title: Decoding the Escherichia coli epitranscriptome

city: Martinsried

first: Kirsten

last: Jung

address: Großhaderner Str. 2-4

institute: LMU

summary: Modifications of RNA, known as the epitranscriptome, affect mRNA stability, translation, and splicing in eukaryotes and have implications for developmental processes, cancer, and viral infections. In prokaryotes, however, the landscape of the epitranscriptome is still poorly understood. To address this knowledge gap, we used direct RNA sequencing with Nanopore technology to study RNA modifications in the model bacterium Escherichia coli. With a single sequencing reaction, we were able to simultaneously identify and map most of the known modification types in rRNA, tRNA, and mRNA. Subsequently, a multifaceted approach integrating different algorithms for data analysis, deletion mutants, mass spectrometry, qPCR, and in vitro methylation was implemented to evaluate the presence of m5C and m6A in E. coli. Known m5C and m6A sites in rRNA were confirmed, but these modifications could not be localized in the mRNA. Nevertheless, based on the sequencing data, modifications were found to be enriched in the coding regions of genes associated with general metabolism and RNA processing. This study provides a useful resource for experimental and bioinformatic approaches to gain new insights into post-transcriptional regulation in a prokaryotic model.

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

overall_design:
Total RNA was extracted from two biological replicates each of wild-type (WT) and ∆rlmF∆rlmJ mutant E. coli cultured under standard growth conditions and from two biological replicates of WT heat-shocked E. coli to measure RNA modifications throughout the transcriptome.
Two replicates of in vitro transcribed RNA were synthesized from WT E. coli DNA template as unmodified controls.
Putative modifications were detected after samples were processed with direct RNA sequencing via Nanopore.

first: Sebastian

last: Riquelme Barrios

first: Leonardo

last: Vasquez Camus

first: Siobhan

last: Cusack

first: Korinna

last: Burdack

first: Dimitar

last: Plamenov Petrov

first: Nur

last: Yeşiltaç

first: Stefanie

last: Kaiser

first: Pascal

last: Giehr


title: Patient derived model of UBA5-associated encephalopathy identifies defects in neurodevelopment and highlights potential therapies [scRNA-seq]

status: Public on Feb 21 2025

city: Memphis

first: Yong-Dong

last: Wang

state: TN

address: 262 Danny Thomas Pl, MS1135

country: USA

institute: St. Jude Children's Research Hospital

type: Expression profiling by high throughput sequencing

summary: UBA5 encodes for the E1 enzyme of the UFMylation cascade, which plays an essential role in ER homeostasis. The clinical phenotypes of UBA5-associated encephalopathy includes developmental delays, epilepsy and intellectual disability. To date, there is no humanized neuronal model to study the cellular and molecular consequences of UBA5 pathogenic variants. We developed and characterized patient-derived cortical organoid cultures and identified defects in GABAergic interneuron development. We demonstrated aberrant neuronal firing and microcephaly phenotypes in affected organoids. Mechanistically, we show that ER homeostasis is perturbed along with exacerbated unfolded protein response pathway in cells expressing UBA5 pathogenic variants. We also assessed two gene expression modalities that augmented UBA5 expression to rescue aberrant molecular and cellular phenotypes. Our study provides a novel humanized model that allows further investigations of UBA5 variants in the brain and highlights novel systemic approaches to alleviate cellular aberrations for this rare, developmental disorder.

overall_design: scRNA-seq samples from proband cortical organoids compared to those from control cortical organoids of parent

first: Helen

last: Chen

first: Christy

middle: W

last: LaFlamme

first: Heather

middle: C

last: Mefford


title: Rapid folding of nascent RNA regulates eukaryotic RNA biogenesis

city: Worcester

first: Leonard

last: Schärfen

state: MA

address: 368 Plantation Street

department: RNA Therapeutics Institute

institute: UMass Chan Medical School

summary: RNA’s catalytic, regulatory, or coding potential depends on structure formation. Because base-pairing occurs during transcription, early structural states can govern RNA processing events and dictate the formation of functional conformations. These co-transcriptional states remain unknown. Here, we develop CoSTseq, which detects nascent RNA base-pairing within and upon exit from RNA polymerases (Pols) transcriptome-wide in living yeast cells. Monitoring each nucleotide’s base-pairing activity during transcription, CoSTseq reveals predominantly rapid pairing – within 25 bp of transcription after addition to the nascent chain. Moreover, ~23% of rRNA nucleotides attain their final base-pairing state near Pol I, while most other nucleotides must undergo changes in pairing status during later steps of ribosome biogenesis. We show that helicases act immediately to remodel structures across the rDNA locus to facilitate ribosome biogenesis. In contrast, nascent pre-mRNAs attain local structures indistinguishable from mature mRNAs, suggesting that refolding behind elongating ribosomes resembles co-transcriptional folding behind Pol II.

overall_design: To investigate how nascent RNA base pairs after exit from polymerases, we developed CoSTseq. CoSTseq is a method that detects base pairing of nascent RNA. We perform CoSTseq in Saccharomyces cerevisiae of several genetic backgrounds and compared it with DMS-MaPseq of the same strains. We include at least three biological replicates for each strain.

first: Isaac

last: Vock

first: Matthew

middle: D

last: Simon

first: Karla

middle: M

last: Neugebauer


title: Recapitulation of endochondral ossification by human pluripotent stem cell-derived SOX9+ sclerotomal progenitors

city: Chengdu

first: Zhonghan

last: Li

state: Sichuan

address: 29 Wangjiang Road

department: College of Life Sciences

country: China

institute: Sichuan University

summary: Regeneration of human cartilage and bone remains a challenge due to the limitations from current stem cell sources. Here, we developed a four-day differentiation strategy that achieved near uniform derivation (>98.5%) of SOX9+ sclerotomal progenitors from multiple human pluripotent stem cells (hPSCs). Importantly, SOX9+ sclerotomal progenitors exhibited typical bipotential of mesenchymal progenitors during skeletal development. Upon lineage-specific induction, they were able to differentiate either into chondroprogenitors that could repair articular cartilage defects and  early chondrocytes that could undergo endochondral ossification for human bone formation. Furthermore, we identified ITGA9 as the specific surface marker that facilitated reporter-independent isolation of SOX9+ sclerotomal progenitors and established an in vitro culture system that could expand these cells for at least 15,000-fold. Collectively, these findings highlight the SOX9+ sclerotomal progenitors as a promising stem cell source for next-generation human cartilage/bone bioengineering.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for human iPSC-derived  SOX9+ sclerotomal progenitors, Sox9- cells and the undifferentiated human iPSCs

first: Jingfei

last: Xiong

first: Yuansong

last: Liao

first: Yanhui

last: Deng

first: Hanyi

first: Ce

last: Shan

first: Runxin

last: Ma

first: Guogen

last: Ye

first: Kun

last: Xie

first: Haoyang

last: Cai

first: Qing

last: Zhu

first: Weihua

last: Guo

first: Yike

last: Yin

first: Yunqiu

last: Zhang

first: Yuqing

first: Yifu


title: Patient derived model of UBA5-associated encephalopathy identifies defects in neurodevelopment and highlights potential therapies [bulk RNA-seq]

first: Yong-Dong

last: Wang

overall_design: Bulk RNA-seq samples from proband cortical organoids compared to those from control cortical organoids of parents

first: Helen

last: Chen

first: Heather

last: Mefford


title: Glucose controls protein-protein interactions and tissue differentiation [RNA-Seq]

city: Stanford

first: Vanessa

last: Lopez-Pajares

state: CA

address: 269 Campus Dr., CCSR 2150

department: Dermatology

institute: Stanford University

summary:
Abstract: Glucose serves as a universal energy currency in living organisms, however, its potential non-energetic biomolecular functions are less well defined. Glucose was among the most increased analytes among >14,000 assessed across epidermal differentiation, an elevation verified in tissue engineered with fluorescent glucose sensors and also observed in differentiating cells from other tissues. Free glucose accumulation, but not its increased metabolism, was essential for epidermal differentiation and required GLUT1, GLUT3, and SLC5A1 transporters. Glucose affinity chromatography and azido-glucose click chemistry revealed direct glucose binding to a variety of regulatory proteins, including the IRF6 transcription factor (TF), whose epidermal knockout confirmed its requirement in glucose-dependent differentiation. Glucose binding mediated IRF6 dimerization, DNA affinity, and genomic targeting. The IRF6R84C mutant found in poorly differentiated cancers was unable to bind glucose. These data demonstrate a non-energetic role for glucose in modulating protein multimerization to control genome dynamics.
Purpose: To assess the global transcriptional program in epidermal organoids where glucose has been modulated and to assess the  transcriptome in the context of IRF6 loss.

overall_design: Epidermal organoids derived from primary human keratinocytes isolated from foreskin grown in normal or low glucose media or CRISPR-mediated IRF6 KO


title: Alteration of global transcription by saccharin in Escherichia coli K12

city: Uxbridge

first: Rubén

last: de Dios

state: Middlesex

address: Kingston Lane

department: Life Sciences

country: United Kingdom

institute: Brunel University London

summary: We perform differential RNA-seq comparing a treatment with 1.4% saccharin to a mock treatment in E. coli K12 in order to describe the alterations in global transcription produced by this artificial sweetener. Three biological replicates  of saturated E. coli K12 cultures were diluted 1/100 in 25 mL LB medium (in duplicate, to obtain 3 treated cultures and 3 mock controls). The cultures were grown (37 ºC, 180 rpm) to approximately OD600 0.3 and they were treated with a final concentration of 1.4% saccharin or a water control. The cultures were further incubated until reacing OD600 0.8. At this point, cells were harvested, treated with RNAlate for preservation of total RNA and stored at -80 C. After that, total RNA was extracted from each sample. As a result, 305 genes appeared upregulated by the saccharin treatment, whereas 419 were downregulated.

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

first: Evgenia

last: Maslova

first: Ronan

middle: R

last: McCarthy


title: Glucose controls protein-protein interactions and tissue differentiation [ATAC-Seq]

type: Genome binding/occupancy profiling by high throughput sequencing

summary:
Abstract: Glucose serves as a universal energy currency in living organisms, however, its potential non-energetic biomolecular functions are less well defined. Glucose was among the most increased analytes among >14,000 assessed across epidermal differentiation, an elevation verified in tissue engineered with fluorescent glucose sensors and also observed in differentiating cells from other tissues. Free glucose accumulation, but not its increased metabolism, was essential for epidermal differentiation and required GLUT1, GLUT3, and SLC5A1 transporters. Glucose affinity chromatography and azido-glucose click chemistry revealed direct glucose binding to a variety of regulatory proteins, including the IRF6 transcription factor (TF), whose epidermal knockout confirmed its requirement in glucose-dependent differentiation. Glucose binding mediated IRF6 dimerization, DNA affinity, and genomic targeting. The IRF6R84C mutant found in poorly differentiated cancers was unable to bind glucose. These data demonstrate a non-energetic role for glucose in modulating protein multimerization to control genome dynamics.
Purpose: To determine the impact of glucose modulation on chromatin accessibility during keratinocyte differentiation and the impact of IRF6 loss on chromatin accessibilty in differentiated keratincoytes

overall_design: Primary human keratinocytes isolated from foreskin maintained as progenitors or differentiated and grown in normal or low glucose media or differentiated keratinocytes with IRF6 KO


title: Glucose controls protein-protein interactions and tissue differentiation [ChIP-Seq]

summary:
Abstract: Glucose serves as a universal energy currency in living organisms, however, its potential non-energetic biomolecular functions are less well defined. Glucose was among the most increased analytes among >14,000 assessed across epidermal differentiation, an elevation verified in tissue engineered with fluorescent glucose sensors and also observed in differentiating cells from other tissues. Free glucose accumulation, but not its increased metabolism, was essential for epidermal differentiation and required GLUT1, GLUT3, and SLC5A1 transporters. Glucose affinity chromatography and azido-glucose click chemistry revealed direct glucose binding to a variety of regulatory proteins, including the IRF6 transcription factor (TF), whose epidermal knockout confirmed its requirement in glucose-dependent differentiation. Glucose binding mediated IRF6 dimerization, DNA affinity, and genomic targeting. The IRF6R84C mutant found in poorly differentiated cancers was unable to bind glucose. These data demonstrate a non-energetic role for glucose in modulating protein multimerization to control genome dynamics.
Purpose: To determine the impact of glucose modulation on IRF6 genomic localization during keratinocyte differentiation

overall_design: IRF6 ChIP profile in differentiated keratinocytes grown in normal (4.5g/L) or low (0.5g/L) glucose


title: Experimental challenge with bovine herpesvirus 1 in dairy calves: cranial lung lobe mRNA transcriptome response

city: Dunsany

first: Dayle

last: Johnston

state: Ireland

address: AGRIC

department: Animal and Bioscience Research Department

country: Ireland

institute: Teagasc

summary: Bovine Herpesvirus 1 (BoHV1) is a leading cause of Bovine Respiratory Disease (BRD) in young calves, which is responsible for substantial morbidity and mortality. Therefore, the objective of the current study was to elucidate the cranial lung lobe mRNA transcriptomic response to an experimental challenge with BoHV1, in dairy calves. Holstein-Friesian calves were either challenged by intranasal atomisation with BoHV1 virus (6.3 x 10^7/mL x 1.35mL) (n=12) or mock challenged with sterile phosphate buffered saline (n=6). Clinical signs were scored daily until euthanasia at day 6 post-challenge. Total RNA was extracted and sequenced from cranial lung lobe tissue (lesioned and healthy) (150 bp paired-end). Sequence reads were aligned to the ARS-UCD1.2 bovine reference genome and differential gene expression analysis was performed using EdgeR. There were 334 differentially expressed genes in the healthy cranial lobe tissue, between BoHV1 challenged and control calves, and there were 67 differentially expressed genes

overall_design: Examination of a single tissue type (cranial lung lobe), in two different conditions, healthy or lesioned, in two different treatment groups, BoHV1 challenged calves (n=12) and control calves (n=6)

first: Bernadette

last: Earley

first: Matthew

middle: S

last: McCabe

first: JaeWoo

last: Kim

first: Jeremy

middle: F

last: Taylor

first: Ken

last: Lemon

first: Catherine

last: Duffy

first: Michael

last: McMenamy

first: S

middle: L

last: Cosby

first: Sinead

last: Waters

first: Stephanie

last: O'Donoghue

first: Derek

last: Morris


first: Zhonghan

last: Li

summary: Regeneration of human cartilage and bone remains a challenge due to the limitations from current stem cell sources. Here, we developed a four-day differentiation strategy that achieved near uniform derivation (>98.5%) of SOX9+ sclerotomal progenitorsfrom multiple human pluripotent stem cells (hPSCs). Importantly, SOX9+ sclerotomal progenitors exhibited typical bipotential of mesenchymal progenitors during skeletal development. Upon lineage-specific induction, they were able to differentiate either into chondroprogenitors that could repair articular cartilage defects or early chondrocytes that could undergo endochondral ossification for human bone formation. Furthermore, we identified ITGA9 as the specific surface marker that facilitated reporter-independent isolation of SOX9+ sclerotomal progenitors and established an in vitro culture system that could expand these cells for at least 15,000-fold . Collectively, these findings highlight the SOX9+ sclerotomal progenitors a promising stem cell source for next-generation human cartilage/bone bioengineering.

overall_design: We performed scRNA sequencing to reconstruct lineage trajectories duing differentiation of iPSCs to SOX9+ sclerotomal progenitors in vitro. Cells of 4 stages including iPSC, PS, PSM, SOX9+ sclerotomal progenitors were first generated respectively before they were barcoded by cell hashing using 4 individual TotalSeq-A antibodies with different barcodes. The mixture of equal number of cells from each stages were then pooled for sc-RNA sequencing on one lane simutaneously.

first: Jingfei

last: Xiong

first: Yuansong

last: Liao

first: Yanhui

last: Deng

first: Hanyi

last: Chen

first: Fanchen

last: Kang

first: Runxin

last: Ma

first: Guogen

last: Ye

first: Kun

last: Xie

first: Haoyang

last: Cai

first: Qing

last: Zhu

first: Weihua

last: Guo


title: Experimental challenge with bovine herpesvirus 1 in dairy calves: bronchial lymph node mRNA transcriptome response

first: Dayle

last: Johnston

summary: Bovine Herpesvirus 1 (BoHV1) is a leading cause of Bovine Respiratory Disease (BRD) in young calves, which is responsible for substantial morbidity and mortality. Therefore, the objective of the current study was to elucidate the bronchial lymph node mRNA transcriptomic response to an experimental challenge with BoHV1, in dairy calves. Holstein-Friesian calves were either challenged by intranasal atomisation with BoHV1 virus (6.3 x 10^7/mL x 1.35mL) (n=12) or mock challenged with sterile phosphate buffered saline (n=6). Clinical signs were scored daily until euthanasia at day 6 post-challenge. Total RNA was extracted and sequenced from bronchial lymph node tissue (150 bp paired-end). Sequence reads were aligned to the ARS-UCD1.2 bovine reference genome and differential gene expression analysis was performed using EdgeR. An MDS plot displayed an obvious separation between BoHV1 challenged and control calves based on bronchial lymph node gene expression changes. There were 337 differentially expressed (DE) genes (p < 0.05, FDR < 0.1, fold change > 2) between the BoHV1challenged and control calves.

overall_design: Examination of a single tissue type (bronchial lymph node) in two different treatment groups, BoHV1 challenged calves (n=12) and control calves (n=6)

first: Bernadette

last: Earley

first: Matthew

last: McCabe

first: JaeWoo

last: Kim

first: Jeremy

last: Taylor

first: Ken

last: Lemon

first: Catherine

last: Duffy

first: Michael

last: McMenamy


title: Swimming induces bone loss via regulating mechanical sensing pathways in bone marrow

status: Public on Feb 20 2025

city: Shanghai

first: Shaotian

last: Fu

address: Minghang

institute: sjtu

summary: Bone is an organ capable of perceiving external mechanical stress in real time and respond dynamically via mechanosensing proteins such as Piezo1 and YAP/TAZ. Upon sensing the mechano-signals, cells within the bone matrix collaborate to coordinate bone formation and resorption, while bone marrow cells are also stimulated and mobilized. High-load exercise stimulates osteoblast differentiation and bone formation. However, the mechanism through which the low-load exercises on bone homeostasis is still unclear. In this work, we established a long-term swimming training model to unload the mechanical stress in mice. Throughout the training model, we observed a significant loss in trabecular bone mass, as evidenced by microCT scanning and histological staining. Single-cell sequencing of the tibial bone marrow tissue revealed a significant increase in the percentage of bone marrow neutrophils, along with alterations in integrins and the ERK1/2 signaling pathway. Notably, the changes in both Integrins and the ERK1/2 signaling pathway in macrophages were more pronounced than in other cell types, which not only suggests a mechanical adaptive response in these cells. Moreover, the involvement of integrins is also critical for the crosstalk between monocyte precusors and macrophages during swimming. Together, this study provide a resource of the alterations of bone marrow cell gene expression profile during swimming and highlights the importance of Integrins and the ERK1/2 signaling pathway in the bone marrow microenvironment after swimming.

overall_design: Twelve 2-month-old male C57bl/6j mice, sourced from Shanghai Ninth People's Hospital, were housed in SPF conditions with free access to food and water, under a 12-hour light/dark cycle (07:00-19:00), at 22°C and 30-70% humidity. They were divided into swimming (n=6) and non-swimming (n=6) groups. The swimming group mice starting with 10 minutes of swimming on the first day and gradually increasing to 60 minutes per day within a week. Afterward, they swam for 60 minutes every day, five times a week, for a total of 12 weeks. All swimming training sessions were conducted in the afternoon. The non-swimming group mice did not undergo swimming training. After the previous training, three mice from the swimming group and three mice from the non-swimming group were euthanized. Bone marrow was flushed out from their tibiae using PBS, respectively. After dispersing the bone marrow tissue, dead cells were removed using a Dead Cell Removal Kit (130-090-101; Miltenyi Biotec) according to the manufacturer's instructions. Single-cell RNA sequencing (scRNA-seq) analysis of the tibial bone marrow cells was then performed using the 10x Genomics Chromium platform. We designed this experiment to explore the changes in bone marrow environment induced by mechanical stress unloading during swimming, and to provide an explanation for bone loss caused by swimming.

first: Yahong

last: Lu

first: Wenkun

last: Sun

first: Wugui

first: Chengshou

last: Lin

first: An

last: Qin


title: Aboral cell types of Clytia and coral larvae have shared features and link taurine to the regulation of settlement [scRNA-Seq]

city: Villefranche-sur-mer

first: Richard

last: Copley

address: 181 chemin du lazaret

country: France

institute: CNRS UMR7009

summary: We performed single-cell RNA sequencing on the the larvae of the cnidarians Clytia hemisphaerica (Hydrozoa), Astroides calycularis (Anthozoa) and Pocillopora damicornis (Anthozoa). We identified cell types that were likely from the aboral end of the planulae and searched for genes in common that may mediate the larval settlement response.

sample_organism:
Astroides calycularis
Clytia hemisphaerica
Pocillopora acuta

overall_design: Planula larvae were dissociated and cells fixed prior to library preparation using the 10X Genomics platform, and Illumina sequencing.

first: Julia

last: Ramon-Mateu

first: Anna

last: Ferraioli

first: Evelyn

last: Houliston


title: IL-32 producing CD8+ memory T cells and Tregs define the IDO1 / PD-L1 niche in human cutaneous leishmaniasis skin lesions

city: York

first: Paul

last: Kaye

state: North Yorkshire

address: University of York, Heslington

institute: Hull York Medical School, University of York

summary: Human cutaneous leishmaniasis (CL) is characterised by chronic skin pathology. Experimental and clinical data suggest that immune checkpoints (ICs) play a crucial role in disease outcome but the cellular and molecular niches that facilitate IC expression during leishmaniasis are ill-defined. We previously showed that in Sri Lankan patients with CL, indoleamine 2,3-dioxygenase 1 (IDO1) and programmed death-ligand 1 (PD-L1) are enriched in lesion skin and that reduced PD-L1 expression early after treatment onset predicted cure rate following antimonial therapy. Here, we use spatial cell interaction mapping to identify IL-32-expressing CD8+ memory cells and regulatory T cells as key components of the IDO1 / PD-L1 niche in Sri Lankan CL patients and in patients with distinct forms of dermal leishmaniasis in Brazil and India. Furthermore, the abundance of IL-32+ cells and IL-32+CD8+ T cells at treatment onset was prognostic for rate of cure in Sri Lankan patients. This study provides a unique spatial perspective on the mechanisms underpinning IC expression during CL and a novel route to identify additional biomarkers of treatment response.

overall_design: Samples: SL2 cohort, BR cohort and IN cohort. Study design Sri Lanka The study design for the Sri Lankan cohort was initially published as part of the validation cohort details in Dey et al, 2021 32. Briefly, individuals who provided written informed consent and exhibited clinically highly suspected CL lesions were enrolled. On day 0, punch biopsies were collected and intralesional sodium stibogluconate (SSG) treatment was administered. Biopsies were processed into formalin-fixed and embedded in paraffin (FFPE) format and then shipped to the University of York (UoY) for subsequent immunological studies described here. These enrolled patients were then monitored for up to 6 months to assess complete clinical cure, and given weekly intralesional SSG injections. Of these, 24/25 patients had achieved complete clinical cure by 6 months, and 1/25 patients at 6.5 months. Out of the 25 patients in this cohort, 23/25 patient sections were included in the neighbourhood image analysis aimed at identifying neighbours of IDO1 and PD-L1 in stained sections; two were excluded due to insufficient material in the blocks. For including samples for 10x Visium analysis, these 23 patients were further screened for the presence of parasites by using an RNA probe against Amastin to detect parasites in 5u thick sections as described previously 32. 6/23 were Amastin+ and were included for Visium spatial analysis (P1-P6). Furthermore, out of the 6 samples included for Visium analysis, only four samples could be included in the CosMx analysis due to the availability of sufficient material in the blocks for follow up studies.  Brazil Patient cohort from Brazil involved 20 patients that agreed to participate by signing the informed consent Form. Patients were diagnosed on the basis of positive polymerase chain reaction and/or indirect immunofluorescence and/or ELISA directed to Leishmania. Depending on the size and location of the lesion and the clinical condition of the patients, treatment was done with systemic pentavalent antimonials (Glucantime) for 20 days; or intralesional pentavalent antimonials (Glucantime) with one application per week for 4 weeks; or liposomal amphotericin B for 10 days. Biopsy samples of the lesion were collected on day 0 when the diagnosis was established and patients were followed up after treatment for 6 months to assess clinical cure. To understand IDO1 and CD274 microenvironments in healing and non-healing chronic skin L.(V)b. lesions, 20 patient samples were screened for double positivity against diagnostic PCR and ELISA (12/20) and were then stratified on the basis of visual treatment score at 3 months; healed if visual score=0 at 3 months and not healed if visual score >0 at 3 months. Of 12 samples, only 2 patients had healing score>0 at 3 months and were included in the study. 2 patients with visual score=0 at 3 months with comparable age, lesion size and lesion duration were then selected from the bigger cohort (12) to include a total of 4 samples that were processed through 10x Visium.  India The initial diagnosis was based on clinical features suggestive of PKDL (presence of papules, nodules and/or hypopigmented macular lesions), a prior history of VL, rK-39 positivity, and/or if they resided in an area endemic for VL. In general, cases with hypopigmented patches were considered as macular PKDL, whereas cases with an assortment of papules, nodules, plaques, and/or macules were termed as polymorphic PKDL. Based on these criteria, individuals who were willing and provided written informed consent were enrolled for this study. The patients were examined by a clinician, clinical history recorded, and biological samples (blood and punch biopsy) collected. Subsequently, the diagnosis was confirmed by qPCR 91. 2 samples with typical histopathology of PKDL lesions and comparable RNA quality were chosen. These two patients were enrolled between November 2019 and August 2021. 5u thick sections were cut and processed through 10x Visium.

first: Nidhi

last: Dey

first: Shoumit

first: Naj

last: Brown

first: Sujai

last: Senarathne

first: Luiza

last: Campos Reis

first: Ritikia

last: Sengupta

first: Jose

last: Angelo L. Lindoso

first: Sally

last: James

first: Lesley

last: Gilbert

first: Mitali

last: Chatterjee

first: Hiro

last: Goto

first: Shalindra

last: Ranasinghe


title: Isogenic comparison of Airn and Xist reveals core principles of Polycomb recruitment by lncRNAs

city: Chapel Hill

first: Joseph

middle: Mauro

last: Calabrese

state: NC

address: 120 Mason Farm Road

institute: UNC Chapel Hill

summary: The mechanisms and biological roles of Polycomb repressive complex (PRC) recruitment by lncRNAs remain unclear. To gain insight, we expressed two lncRNAs that recruit PRCs to multi-megabase domains, Airn and Xist, from an ectopic locus and compared effects. Unexpectedly, ectopic Airn recruited PRC1 and PRC2 to chromatin with a potency resembling Xist yet did not repress genes. Compared to PRC2, PRC1 was more proximal to Airn and Xist, where its enrichment over C-rich elements required the RNA-binding protein HNRNPK. Fusing Airn to Repeat A, the domain required for gene silencing by Xist, enabled gene silencing and altered local patterns but not relative levels of PRC-directed modifications. Our data suggest that endogenously, Airn recruits PRCs to maintain rather than initiate gene silencing; that PRC recruitment occurs independently of Xist Repeat A; and that protein-bridged interactions, not direct RNA contacts, underlie PRC recruitment by Airn, Xist, and other lncRNAs.

overall_design:
Chromatin Immunoprecipitation DNA (ChIP-) and total RNA sequencing (RNA-Seq) in RMCE ESCs and TSCs.
RNA immunoprecipitation-sequencing (RIP-Seq) in RMCE ESCs.
Chromatin immunoprecipitation DNA-sequencing (ChIP-Seq) in RMCE ESCs.
Total RNA sequencing (RNA-Seq) in RMCE ESCs and TSCs. Nuclear, chromatin, and Cytoplasm RNA-seq from Cellular fractionation of TSCs.
RNA immunoprecipitation-sequencing (RIP-Seq) in TSCs.

first: Jackson

first: Elizabeth

last: Abrash

first: McKenzie

last: Murvin

first: Aki

middle: K

last: Braceros

first: Shuang

last: Li

first: Samuel

middle: P

last: Boyson

first: Ryan

last: Salcido

first: Rachel

middle: E

last: Cherney

first: Steven

last: Bischoff

first: Kyle

last: Kaufmann

first: Quinn

last: Eberhard

first: Zhiyue

last: Zhang

first: Dale

middle: O

last: Cowley

first: Mauro


title: Aboral cell types of Clytia and coral larvae have shared features and link taurine to the regulation of settlement [bulk_RNAseq]

first: Richard

last: Copley

summary: We performed bulk RNA sequencing on the aboral (anterior) and the oral (posterior) ends of larvae from the cnidarians Clytia hemisphaerica (Hydrozoa), Astroides calycularis (Anthozoa) , and Pocillopora acuta (Anthozoa). Our analysis identified genes enriched at the aboral end of the planulae and revealed shared genes as potential candidates to play a role in mediating the larval settlement response.

platform_organism:
Astroides calycularis
Clytia hemisphaerica
Pocillopora acuta

overall_design: Whole-tissue RNA was extracted from bisected planula larva aboral and oral ends,  and cDNA libraries were prepared for DNBseq sequencing.

first: Julia

last: Ramon-Mateu

first: Anna

last: Ferraioli

first: Evelyn

last: Houliston


title: Binding domain mutations provide insight into CTCF's relationship with chromatin and its contribution to gene regulation (RNA-seq, exit from pluripotency)

city: new york

first: Jane

middle: A

last: Skok

state: NEW YORK

address: Smilow Research Center, Third floor, Room 307, 522 First Avenue

department: Pathology

institute: NYU

summary: Here we used a series of CTCF mutations to explore CTCF’s relationship with chromatin and its contribution to gene regulation. CTCF’s impact depends on the genomic context of bound sites and the unique binding properties of WT and mutant CTCF proteins. Specifically, CTCF’s signal strength is linked to changes in accessibility, and the ability to block cohesin is linked to its binding stability. Multivariate modelling reveals that both CTCF and accessibility contribute independently to cohesin binding and insulation, however CTCF signal strength has a stronger effect. CTCF and chromatin have a bidirectional relationship such that at CTCF sites, accessibility is reduced in a cohesin-dependent, mutant specific fashion. In addition, each mutant alters TF binding and accessibility in an indirect manner, changes which impart the most influence on rewiring transcriptional networks and the cell’s ability to differentiate. Collectively, the mutant perturbations provide a rich resource for determining CTCF’s site-specific effects.

overall_design:
To study the impact of CTCF mutations on lineage commitment, we cultured cell lines wherein the mESC degron cell line was modified to express either a stable doxycycline-inducible control wild-type Ctcf or a mutant Ctcf (mCtcf) transgene in the absence of endogenous CTCF in cell media with or without LIF (leukemia inhibitory factor).
We then performed  gene expression profiling using data generated from total RNA-seq. Pluripotent mESCs expressing transgene CTCF WT or mutant (indole acetic acid, IAA, an analogue of auxin + doxycycline, ID condition) cultured in LIF-containing media were compared to mESCs undergoing exit from pluripoitency cultured in no LIF media for 3 days. Two replicates per condition were assessed.

first: Catherine

last: Do

first: Guimei

last: Jiang

first: Giulia

last: Cova

first: Christos C.

last: Katsifis

first: Domenic N.

last: Narducci

first: Theodore

last: Sakellaropoulos

first: Raphael

last: Vidal

first: Priscillia

last: Lhoumaud

first: Aristotelis

last: Tsirigos

first: Faye Fara D.

last: Regis

first: Nata

last: Kakabadze

first: Elphege P.

last: Nora

first: Marcus

last: Noyes

first: Anders S.

last: Hansen

first: Jane A.


title: Binding domain mutations provide insight into CTCF's relationship with chromatin and its contribution to gene regulation (Hi-C)

overall_design:
To study the impact of CTCF mutations, we established 10 cell lines wherein the mESC degron cell line was modified to express either a stable doxycycline-inducible control wild-type Ctcf or a mutant Ctcf (mCtcf) transgene in the absence of endogenous CTCF.
We then performed Hi-C. Cells expressing transgene CTCF mutant (indole acetic acid, IAA, an analogue of auxin + doxycycline, ID condition) were compared to cells expressing the transgene WT CTCF (WT, ID condition). Two replicates per condition were assessed.

first: Christos

first: Domenic

middle: N

first: Faye

first: Elphege

first: Anders


title: Binding domain mutations provide insight into CTCF's relationship with chromatin and its contribution to gene regulation (Omni-ATAC)

first: Jane

overall_design:
To study the impact of CTCF mutations, we established 10 cell lines wherein the mESC degron cell line was modified to express either a stable doxycycline-inducible control wild-type Ctcf or a mutant Ctcf (mCtcf) transgene in the absence of endogenous CTCF.
We then performed  ATAC-seq. Cells expressing transgene CTCF mutant (indole acetic acid, IAA, an analogue of auxin + doxycycline, ID condition) were compared to cells expressing the transgene WT CTCF (WT, ID condition). Two replicates per condition were assessed.

first: Catherine

first: Guimei

first: Giulia


title: Effect of fatty acid synthesis inhibition in IL-17-producing gamma delta T (γδT17) cells upon psoriatic inflammation

city: Mainz

first: Matthias

last: Klein

address: Langenbeckstrasse 1

department: Inst. for Immunology

country: Germany

institute: University Medical Center Mainz

summary: γδ T cells represent the primary innate source of IL-17 (γδT17) and are known to play a critical role in autoimmune and inflammatory diseases such as psoriasis. We here reported that psoriatic condition (IL-1b and IL-23) reshaped γδT17 cell metabolic signatures and promoted cytokine expression levels compared to homeostatic condition (IL-7). Acetyl-CoA carboxylase 1 (ACC1), a rate-limiting enzyme of fatty acid synthesis (FAS), directs the fate of IL-17-producing CD4 T cells (Th17) differentiation. However, little is known about the role of ACC1-mediated FAS in their innate IL-17-producer counterpart, γδT17 cells. We further investigated the role of FAS in γδT17 by comparing the effect of pharmacological inhibitor Soraphen A (SorA) on DMSO vehicle under psoriatic conditions (IL-1b and IL-23). Interestingly, ACC inhibition shifts the lipid metabolism in γδT17 cells, which allows them to cope with lipid demand without affecting cellular viability.

overall_design: CD27−γδ T cells were expanded in vitro as previously described (McKenzie et al., 2017). This protocol was adapted to have pure CD27−γδ T cells asγδT17 by sorting out expanded CD27-γδTCR+, γδT17 cells. CD27+ γδ T cells were separated and maintained by 20 ng ml–1 IL-7 as γδIFN culture. Sorted γδT17 cells were expanded for three days by 20 ng ml–1IL-7. Then, the cells were harvested and plated at 1 × 106 cells ml–1 in 96-well U-bottom plates to be stimulated with indicated cytokines (5 ng ml–1murine IL-1β/IL-23) in the presence or absence of 2000 nM Soraphen A for 3 hours.

first: Sparwasser

last: Tim

first: Yu-San

last: Kao


title: Binding domain mutations provide insight into CTCF's relationship with chromatin and its contribution to gene regulation  (RNA-seq)

last: Skok

summary: Although only a fraction of CTCF motifs are bound in any cell type, and approximately half of the occupied sites overlap cohesin, the mechanisms underlying cell-type specific attachment and ability to function as a chromatin organizer remain unknown. To investigate the relationship between CTCF and chromatin we applied a combination of imaging, structural and molecular approaches, using a series of brain and cancer associated CTCF mutations that act as CTCF perturbations. We demonstrate that binding and the functional impact of WT and mutant CTCF depend not only on the unique properties of each protein, but also on the genomic context of bound sites. Our studies also highlight the reciprocal relationship between CTCF and chromatin, demonstrating that the unique binding properties of WT and mutant proteins have a distinct impact on accessibility, TF binding, cohesin overlap, chromatin interactivity and gene expression programs, providing insight into their cancer and brain related effects.

overall_design:
To study the impact of CTCF mutations, we established 10 cell lines wherein the mESC degron cell line was modified to express either a stable doxycycline-inducible control wild-type Ctcf or a mutant Ctcf (mCtcf) transgene in the absence of endogenous CTCF.
We then performed  gene expression profiling using data generated from total RNA-seq. Cells expressing transgene CTCF mutant (indole acetic acid, IAA, an analogue of auxin + doxycycline, ID condition) were compared to cells expressing the transgene WT CTCF (WT, ID condition). To assess the rescue by CTCF WT transgene, we also performed RNA-seq in WT untreated cells. Two replicates per condition were assessed.

last: Do

first: Christos C.

first: Domenic N.

first: Jie

last: Yang

first: Theodore

first: Raphael

first: Priscillia

first: Aristotelis

first: Faye Fara D.

first: Nata

first: Xiaodong

last: Cheng


title: Examination of the pharyngeal tonsil transcriptome response to Bovine herpesvirus 1 (BoHV-1) infection in dairy calves

city: Grange, Dunsany

first: Stephanie

last: O'Donoghue

state: Meath

address: Animal Bioscience Research Centre

department: Animal & Bioscience Research Department

summary: Bovine Herpesvirus 1 (BoHV1) is a leading cause of Bovine Respiratory Disease (BRD) in young calves, which is responsible for substantial morbidity and mortality. Therefore, the objective of the current study was to elucidate the mRNA transcriptomic response in pharyngeal tonsil tissue to an experimental challenge with BoHV1, in dairy calves. Holstein-Friesian calves were either challenged by intranasal atomisation with BoHV1 virus (6.3 x 10^7/mL x 1.35mL) (n=12) or mock challenged with sterile phosphate buffered saline (n=6). Clinical signs were scored daily until euthanasia at day 6 post-challenge. Total RNA was extracted and sequenced from pharyngeal tonsil (100 bp paired-end). Sequence reads were aligned to the ARS-UCD1.2 bovine reference genome and differential gene expression analysis was performed using EdgeR. Anlysis uncovered a total of 1833 differentially expressed genes (p < 0.05, FDR < 0.1, fold change > 2) between the BoHV-1 challenged and control calves.

overall_design: Examination of a single tissue type (pharyngeal tonsil) in two different treatment groups, BoHV-1 challenged calves (n=12) and control calves (n=6)

first: Bernadette

last: Earley

first: Dayle

last: Johnston

first: Jeremy

last: Taylor

first: S

last: Cosby

first: Ken

last: Lemon

first: Derek

last: Morris

first: Sinead

last: Waters


title: Examination of the mediastinal lymph node transcriptome response to Bovine herpesvirus 1 (BoHV-1) infection in dairy calves

summary: Bovine Herpesvirus 1 (BoHV1) is a leading cause of Bovine Respiratory Disease (BRD) in young calves, which is responsible for substantial morbidity and mortality. Therefore, the objective of the current study was to elucidate the mRNA transcriptomic response in mediastinal lymph node tissue to an experimental challenge with BoHV1, in dairy calves. Holstein-Friesian calves were either challenged by intranasal atomisation with BoHV1 virus (6.3 x 10^7/mL x 1.35mL) (n=12) or mock challenged with sterile phosphate buffered saline (n=6). Clinical signs were scored daily until euthanasia at day 6 post-challenge. Total RNA was extracted and sequenced from mediastinal lymph node tissue (100 bp paired-end). Sequence reads were aligned to the ARS-UCD1.2 bovine reference genome and differential gene expression analysis was performed using EdgeR. Anlysis uncovered a total of 81 differentially expressed genes (p < 0.05, FDR < 0.1, fold change > 2) between the BoHV-1 challenged and control calves.

overall_design: Examination of a single tissue type (mediastinal lymph node) in two different treatment groups, BoHV-1 challenged calves (n=12) and control calves (n=6)


title: Sequencing of microglia and monocytes during focal remyelination of the corpus callosum

city: Edinburgh

first: Veronique

last: Miron

address: 49 Little France Crescent

department: UK DRI

institute: The University of Edinburgh

summary: The goal of the study was to determine the gene expression of monocytes vs microglia during remyelination.

overall_design: 10 week-old male C57Bl/6J mice were anaesthetized with isoflurane before stereotaxic injection of 2 μl of 1% lysolecithin (LPC; vol/vol) into the corpus callosum and sacrificed 10 days later.

first: Claire

last: Davies

first: Stefka

last: Gyoneva

first: Richard

last: Ransohoff

first: Brian

last: Wipke


title: Krüppel-like Factor family members control expression of genes required for serous cavity and alveolar macrophage identities [CUT&Run]

status: Public on Feb 19 2025

city: Berkeley

first: Gregory

last: Barton

department: Molecular and Cell Biology

institute: UC Berkeley

summary: Tissue-resident macrophages adopt distinct gene expression profiles and exhibit functional specialization based on their tissue of residence. Recent studies have begun to define the signals and transcription factors that induce these identities. Here we describe an unexpected and specific role for the broadly expressed transcription factor Kruppel-like Factor 2 (KLF2) in the development of embryonically derived Large Cavity Macrophages (LCM) in the serous cavities. KLF2 not only directly regulates the transcription of genes previously shown to specify LCM identity, such as retinoic acid receptors and GATA6, but also is required for induction of many other transcripts that define the identity of these cells. We identify a similar role for KLF4 in regulating the identity of alveolar macrophages in the lung. These data demonstrate that broadly expressed transcription factors, such as Group 2 KLFs, can play important roles in the specification of distinct identities of tissue-resident macrophages.

overall_design: CUT&RUN sequencing of anti-GFP vs IgG of sorted large peritoneal cavity macrophages from GFP-KLF2 fusion knock-in mice. Anti-GFP and IgG are paired across three replicates from individual mice.

first: Kathleen

last: Pestal

first: Leianna

last: Slayden


title: An inducible mouse model of OI type V reveals aberrant osteogenesis caused by Ifitm5 c.-14C>T mutation

city: Shenzhen

first: Zhijia

last: Tan

address: 1st Haiyuan Road, Futian district

department: Orthopaedic and Traumatology

institute: The University of Hong Kong-Shenzhen Hospital

summary: Osteogenesis imperfecta (OI) Type V is typically characterized by radial head dislocation, calcification of interosseous membrane and post-fracture hyperplastic callus. It is caused by the c.-14C>T mutation in the 5’UTR of the IFITM5 gene, adding five amino acids (MALEP) to the N-terminal of IFITM5 protein. Previous studies have suggested a neomorphic function of the MALEP-IFITM5 protein. However, the underlying mechanisms remain unclear due to embryonic lethality in previous models. Therefore, we developed an inducible animal model of Ifitm5flox c.-14C>T that could be induced by Cre expressing at different developmental stages to explore the pathogenic effects of the neomorphic MALEP-IFITM5. Specifically, Prx1-Cre; Ifitm5flox c.-14C>T mutant mice were born with fractures in all limbs, showing deficient ossification and enhanced chondrogenesis associated with increased SOX9 abundance. We isolated skeletal cells from tibia of Ifitm5 mutant mouse at P6 stage and conducted single RNA sequencing (10X Genomics). The data were compared to the RNA-seq data generated from the tibia of control mouse at the same stage (GSE159544).

overall_design: The protocol to isolate the mutant skeletal cells for single cell RNA sequencing was the same as the control sample (GSE159544). Right and left tibia from Prx1-Cre; Ifitm5flox c.-14C>T mutant mouse at P6 stage were dissected with soft tissues removed. Bone tissue was cut into small pieces and blood cells were removed with TrypLE Express digession for 10 minutes at 37 degree. The remaining bone tissue was digested with type II collagenase (0.25%) and Dispase (0.25%) in HBSS for 1.5 hours. Cells were collected every half an hour, and digestion medium was changed each time. Red blood cells were lysised and dead cells were removed using kits. The remaining cells were tested for cell viability (>80%). Single cells were encapsulated for library preparation and sequencing using the Chromium single cell platform (10X Genomics Inc)according to the manufacturer’s protocol. Library size and concentration were determined by Qubit, quantitative PCR, and Bioanalyzer assays. A raw total input of 15000 cells was estimated for each sample. Library preparation and Illumina (NovaSeq 6000) sequencing (151 bp paired end) were performed at HKU CPOS, at 150 Gbp throughput per sample.

first: Hiu

middle: T

last: Shek

first: Zeluan

last: Li


title: Binding domain mutations provide insight into CTCF's relationship with chromatin and its contribution to gene regulation  (chip-seq)

first: Jane

last: Skok

overall_design:
To study the impact of CTCF mutations, we established 10 cell lines wherein the mESC degron cell line was modified to express either a stable doxycycline-inducible control wild-type Ctcf or a mutant Ctcf (mCtcf) transgene in the absence of endogenous CTCF.
We then performed  chipmentation  for cohesin (SMC3) and for transgene CTCF (FLAG). Cells expressing transgene CTCF mutant (indole acetic acid, IAA, an analogue of auxin + doxycycline, ID condition) were compared to cells expressing the transgene WT CTCF (WT, ID condition). To assess the rescue by the transgene WT CTCF, we performed chipmentation using CTCF antibody in WT untreated and WT ID conditions. Two replicates per condition were assessed.

first: Catherine

last: Do

first: Guimei

last: Jiang

first: Giulia

last: Cova

first: Christos C.

last: Katsifis

first: Domenic N.

last: Narducci

first: Jie

last: Yang

first: Theodore

last: Sakellaropoulos

first: Raphael

last: Vidal

first: Priscillia

last: Lhoumaud

first: Aristotelis

last: Tsirigos

first: Faye Fara D.

last: Regis

first: Nata

last: Kakabadze

first: Elphege P.

last: Nora

first: Marcus

last: Noyes

first: Xiaodong

last: Cheng

first: Anders S.

last: Hansen

first: Jane A.


title: Adaptive evolution of KoRV-A  retroviral silencing in wild koalas [small RNA-seq]

first: Tianxiong

last: Yu

address: 364 Plantation St

institute: Umass Med

type: Non-coding RNA profiling by high throughput sequencing

summary: Koala Retrovirus-A is spreading through wild koalas in a north to south wave while transducing the germline, generating heritable genetic modifications as it transitions to an endogenous retrovirus.  In animals south of Brisbane, we previously found that KoRV-A is expressed in the germline, but the resulting unspliced genomic transcripts are processed into sense strand piRNAs, post-transcriptionally suppressing replication.   Here we show that this initial “innate” response is prevalent in animals south of the Brisbane River, but KoRV-A promoters are methylated and the virus is transcriptionally silenced in a subpopulation of animals north of the river.  Transcriptional silencing correlates with capture of an anti-sense KoRV-A provirus by the 3’UTR of the MAP4K4 gene.  The resulting hybrid transcripts are processed into anti-sense piRNAs, which transcriptionally silence established endogenous retroviruses in the germline.  This provirus is sweeping through northern koalas, supporting a direct role in adaptive evolution of KoRV-A transcriptional silencing.

overall_design: To investigate the evolution of retroviral repression in the germline, we performed long- and short-read DNA-seq, RNA-seq, and small RNA-seq.

first: Milky

last: Abojorga

first: Birgit

last: Koppetsch

first: Michaela

last: Blyton

first: Keith

last: Chappell

first: Jeremy

last: Luban

first: Zhiping

last: Weng

first: William

last: Theurkauf


title: Krüppel-like Factor family members control expression of genes required for serous cavity and alveolar macrophage identities [RNA-seq]

first: Gregory

last: Barton

overall_design:
RNA-Seq of KLF2flox or LysMCre/+ KLF2flox BMMs before or after 7 days in the peritoneal cavity.
RNA-Seq of retrovirally transduced BMMs overexpressing empty vector (EV), KLF2, or GATA6. RNA-Seq of KLF4flox or LysMCre/+ KLF4flox sorted alveolar macrophages.
RNA-Seq of KLF4flox or LysMCre/+ KLF4flox sorted alveolar macrophages.

first: Kathleen

last: Pestal

first: Leianna

last: Slayden


title: P0 kidney samples

city: Cincinnati

first: Mike

last: Adam

state: Ohio

address: 240 Albert Sabin Way

department: CBDI

institute: Cincinnati Children's Hospital Research Foundation

summary: The goal of our study is to analyze formation/development of nephron segments.

overall_design: Mouse kidneys at P0 were collected and subjected to scRNA-seq analysis.

first: Joo-seop

last: Park


title: Mapping GATA2 Occupancy in Uterine Serous Carcinoma

city: Madison

first: Daniel

last: Matson

address: 1111 Highland Avenue

institute: University of Wisconsin

summary: GATA2 expression predicts uterine serous carcinoma (USC) behavior and patient outcome. To identify putative GATA2 target genes, we performed anti-GATA2 ChIP-seq in Ark1 USC cells.

overall_design: Chromatin immunoprecipitation DNA sequencing (ChIP-seq) for GATA2 or IgG control in Ark1 USC cells.

first: Apoorva

last: Patil

first: Usha

last: Polaki

first: Peng

last: Liu


title: The effects of spermidine supplementation on transcriptome profiles in aging Drosophila brains

city: Berlin

first: Stephan

middle: J

last: Sigrist

state: Berlin

address: Takustr. 6, 2OG, AG Sigrist

department: Biology, Chemistry and Pharmacy,

institute: Freie Universität Berlin

summary: We report the trancriptome comparison of 15-day-old Drosophia brains fed with and without spermidine

overall_design: Transcriptome profiles in aging wild type Drosophila brains with and without spermidine supplementation

first: YongTian

last: Liang


title: An ISWI-related chromatin remodeler orchestrates parasite lifecycle progression by insulating gene expression in a densely packed genome [ChIP-seq]

city: La Tronche

first: Christopher

last: Swale

address: Batiment Jean Roger

department: U1209 - Institute of Advanced Biosciences

institute: INSERM

summary: In eukaryotic cells, ATP-dependent chromatin remodelers are specialized multiprotein machines that organize the genome and rule its accessibility by repositioning, ejecting or modifying nucleosomes, but their role in Toxoplasma gondii is still poorly understood. Phylogenetic analyzes show that the parasite retains the four commonly recognized remodeler families and has evolved two divergent proteins within the ISWI family: TgSNF2h and TgSNF2L. These proteins are structurally divergent and form distinct complexes. TgSNF2h specifically forms a core complex with the transcription factor AP2VIII-2 and the scaffold protein TgRFTS. Depletion of TgRFTS phenocopies the knockdown of TgSNF2h, restricting access to chromatin and altering local gene expression. At the genomic level, TgSNF2h actively insulates highly transcribed genes from their minimally expressed or silenced neighbors. This ISWI complex establishes and maintains stage-specific gene expression by regulating the chromatin accessibility to transcription factors. Using MORC as a proxy, we have shown that TgSNF2h rules DNA accessibility and thereby exerts an epistatic control over a key regulator of sexual commitment. In a parasite characterized by high phenotypic plasticity and a compact genome, this dedicated ISWI complex orchestrates the partitioning of developmental genes and ensures transcriptional fidelity throughout its life cycle.

overall_design: We report genome-wide SNF2h and MORC, profiles in Toxoplasma gondii using ChIP-seq in the context of SNF2h protein knock-down

first: Belen

last: Pachano

first: Martina

last: Shahinas

first: Jill

last: von Velsen

first: Charlotte

last: Corrao

first: Lucid

last: Belmudes

first: Caroline

last: Mas

first: Yohann

last: Couté

first: Matthew

middle: W

last: Bowler

first: Alexandre

last: Bougdour

first: Mohamed-Ali

last: Hakimi


title: An ISWI-related chromatin remodeler orchestrates parasite lifecycle progression by insulating gene expression in a densely packed genome [ATAC-seq]

overall_design: We report Tn5 transposase accessiblity data (ATAC-Sequencing) on the RHK80 SNF2h KD, RHK80 RFTS KD, RHK80 SNF2L KD  and RHK80  MORC KD strains in an untreated condition (UT) and induced knockdown (IAA 24h)


title: Leukemogenic Kras mutation reprograms multipotent progenitors to facilitate its spread through the hematopoietic system

city: New York City

first: Boris

last: Reizis

state: New York

address: 550 1st Ave

institute: NYU Grossman School of Medicine

summary: We studied how leukemia-associated mutations subvert normal adult hematopoiesis by genetically inducing them in specifically in murine hematopoietic stem cells (HSC) followed by lineage tracing in native hosts. Leukemia-initiating mutations such as Tet2 or Dnmt3a deletions did not increase the net HSC contribution in the steady state, whereasTet2 deletion slightly accelerated HSC contribution to myeloid lineages following microbial stimulation. In contrast, the leukemia-transforming KrasG12D mutation dramatically accelerated the contribution of mutant HSC to all hematopoietic lineages, rapidly creating a pre-leukemic state. The acceleration was mediated by KrasG12D-carrying multipotent progenitors (MPP) that rapidly outcompeted normal MPP and showed increased proliferation but lacked self-renewal capacity. In addition, mutant progenitor cells expressed the secreted stem/progenitor regulator osteopontin and showed enhanced CXCR4-dependent motility, both of which contributed to the accelerated spread of the mutation to mature cells. Thus, transforming mutations may facilitate their spread from HSC to mature cells via hypercompetitive progenitors, creating a two-component stem/progenitor circuit that underlies leukemogenesis.

overall_design: Pdzk1ip1-CreER R26Tom or Pdzk1ip1-CreER R26Tom KrasG12D mice were treated with 0.5mg of tamoxifen. After 3 weeks post-induction, bone marrow cells were stained with hashtags FACS antibodies. For KrasG12D group, Tom+ Lin- Sca-1+ c-Kit+ (LSK) cells were sorted and for WT group, total LSK cells were sorted.

first: Geunhyo

last: Jang

first: Eduardo

last: Esteva


title: An ISWI-related chromatin remodeler orchestrates parasite lifecycle progression by insulating gene expression in a densely packed genome [RNA-seq]

overall_design: We report transcriptome of RH Ku80 SNF2h KD and  RH Ku80 RFTS KD in Toxoplasma gondii tachyzoites left untreated or after 24 h of IAA treatment using Nanopore Direct RNA sequencing and Illumina RNA sequencing.


title: Regulatory T cells are essential for controlling autoimmune lacrimal gland pathogenic cells

city: Houston

first: Cintia

middle: Sade

last: de Paiva

state: Texas

address: One Baylor Plaza

department: Ophthalmology

institute: Baylor College of Medicine

summary: Purpose: CD25KO mice have severe autoimmune disease and are used as a model of Sjögren disease, a prototype autoimmune dry eye. CD25KO mice have severe inflammation and infiltrating lymphocytes to multiple self-tissues, including the exocrine glands. We investigated the immunological profile of CD25KO mice and compared it to wild-type (WT) mice. Methods: 8-week-old WT and CD25KO mice were used. LGs (n=5) were submitted for RNA bulk sequencing. Tears washes were collected from mice and analyzed for protein expression using a luminex assay. A time course of adoptive transfer of CD25KO CD4+T cells was performed and LGs were evaluated by histology and qPCR. Co-adoptive transfer of CD25KO CD4+T cells with either 2M or 20M B6 CD25+GITR+T regulatory cells (Tregs) was performed in RAG1KO mice. LGs were evaluated in H&E-stained sections and by qPCR. Results: Investigation by RNA bulk sequencing found 3481 genes differentially expressed in CD25KO mice compared to B6, including upregulation of genes involved in T cell activation and Type I interferon signaling. CD25KO mice produced the same volume of tears as wild-type littermate controls, but had elevated protein expression of TNF, IFN-γ, and CCL5 and decreased protein expression of IL-12p40 and VEGF-A. Adoptive transfer recipients begin to develop inflammation and disease three weeks after adoptive transfer. In mice that received adoptive transfer of CD25KO CD4+ T cells, LGs had inflammation scores significantly higher than naive mice. They had increased expression of transcripts for inflammatory cytokines and chemokines. However, in recipient mice that received CD25KO CD4+T cells + 2M B6 Tregs, LGs had significantly reduced inflammatory scores and reduced expression of some inflammatory cytokines. Recipients of CD25KO CD4+T cells + 20M B6 Tregs had more inflamed lacrimal glands by both histology and qPCR, suggesting aged Tregs have less suppressive capacity.Conclusions: CD25KO mice have phenotypic and genetic changes resulting in increased inflammation, systemic autoimmunity and severe lymphocytic infiltration to the exocrine glands. These results are prominent in the tear film and LG. However, this autoimmunity can be controlled by addition of Tregs from healthy young mice, suggesting a possible avenue for therapeutic research.

overall_design: To explore differences between wild-type littermates and CD25KO mice's lacrimal glands, we excised whole glands from both cohorts of mice at 8 weeks of age. All mice were female, because SjD affects more women than men at a ratio of 9:1. Glands were homogenized and RNA extracted from lysates for submission for bulk RNA sequencing.

first: Kaitlin

last: Scholand

first: Laura

last: Schaefer

first: Gowthaman

last: Govindarajan

first: Zhiyuan

last: Yu

first: Jeremias

last: Galletti


title: Piezo2 in sensory neurons regulates systemic and adipose tissue metabolism

city: Rehovot

first: FABIAN

middle: Simone

last: PASSINI

address: Herzl Street 234

country: Israel

institute: Weizmann Institute

summary: Systemic metabolism ensures energy homeostasis through inter-organ crosstalk regulating thermogenic adipose tissue. Unlike the well-described inductive role of the sympathetic system, the inhibitory signal ensuring energy preservation remains poorly understood. Here, we show that, via the mechanosensor Piezo2, sensory neurons regulate morphological and physiological properties of brown and beige fat and prevent systemic hypermetabolism. Targeting runt-related transcription factor 3 (Runx3)/parvalbumin (PV) sensory neurons in independent genetic mouse models resulted in a systemic metabolic phenotype characterized by reduced body fat and increased insulin sensitivity and glucose tolerance. Deletion of Piezo2 in PV sensory neurons reproduced the phenotype, protected against high-fat-diet-induced obesity, and caused adipose tissue browning and beiging, likely driven by elevated norepinephrine levels. Finding that brown and beige fat are innervated by Runx3/PV sensory neurons expressing Piezo2 suggests a model in which mechanical signals, sensed by Piezo2 in sensory neurons, protect energy storage and prevent a systemic hypermetabolic phenotype.

overall_design: The RNA seq experiment was performed to assess the difference in muscle transcriptome between control and Piezo2 cKO mice. Thus, we collected the soleus and the gastrocnemius muscles from control and Piezo2cKO mice and extracted RNA from these samples.

first: Fabian

last: Passini

first: Elazar

last: Zelzer

first: Noa

last: Wigoda


title: Single cell RNA-seq analysis of the Rbpj mutant mouse kidney by Foxd1Cre

first: Mike

last: Adam

summary: The goal of our study is to determine if interstitial Notch signaling is required for the formation/development of nephron segments.

overall_design: Rbpj flox allele was deleted in the mouse renal interstitium using Foxd1Cre. Mouse kidneys at P0 were collected and subjected to scRNA-seq analysis. Examination of four groups: two controls (Rbpj c/+ ; Foxd1cre/+) and two mutants (Rbpj c/- ; Foxd1cre/+)

first: Joo-seop

last: Park


title: mTORC1 cooperates with tRNA wobble modification to sustain the protein synthesis machinery

city: Heidelberg

first: Andres

last: Elia

state: Baden-Württemberg

address: Im Neuenheimer Feld 223

institute: DKFZ

summary: Synthesizing the cellular proteome is a demanding process that is regulated by numerous signaling pathways and RNA modifications. How these orchestrate the protein synthesis machinery to generate specific proteome subsets remains unclear. We found when mTORC1 was inactive, tRNA wobble uridine-modifying enzymes (U34-enzymes) Elongator and Ctu1/2 became essential for cell growth in vitro and in tumors. Using ribosome profiling, RNA seq and other methods, we interrogated the functional interplay between U34-enzymes and mTORC1.

overall_design:
Ribosome Profiling and RNAseq from EPP2 murine pancreatic cancer cell lines, with induced KO of Ctu1, were carried out to better understand the role of RNA modifications and mTORC1 in codon-specific translation.
For this EPP2 cells were treated with 300 nM Torin1 or DMSO for 2 hours. For every replicate, RiboSeq and RNAseq was performed from the same sample.

first: Julia

last: Hermann

first: Alexander

last: Kowar

first: Fabricio

last: Loayza-Puch

first: Wilhelm

last: Palm


title: Single nucleus RNAseq of aldosterone-induced left atrial myopathy

status: Public on Feb 18 2025

city: Freiburg

first: Achim

last: Lother

state: --- Select One ---

address: Albertstr. 25

department: Institute of experimental and clinical Pharmacology and Toxicology,

institute: University of Freiburg

summary: Atrial fibrillation is associated with stuctural remodelling of the atria that involves various cell types, including cardiac myocytes, endothelial cells, and immune cells. Atrial myopathy forms the substrate for an increased risk for AF onset and on the other hand AF drives atrial myopathy. Atrial myopathy is linked to risk factors such as aging, hypertension, obesity, or heart failure. Aldosterone and the mineralocorticoid receptor are drivers of pathological remodeling in atrial myopathy. In this study, we investigated the effect of aldosterone on left atrial gene expression and cell-cell communication.

overall_design: single nucleus RNAseq of mouse left atrial tissue after aldosterone infusion or untreated control


title: KRAS mutants confer platinum resistance by regulating ALKBH5 posttranslational modifications in lung cancer

city: Milwaukee

first: Tongjun

last: Gu

address: 8727 W Watertown Plank Rd

institute: VERSITI WISCONSIN, INC.

type: Methylation profiling by high throughput sequencing

summary: Constitutively active mutations of KRAS frequently occur in non-small cell lung adenocarcinoma cells (NSCLC). However, the relationship between KRAS constitutive mutations and platinum drug resistance in NSCLC, as well as the underlying mechanism, remains unclear. In this study, we have identified that KRAS constitutive mutations confer resistance to platinum-based drugs in NSCLC. Mechanistically, KRAS mediates platinum resistance in NSCLC cells by enhancing ERK/JNK signaling, which controls post-translational modifications (PTMs) of ALKBH5. This, in turn, leads to an upregulation of m6A methylation in mRNA transcripts related to DNA repair genes. Consequently, the increased m6A methylation of these transcripts results in their mRNA stabilization, thereby protecting NSCLC cells from platinum-induced DNA damage and apoptosis, ultimately conferring platinum drug resistance in NSCLC. Furthermore, our observations indicate that blocking the KRAS mutation-induced increase in m6A methylation of DNA repair-related genes can sensitize NSCLC cells harboring KRAS mutations to platinum exposure. This can be achieved either by overexpressing a SUMOylation-deficient mutant of ALKBH5 or by pharmacologically inhibiting METTL3 enzymatic activity using a small molecule. These interventions significantly enhance the sensitivity of KRAS mutant-bearing NSCLC cells to platinum-based treatments both in vitro and in vivo. In summary, our study uncovers a previously unrecognized mechanism involving KRAS mutations, ALKBH5 PTMs, and increased mRNA m6A levels in DNA repair-related genes, which collectively promote platinum resistance in NSCLC cells.

overall_design: RNA-seq and m6A-seq were performed in NCI-H522 cells stably expressing empty vector and KRAS G12V

first: Fang

first: Shikan

last: Zheng

first: Zhijian

last: Qian


title: Epitranscriptomic profiling of m6A RNA modifications in THP-1 monocytic cell line.

city: Guangzhou

first: Yin

last: Zhang

address: No. 135, Xingang Xi Road

institute: SYSU

summary: Allele-specific regulatory events are essential in many fundamental biological processes. In this study, we utilized the human monocytic cell line THP-1 and performed RNA immunoprecipitation sequencing (MeRIP-seq) to explore allele-specific regulation of N6-methyladenosine (m6A) RNA modifications. Recent research highlights the importance of allele-specific regulation of m6A (ASm6A) and the need for precise detection methods to elucidate its underlying mechanisms. To address limitations in current approaches, which often emphasize genetic variations rather than comprehensive regulatory mechanisms, we applied M6Allele, a meta-analysis framework based on hierarchical Bayesian models. This method enables accurate detection of ASm6A events at the peak level, providing new insights into allele-specific regulatory mechanisms of m6A modifications. Our findings from the THP-1 cell line contribute to a deeper understanding of ASm6A and its biological significance.

overall_design: MeRIP-seq of wild-type THP-1 cells, including input and m6A-immunoprecipitated (IP) RNA samples, without replicates.


title: Binding domain mutations provide insight into CTCF's relationship with chromatin and its contribution to gene regulation

city: new york

first: Jane

last: Skok

address: Smilow Research Center, Third floor, Room 307, 522 First Avenue

institute: NYU

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

overall_design: Refer to individual Series


title: Interleukin-4 modulates type I interferon to augment anti-tumour immunity

city: Perth

first: Jesse

middle: David

last: Armitage

state: WA

address: 35 Stirling Hwy, Crawley

department: School of Biomedical Sciences

country: Australia

institute: University of Western Australia

summary: Despite advances in immunotherapy, metastatic melanoma remains a significant therapeutic challenge due to the complexity of the tumour microenvironment. Intratumoural type I interferon (IFN-I) has long been associated with improved clinical outcomes. However, several IFN-I subtypes can also paradoxically promote tumour growth in some contexts. We investigated this further by engineering murine B16 melanoma cells to overexpress various IFN-I subtypes, where a spectrum of outcomes was observed. Characterisation of these tumours by RNA-sequencing revealed a tumour immune phenotype, where potent IFN-I signalling concomitant with diminished type 2 inflammation failed to confer durable tumour control. T cell-mediated rejection of these tumours was restored by introducing interleukin (IL)-4 into the tumour microenvironment, either through ectopic expression or in a pre-clinical adoptive T cell therapy model. Collectively, our findings highlight the IFN-I/IL-4 axis in promoting anti-tumour immunity, which could be harnessed to target and stratify solid tumours that are non-responsive to frontline therapies.

overall_design: Bulk RNA-seq profiling of whole melanoma B16-GFP, B16-IFNA4 and B16-IFNA9 tumours harvested 8 days post-engraftment (n=10 per group).

middle: D


title: Interleukin-4 modulates type I interferon to augment anti-tumour immunity [scRNA-Seq]

overall_design: Single cell RNA-seq profiling of whole melanoma B16-GFP, B16-IFNA4 and B16-IFNA9 tumours harvested 8 days post-engraftment (n=3 mice per group, 4 replicates per mouse).


title: Immunotherapy-related cognitive impairment after CAR T cell therapy in mice

city: Palo Alto

first: Michelle

last: Monje

state: California

address: 265 Campus Dr.

institute: Stanford University

summary: Persistent central nervous system (CNS) immune dysregulation and consequent dysfunction of multiple neural cell types is central to the neurobiological underpinnings of a cognitive impairment syndrome, colloquially referred to as “brain fog”, that can occur following traditional cancer therapies or certain infections. Immunotherapies have revolutionized cancer care for many tumor types, but the potential long-term cognitive sequelae are incompletely understood. Here, we demonstrate in mouse models that chimeric antigen receptor (CAR) T-cell therapy for both CNS and non-CNS cancers can impair cognitive function and induce a persistent CNS immune response characterized by white matter microglial reactivity, microglial chemokine expression, and elevated cerebrospinal fluid (CSF) cytokines and chemokines. Consequently, oligodendroglial homeostasis and hippocampal neurogenesis are disrupted. Single nucleus sequencing studies of human frontal cortex and subcortical white matter from brain tumor patients with or without previous CAR T-cell therapy confirm reactive states of microglia and oligodendrocytes in patients treated with CAR T cell therapy. In mice, transient microglial depletion or CCR3 chemokine receptor blockade rescues oligodendroglial deficits and cognitive performance in a behavioral test of attention and short-term memory function following CAR T-cell therapy. Taken together, these findings illustrate similar mechanisms underlying immunotherapy-related cognitive impairment (IRCI) and cognitive impairment following traditional cancer therapies and other immune challenges.

overall_design: Bulk RNA seq was performed on mouse microglia extracted from full brains of mice that either received tumor clearing CAR T treatment or mock treated controls. 5 mice per treatment, analyzed one month after CAR T cell therapy

first: Geraghty

last: Anna

first: Mancusi

last: Rebecca

first: Truvedi

last: Vrunda


title: Roles of P-body factors in Candida albicans filamentation and stress response

city: Pittsburgh

first: Charles

middle: Joel

last: McManus

state: Pennsylvania

address: 4400 Fifth Ave

department: Biological Sciences

institute: Carnegie Mellon University

summary: Hyphal growth is strongly associated with virulence in the human fungal pathogen Candida albicans. While hyphal transcriptional networks have been the subject of intense study, relatively little is known about post-transcriptional regulation. Previous work reported that P-Body (PB) factors Dhh1 and Edc3 were required for C. albicans virulence and filamentation, suggesting an essential role for post-transcriptional regulation of these processes. However, the molecular roles of these factors have not been determined. To further study the function of PB factors in filamentation, we generated homozygous deletions of DHH1 and EDC3 in diverse prototrophic clinical strains using transient CRISPR-Cas9. Homozygous DHH1 deletion strongly impaired growth, altered filamentation, and exhibited unusual colony morphology in response to heat stress in five strain backgrounds. Using RNA-seq, we found DHH1 deletion disrupts the regulation of thousands of genes under both yeast and hyphal growth conditions in SC5314 and P57055. This included upregulation of many stress response genes in the absence of external stress, similar to deletion of the S. cerevisiae DHH1 homolog. In contrast, we found EDC3 was not required for heat tolerance or filamentation in diverse strains. These results support a model in which DHH1, but not EDC3, represses hyphal stress response transcripts in yeast and remodels the transcriptome during filamentation. Our work supports distinct requirements for specific mRNA decay factors, bolstering evidence for post-transcriptional regulation of filamentation in C. albicans.

overall_design: RNA-seq was used to compare gene expression in yeast (YPD, 30C) and hyphae (RPMI + serum, 37C) from wildtype and Dhh1-deletion strains of C. albicans. Both SC5314 and P57055 wildtype and deletion strains were used in this study.

first: Melissa

last: Tosiano


title: Flow-cytometric quantification of urine kidney epithelial cells specifically reflects tubular damage in acute kidney diseases

first: Jan

last: Klocke

address: Charitéplatz 1

department: Nephrology and Intesive Care

institute: Charité Universitätsmedizin Berlin

summary: Despite tubular injury being one of the main mechanisms driving acute kidney injury (AKI), clinicians still have a limited diagnostic repertoire to precisely monitor damage to tubular epithelial cells (TEC). In our previous work we used single cell sequencing to identify TEC subsets as main component of the urine signature in AKI. The aim of this study was to establish TEC as a clinical marker for tubular damage. In total 243  patients were analyzed. For sequencing, we collected eight urine samples of  patients with AKI and glomerular disease. By aligning urinary single-cell transcriptomes and TEC-surface proteins using Cellular Indexing of Transcriptome and Epitope Sequencing (CITE-Seq), we developed a protocol for flow cytometric quantification of CD10/CD13+ proximal and CD227/CD326+ distal TEC in urine. The marker combinations were confirmed in kidney biopsies. We validated our approach across four cohorts totaling 235  patients, consisting of patients with AKI (n=63), SARS-CoV-2 infection (n=47), ANCA-associated vasculitis (AAV) with active disease and stable remission (n=110) and healthy controls (n=15). Our findings demonstrate that CD10/CD13 and CD227/CD326 adequately identify proximal and distal urinary TEC, respectively. Distal urinary TEC counts correlate with the severity of AKI based on KDIGO stage and acute  eGFR loss in two separate cohorts and can successfully discriminate AKI from healthy controls as well as glomerular disease. We propose urinary CD227/CD326+ TEC counts as a specific, non-invasive marker for tubular injury in AKI. Our protocol provides the basis for a deeper phenotypic analysis of urinary TECs

overall_design: Single-cell RNAseq of flow cytometry sorted viable cells from 8  human urine specimen of patients with acute kidney injury (n=6), membranous nephropathy  (n=1) and active lupus nephritis class IV (n=1)

first: Leonie

last: Wagner

first: Christopher

last: Skopnik

first: Philipp

last: Enghard


title: Immunotherapy-related cognitive impairment after CAR T cell therapy in mice [scRNA-seq]

overall_design: Single cell nucleus sequencing was performed on cortex autopsy samples from patients with DIPG that either received CAR T cell therapy or did not.

first: Anna

last: Geraghty

first: Michael

last: O'Dea

first: Lehi

last: Acosta-Alvarez

first: Daniel

last: Zamler

first: Kiarash

last: Shamardani


title: Loss of Socs2 improves molecular responses to IFNα in a mouse model of myeloproliferative neoplasms driven by JAK2-V617F

city: Basel

first: DBM Bioinformatics

last: Core Facility

state: BS

address: Hebelstrasse 20

department: Departement of Biomedicine

country: Switzerland

institute: University of Basel

summary: Therapy with pegylated interferon alpha (pegIFNα) can induce a deep molecular response in a subset of patients with myeloproliferative neoplasms (MPN). Here we investigated the role of Socs2, a negative regulator of cytokine signaling, in modulating the response to pegIFNα in a JAK2-V617F mouse model of MPN. Deleting Socs2 in JAK2-V617F mice resulted in increased sensitivity to cytokines, without causing significant alterations in the MPN phenotype. When subjected to pegIFNα, the loss of Socs2 enhanced the depletion of JAK2-mutant hematopoietic stem cells (HSCs), evidenced by reduced chimerism in peripheral blood and bone marrow compared to vehicle controls. Additionally, pegIFNα-treated Socs2-deficient JAK2-mutant HSCs exhibited functional impairments in secondary transplantations, reflecting long-term detrimental decline of their stemness. These findings demonstrate that loss of Socs2 enhances the effectiveness of pegIFNα in depleting the JAK2-mutant HSC clone. In line with the genetic ablation of Socs2, the SOCS2 inhibitor MN714 combined with IFNα exhibited better efficacy than IFNα alone in reducing the output of CD34+ cells from PV patients in vitro. Targeting SOCS2 could therefore improve therapeutic responsiveness in MPN patients receiving interferon therapy.

overall_design: We analyzed bone marrow c-kit+ cells from tamoxifen-inducible conditional JAK2-V617F transgenic mice (Scl;VF) and wild-type mice (WT). Individual mice were multiplexed using a hashtag oligonucleotide labeling strategy (TotalSeq-B). Sample demultiplexing can be performed using the information provided in the processed data file HTO_full_info.txt.

first: Marc

last: Usart

first: Radek

last: Skoda

first: Julien

last: Roux

first: Athimed

last: El Taher


title: Distinct CD8+ T cell dynamics associate with response to neoadjuvant cancer immunotherapies

status: Public on Feb 24 2025

city: Chapel Hill

first: Robert

last: Ferris

address: 450 West Drive

institute: UNC Lineberger Comprehensive Cancer Center

summary: We leverage a clinical trial (NCT04080804) that compared neoadjuvant anti-PD-1, anti-PD-1+CTLA-4, and anti-PD-1+LAG-3 therapies in head and neck squamous cell carcinoma patients. Combination therapies promote higher pathologic response rates versus monotherapy, and major pathologic response is associated with better survival. To address whether successful immune checkpoint inhibitor (ICI) regimens act through similar or distinct pathways, we robustly and longitudinally characterize transcriptional and proteomic dynamics of CD8+ tumor-infiltrating lymphocytes (TILs) in a clonal manner. Anti-PD-1+LAG-3 reprograms CD8+ TIL with type-I interferon response and exhaustion gene programs into effector memory and resident memory (TEM/TRM). In contrast, anti-PD-1+CTLA-4 activates and expands pre-existing TEM/TRM CD8+ TIL, but does not rejuvenate exhausted phenotypes into T effector cells. Anti-PD-1+LAG-3, but not anti-PD-1+CTLA-4, induces widespread TCR sharing among the different transcriptional states, as well as increased TCR diversity in responding patients. Our data suggest doublet regimen-specific transcriptional and clonal dynamics of tumor-reactive CD8+ T cells.

overall_design: FACS sorted CD45+ CD3+ cells (or CD45+ only in case of low cell numbers) were isolated from baseline and post immunotherapy treatment tumors of HNSCC patients were collected. These samples were run through the 10x genomics protocols and scRNASeq, CITE-Seq and TCR-Seq libraries were generated

first: Housaiyin

last: Li

first: Dan

last: Zandberg

first: Aditi

last: Kulkarni

first: Lazar

last: Vujanovic


title: single-cell RNA-seq analysis for high-dose testosterone treated LuCaP 96CR PDX tumors

city: Boston

first: Maryam

last: Labaf

state: MA

address: 100 William T Morrissey Blvd

department: Biology

institute: University of Massachusetts Boston

summary: Our single cell RNA sequencng (scRNA-seq) analysis of high-dose testosterone (Hi-T) treated patient derived xenograft model 96CR shows RB1 positive cells displayed a robust response to Hi-T while RB1 negative cells are partiallly resistant to Hi-T.

overall_design: Cells were treated with 40mg/kg testosterone ip everyday for 7 days

first: Wanting

last: Han

first: Han

last: Dong

first: Changmeng

last: Cai


title: RNA sequencing of psoriasis and healthy keratinocytes

city: Uppsala

first: Enikö

last: Sonkoly

address: Entrance 40, 5th floor

department: University Hospital

country: Sweden

institute: Uppsala University

summary: Psoriasis is a common chronic inflammatory skin disease determined by genetic and environmental factors, resulting in IL-23/IL-17-mediated immune activation and epidermal hyperproliferation and activation. Here, we performed RNA sequencing of CD45-negative epidermal cells from psoriasis lesions and healthy skin to characterize the landscape of coding and non-coding transcripts in keratinocytes in psoriasis and healthy skin.

overall_design: Eight individuals with chronic plaque psoriasis and eight healthy controls were enrolled. 4 mm punch biopsies were obtained, the epidermis and dermis separated following dispase treatment and the epidermis was dissociated using trypsin and negatively sorted for CD45 using magnetic beads to enrich epidermal keratinocytes

first: Longlong

last: Luo

first: Jan

middle: C

last: Freisenhausen


title: Denosumab Enhances antitumour Immunity by Suppressing SPP1 and Boosting Cytotoxic T Cells

city: Hong Kong

first: Zezhuo

last: Su

address: No. 21 Sasson Rd, Pokfulam

institute: The University of Hong Kong

summary: Comprehensive single-cell RNA sequencing data set of human giant cell tumour of bone

overall_design: Single-cell RNA sequencing from benign GCTB with or without denosumab treatment and malignant GCTB naïve to denosumab treatment

last: SU

first: Kelvin

last: CHEUNG


title: DNMT3A regulates megakaryocyte-biased hematopoietic stem cell fate decisions [RNA-Seq]

status: Public on Feb 17 2025

first: Margaret

last: Goodell

state: TX

address: One Baylor Plaza, N1030

summary: Hematopoietic stem cells (HSCs) are defined by their capacity to regenerate all main components of the peripheral blood, but individual HSCs exhibit a range of preferences for generating downstream cell types. Their propensities are thought to be epigenetically encoded, but no differential regulatory mechanisms have been identified. Here, we considered whether DNA methyltransferase 3A (DNMT3A) has a role in determining the differentiation choice of HSCs. We find that the most primitive megakaryocyte-biased HSCs depend most on DNMT3A for efficient hematopoietic regeneration, particularly of the lymphoid lineages. Reduced DNMT3A amplifies the biased HSC behavior. DNMT3A also regulates megakaryocyte and platelet output in part through DNA methylation at key megakaryocyte lineage loci. Together, our findings establish the role of epigenetic regulation in the fate of megakaryocyte-biased HSCs and their downstream progeny and suggest that the outcomes of DNMT3A-mutant clonal hematopoiesis and malignancies might vary depending on the identity of the HSC that acquires the mutation.

overall_design: To compare the transcriptomic profiles in WT and Dnmt3a+/– megakaryocyte-biased (Vwf+) and non-megakaryocyte-biased (Vwf-) HSCs: HSCs were sorted by FACS from mouse bone marrow using the markers Lineage–CD117+Sca1+CD150hiCD48lo and either Vwf-GFP+ or Vwf-GFP–. To obtain enough material, 3-4 mice were pooled per replicate.

first: Sarah

last: Waldvogel

first: Virginia

last: Camacho

first: Dandan

last: Fan

first: Anna

middle: G

last: Guzman

first: Alejandra

last: Garcia-Martell

first: Elmira

last: Khabusheva

first: Jacey

last: Rodriguez-Pridgen

first: Josephine

last: De La Fuente

first: Rachel

last: Rau

first: Ashlyn

last: Laidman

first: Maria

last: Barrachina

first: Estelle

last: Carminita

first: Justin

last: Courson

first: Michael

middle: R

last: Williamson

first: Joanne

middle: I

last: Hsu

first: Chun-Wei

last: Chen

first: Jaime

last: Reyes

first: Subhashree

last: Pradhan

first: Rolando

last: Rumbaut

first: Alan

last: Burns

first: Jianzhong

first: Benjamin

last: Deneen

first: Kellie

last: Machlus


title: CD9-Enriched Extracellular Vesicles from Salivary Gland Basal Progenitors to Promote Regeneration in Salivary Gland Fibrosis

city: SKT

first: Sunyoung

last: Park

state: Seoul

country: South Korea

institute: Yonsei university

summary: Effect of miRNAs from CD9+ EVs in salivary gland epithelail cell for anti-fibrotic regeneration in obstructive sialadenitis.

overall_design: Comparative gene expression profiling analysis of miRNA-seq data for total EV, CD9+EV, and CD9-EV

first: Yeo-Jun

last: Yoon

first: Hyo-Il

last: Jung

first: Jae-Yol

last: Lim


title: DNMT3A regulates megakaryocyte-biased hematopoietic stem cell fate decisions [EM-Seq]

first: Margaret

last: Goodell

institute: Baylor College of Medicine

overall_design: To compare the genome-wide methylation patterns in WT and Dnmt3a+/– megakaryocyte-biased (Vwf+) and non-megakaryocyte-biased (Vwf-) HSCs: HSCs were sorted by FACS from mouse bone marrow using the markers Lineage–CD117+Sca1+CD150hiCD48lo and either Vwf-GFP+ or Vwf-GFP–. To obtain enough material, 3-4 mice were pooled per replicate.

first: Sarah

last: Waldvogel

first: Virginia

last: Camacho

first: Dandan

last: Fan

first: Anna

last: Guzman

first: Alejandra

last: Garcia-Martell

first: Elmira

last: Khabusheva

first: Jacey

last: Rodriguez-Pridgen

first: Josephine

last: De La Fuente

first: Rachel

last: Rau

first: Ashlyn

last: Laidman

first: Maria

last: Su


title: Gene expression profile at single cell level of wildtype and BubR1 knockout embryonic E10.5 hearts

city: Omaha

first: Brian

last: North

state: NE

address: 2500 California Plaza

department: Biomedical Science

institute: Creighton University School of Medicine

summary: Congenital heart defects (CHDs) are the most common type of congenital defects in humans and have consequences across the lifespan. Mutations in BubR1 cause mosaic variegated aneuploidy (MVA) syndrome in which patients develop CHDs such as atrial and ventricular septal defects. We observed that cardiac-specific BubR1 deletion causes embryonic lethality due to defects at the structural and cellular defects. We conducted single cell RNA sequencing (ScRNA-seq) to determine the defects that occur in the absence of BubR1 at the molecular level in the developing myocardium.

overall_design: Hearts were isolated from E10.5  wildtype and BubR1 knockout embryos followed by digestion to make single cell suspensions. The samples were then processed according to 10x genomics pipeline for library preparation and sequencing.

first: Renju

last: Pun

first: Aradhana

last: Thapa

first: Sylar

last: Takafuji

first: Rexton

middle: M

last: Suzuki

first: Gabrielle

middle: F

last: Kay

first: Toni

last: Howard

first: Michael

middle: H

last: Kim


title: Intracellular HIV-1 Tat regulator induces epigenetic changes in the DNA methylation landscape [RNA-seq]

city: Majadahonda

first: Ruben

last: Ayala-Suárez

state: Madrid

address: Crta Pozuelo Km 2

department: AIDS Immunopathology Unit

country: Spain

institute: Instituto de Salud Carlos III

summary: The HIV regulatory protein Tat enhances viral transcription but also modifies host gene expression, affecting cell functions like cell cycle and apoptosis. Cohort studies indicate that, despite virological suppression, people with HIV (PWH) are at increased risk of comorbidities linked to chronic inflammation, accelerated immune-aging and cellular senescence, sometimes associated with abnormal genomic methylation patterns. In this study, we analyzed whether Tat influences DNA methylation and subsequently impact the transcriptional signature, contributing to inflammation and accelerated aging. To this aim, Jurkat cells were transfected with full-length Tat (Tat101), Tat’s first exon (Tat72), or an empty vector (TetOFF) and DNA methylation changes were assessed. Differentially expressed genes (DEG) were identified via RNA-seq. Results showed that Tat101 expression resulted in major hyper- and hypomethylations changes at individual CpG sites resulting in a slightly global DNA hypermethylation. Methylation changes at gene promoters and bodies resulted in altered gene expression, specifically regulating gene transcription in 5.1% of DEG in Tat101 expressing cells. In contrast Tat72 had a minimal impact on this epigenetic process. The observed differentially methylated and expressed genes were involved in inflammatory responses, lipid antigen presentation, and apoptosis, and may constitute a key epigenetic mechanism contributing to HIV pathogenesis and chronic inflammation.

overall_design: We aimed to profile the transcriptional changes together with DNA methylation patterns induced by HIV protein Tat to investigate its impact in gene regulation. Jurkat TetOFF cell line from Clontech was stably transfected with a pTRE2hyg plasmid encoding full-length HIV-Tat cDNA (101aa; Tat101), a pTRE2hyg plasmid encoding only the first exon of HIV-Tat (72aa; Tat72), or an empty pTRE2hyg vector as a control (TetOFF). In this model, the administration of 1 μg/ml doxycycline  to the culture medium for 48h repress Tat expression. We performed RNASeq by triplicates in the three experimental conditions and in Tat101 transfected cells exposed 48h to doxycycline.

first: Francisco

last: Diez-Fuertes

first: Andrea

last: Rodriguez-Agustin

first: José

last: Alcamí

first: Núria

last: Climent


title: Intracellular HIV-1 Tat regulator induces epigenetic changes in the DNA methylation landscape

type: Methylation profiling by genome tiling array

overall_design: Refer to individual Series

first: Rubén

last: Díez-Fuertes

last: Rodríguez-Agustín

first: Angelika

last: Merkel

first: Izar

last: de Villasante


title: Lysine acetylation of aquaporin-3 promotes water reabsorption but is not essential for urine concentration

city: Birmingham

first: Kelly

middle: Anne

last: Hyndman

state: Al

address: 1918 University BLVD

department: Medicine

institute: University of Alabama at Birmingham

summary: The current study aimed to decipher the role of previously discovered K282 acetylation of aquaporin-3 (AQP3) in renal water homeostasis. C57BL/6J mice harboring amino acid subsitutions to model lysine acetylation (AQP3 K282Q, termed Q mutant) or deacetylation (AQP3 K282R, termed R mutant)  and wild type (WT) controls were used to determine if these mutations affected the kidney transcriptome under ad libitum pelled chow and water (standard housing condictions). Male and female mice were included in the transcriptomic analyses, and all samples were collected between 8-11 am (Zeitgeber 2-5).

overall_design: Wild Type (WT), acetylated mimetic (Q), and deacetylated mimetic (R), male and  female mice were kept in standard housing conditions (pelleted food and tap water ad libitum; 12 h light: 12 h dark cycle).  Mice were euthanized between ZT2-5 and kidneys excised, decapsulted, and snap frozen for RNA analyses.

first: K

middle: A

first: N

middle: V

last: Huynh


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

city: London

first: Probir

last: Chakravarty

address: 1 Midland Road

institute: Francis Crick Institute

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

middle: J

last: Henderson


title: Patterning effects of FGF17 and cAMP on generation of dopaminergic progenitors for cell replacement therapy in Parkinson’s disease. [RNA-Seq]

city: Copenhagen

first: Erno

last: Hänninen

address: Blegdamsvej 3B

department: reNEW, Novo Nordisk Foundation Center for Stem Cell Medicine

country: Denmark

institute: University of Copenhagen

summary: Cell replacement therapies using human pluripotent stem cell-derived dopaminergic (DA) progenitors are currently in clinical trials for treatment of Parkinson’s disease (PD). Recapitulating developmental patterning cues, such as fibroblast growth factor 8 (FGF8), secreted at the midbrain-hindbrain boundary (MHB), is critical for the in-vitro production of authentic midbrain DA progenitors. Here, we explore the application of alternative MHB-secreted FGF-family members, FGF17 and FGF18, for DA progenitor patterning. We show that while FGF17 and FGF18 both recapitulate DA progenitor patterning events, FGF17 induced expression of key DA progenitor markers at higher levels than FGF8 and transplanted FGF17-patterned progenitors fully reversed motor deficits in a rat PD model. Early activation of the cAMP pathway mimicked FGF17-induced patterning, although strong cAMP activation also came at the expense of EN1 expression. In summary, we identified FGF17 as a promising candidate for more precise and robust DA progenitor patterning, with potential to improve cell products for treatment of PD.

overall_design: To compare the ability of different MHB-associated FGFs from the FGF8 subfamily to fine-tune hPSC-derived ventral midbrain (VM) DA progenitors, we added either FGF8 or FGF17 from day 9 to 16 for the production of DA progenitors. On day 9 the cells were treated either with FGF8 or FGF17 for 15 min, 1 h, 4 h, and 24 h (n=3). We performed bulk RNA sequencing for these treatments to investigate immediate differences in FGF response pathways. On differentiation day 16 we performed scRNA-seq on hashtag-labeled (hto) DA progenitors patterned with either FGF8 or FGF17 (n=4).

first: Agnete

last: Kirkeby