as a molecular dependency across multiple models of adverse-risk AML. Inhibition of 2-oxoglutarate dehydrogenase (OGDH), the E1 subunit of the aKG dehydrogenase complex, impaired AML progression and drove differentiation. Mechanistically, hindrance of aKG flux through the tricarboxylic acid (TCA) cycle resulted in rapid exhaustion of aspartate pools and blockade of de novo nucleotide biosynthesis, while cellular bioenergetics was largely preserved. Additionally, increased aKG levels following OGDH inhibition impacted the biosynthesis of other critical amino acids. Thus, this work has identified a previously undescribed, functional link between certain TCA cycle components and nucleotide biosynthesis enzymes across AML. This metabolic node may serve as a cancer-specific vulnerability amenable to therapeutic targeting in AML and perhaps in other cancers with similar metabolic wiring.

overall_design: RNA sequencing of murine p53-mutant AML cell lines was perfored at days 0, 2, and/or 4 after induction of the indicated BFP-linked shRNAs with doxycycline. Two independent shRNAs targeting Ogdh (shOgdh346 and shOgdh2081) and a control shRNA (shRen713) were used. Sorted, induced (BFPplus) cells are included for all shRNAs. Undinduced (BFPmin) cells at each timepoint are included for control samples.

first: Scott

last: Millman

first: Almudena

last: Chaves-Perez

first: Sudha

last: Janaki-Raman

first: John

last: Morris IV

first: Varun

last: Narendra

first: Chi-Chao

first: Benjamin

middle: T

last: Jackson

first: Jossie

last: Yashinskie

first: Riccardo

last: Mezzadra

first: Tessa

last: Devine

first: Valentin J

middle: A

last: Barthet

first: Michelle

last: Saoi

first: Timour

last: Baslan

first: Sha

last: Tian

first: Zohar

last: Sachs

first: Lydia W

middle: S

last: Finley

first: Justin

middle: R

last: Cross

middle: W

last: Lowe


title: Single-cell multi-omics profiling of human  preimplantation embryos identifies  cytoskeletal defects during embryonic arrest

status: Public on Nov 17 2023

city: GuangZhou

first: Teng

last: Wang

address: jingxiStreet

institute: Southern Medical University

summary: Human in vitro-fertilized (IVF) embryos exhibit low developmental capabilities. However, the mechanisms underlying human embryonic arrest remain unclear. Here, using single-cell multi-omics sequencing approach, we simultaneously dissected the alteration of transcriptome, chromatin accessibility and DNA methylome in human early embryos arrested for unexplained reasons. Human arrested embryos displayed transcriptome disorders, including distorted microtubule cytoskeleton, increased genomic instability and impaired glycolysis, which were coordinated with multiple epigenetic reprogramming defects. Moreover, we identified Aurora A kinase (AURKA) repression was a reason for human embryonic arrest. Mechanistically, human arrested embryos from AURKA inhibited group resembled the reprogramming abnormalties of clinically arrested embryos in terms of transcriptome, DNA methylome, chromatin accessibility and H3K4me3. Through mitosis-independent sequential activation of zygotic genome in human arrested embryos, we found YY1 contributed to human major zygotic genome activation (ZGA). Collectively, our study decodes the reprogramming abnormalities and mechanisms for human embryonic arrest and key regulators of human ZGA.

overall_design:
We collected human arrested zygote, 2-cell, 4-cell and 8-cell embryos that were with no further development for another two days, followed by performing single-cell multi-omics sequencing including transcriptome, DNA methylome and chromatin accessibility. Additionally, we performed single-embryo multi-omics sequencing of human 8-cell embryos (3PN) from YY1 knockdown group and control group, single-embryo RNA sequencing of human 8-cell embryos (3PN) from KDOAM25 treated group and control group, single-cell multi-omics sequencing of human arrested zygote (3PN) from AURKA inhibited group and control zygotes, and single-cell RNA sequencing of mouse arrested zygote from AURKA inhibited group and control embryos (raw sequencing data of mouse embryos have been deposited in Genome Sequence Archive (GSA) database with the accession number CRA012505; processed data for mouse samples are available in the OMIX database (China National Center for Bioinformation/Beijing Institute of Genomics, Chinese Academy of Sciences) with accession number OMIX004867).

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Due to the relevant regulation from China's Ministry of Science and Technology related to the export of genetic information and materials, all raw sequencing data of human embryos reported in this study have been deposited in Genome Sequence Archive (GSA) for human database with the accession number HRA003366.
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first: Lin

last: Li

first: Lei

first: Junhua

last: Peng

first: Jiaqi

last: Fan


title: An integrative single-cell multiomics profiling of human pancreatic islets identifies T1D associated genes, regulatory regions, and single nucleotide polymorphism

status: Public on Nov 13 2023

first: Shuibing

last: Chen

state: NY

address: A 827B, 1300 York Ave

department: Surgery

institute: Weill Cornell Medical College