title: Endometrial extracellular vesicles regulate processes related to embryo development and implantation in human blastocysts.

city: Valencia

first: Hortensia

last: Ferrero

state: Valencia

address: Avenida Fernando Abril Martorell 106, Torre A

department: Reproductive Medicine

country: Spain

institute: Instituto de Investigación Sanitaria La Fe / Fundación IVI

summary: Extracellular vesicles (EVs) mediate intercellular communication by transporting various molecules, and endometrial EVs have been postulated to be involved in the molecular regulation of embryo implantation. The objective of this study was to investigate the transcriptomic response of human blastocysts following internalization of endometrial EVs in vitro. Primary human endometrial epithelial cells, derived from endometrial biopsies collected from fertile oocyte donors, were cultured in vitro to isolate secreted EVs. Following EV characterization, day 5 human blastocysts were cultured in the presence or absence of the EVs for 24 hours and evaluated by RNA-sequencing. Among the differentially expressed genes (DEGs) in blastocysts co-cultured with EVs, we found 519 were significantly upregulated and 395 were significantly downregulated. These DEGs were significantly enriched in upregulated functions related to embryonic development, cellular invasion and migration, cell cycle, cellular organization and assembly, gene expression and cell viability; and downregulated functions related to cell death and DNA fragmentation. Further, the intracellular signaling pathways regulated by the internalization of endometrial EVs were previously related to early embryo development and implantation potential, for their role in pluripotency, cellular homeostasis, early embryogenesis and implantation-related processes. Finally, integrating data from miRNA cargo of EVs, we found that the miRNAs carried by endometrial EVs targeted nearly 80% of the DEGs in human blastocysts. This study provides novel insights into the functional relevance of EVs secreted by the human endometrium, and particularly the role of EV-miRNA regulation on global transcriptome behavior of human blastocysts during early embryogenesis and embryo implantation.

overall_design: To investigate endometrium-embryo crosstalk during implantation, this study uses next-generation sequencing and compares the differential gene expression profiles of day 5 human embryos, cultured with and without extracellular vesicles secreted by primary human endometrial epithelial cells in a secretory-phase hormonal environment.

first: Marina

last: Segura-Benítez

first: Maria

middle: C

last: Carbajo-García

first: Alicia

last: Quiñonero

last: De Los Santos

first: Antonio

last: Pellicer

first: Irene

last: Cervelló


title: Developmental Programming: Sex-specific Effects of Prenatal Exposure to a Real-Life Mixture of Environmental Chemicals on Liver Function and Transcriptome in Sheep.

status: Public on Jan 12 2025

city: Ann Arbor

first: Vasantha

last: Padmanabhan

state: MI

address: 1150, W. Medical Centre Drive

country: USA

institute: university of Michigan

type: Expression profiling by high throughput sequencing

summary: Humans are chronically exposed to a mixture of environmental chemicals (ECs), many with endocrine disrupting potential, contributing to the development of non-communicable diseases. The adverse health effects of exposure to single ECs have been extensively studied but less is known about the impact of chronic exposure to low levels of a mixture of ECs, which is the real-life exposure scenario. Biosolids, derived from human wastewater treatment, closely reflects the human exposome in both the array and concentration of ECs and is a novel, real-life model to investigate the health risks posed by developmental exposure to mixtures of ECs. Previous studies using the prenatal biosolids exposure model found an increased accumulation of the endocrine disrupting chemical diethylhexylphthalate in the fetal liver and metabolic perturbations including increased thyroid gland weight in the male fetus and reduced bodyweight in prepubertal male lambs. To understand the basis of this sexually dimorphic metabolic phenotype we investigated the transcriptome of liver, the primary metabolic organ, in lambs of both sexes. Since many liver genes show sexual dimorphism, with sex-biased genes playing a pivotal role in lipid and drug metabolism, sex-differences in liver gene expression and the impact of prenatal biosolids exposure on it was examined. We hypothesized that maternal preconceptional and gestational exposure of sheep to biosolids programs sex-specific transcriptional changes in the offspring liver. Ewes (F0) were grazed on either inorganic fertilizer (Control) or biosolids-treated pastures (Biosolids) from 1 month before mating till parturition. All lambs (F1) were raised on control pastures after lambing until euthanasia at 9.5 weeks of age. RNA from liver from Control male (n=7), Control female (n=8), BTP male (n=7) and BTP female (n=8) lambs were subjected to next generation sequencing and DESeq2 R package was used to identify differentially expressed genes due to prenatal biosolids exposure and sex-differentially expressed genes .

overall_design: To understand the effect of preconceptional and gestational exposure to biosolids on the liver transcriptome, RNA from liver tissue of F1 lambs were sequenced. Differential gene expression analysis was done between the Control (n=15) and Biosolids (n=15) groups, comprising both male (n=7/group) and female (n=8/group) lambs.


title: Transcriptome analysis of cruabcabc (a triple cruabcciferin (cruabc) mutant of cruabcA, cruabcB, and cruabcC) during seed maturation

city: Columbia

first: Ruthie

last: Angelovici

state: MO

address: 371D Christopher S. Bond Life Sciences Center, 1201 Rollins St

department: Division of Biological Sciences

institute: University of Missouri

summary: An in-depth analysis of the cruabc triple mutant showed the importance of translational machinery and response to oxidative stress in proteome rebalancing during the early onset of maturation.

overall_design: Seeds from Col-0 and cruabc were grown in growth chambers at 22°C/24°C (day/night) under long-day conditions (16 h of light/8 h of dark) and watering was provided to keep plants hydrated at all times. Seeds were harvested at 10, 12, 14, 16, 18, and 20 days after flowering (DAF). The treatment consists of comparing the triple mutant cruabc (T-DNA insertions targeting CRUA (At5g44120.3), CRUB (At1g03880.1), and CRUC (At4g28520.1)) with Col-0 during seed maturation and normal growth conditions.


title: Mechanism study of VEGF-B maintaining genomic stability in HUVECs using mRNA sequencing

city: Guangzhou

first: Yi

last: Liu

state: Guangdong

address: No.7 Jinsui Rd.

department: Vascular biology

country: China

institute: Zhongshan Ophthalmic Center Sun Yat-Sen University

summary: The goal of this study is to dig out DNA damage pathways mediated by VEGF-B. Using RNA-seq differential expression analysis, we observed genes involved in genomic stability maintainence were upregulated in VEGF-B treated HUVECs. Increased expression of 7 key DNA damage processing genes was confirmed by qRT–PCR. Our study displays the transcriptomic change of HUVECs in the early stage of VEGF-B treatment without strong stress and suggests that VEGF-B signal upregulates genes of DNA damage processing pathway in HUVECs.

overall_design: Collecting HUVECs treated with BSA or VEGF-B after 6 hours for mRNA sequencing and comparing the expression changes between the treatments.

first: Jianing

last: Zhang

first: Xuri

last: Li


title: ATAC-seq of CD8 T cells responding to acute and chronic infections

status: Public on Jan 11 2025

city: Atlanta

first: Chris

last: Scharer

state: GA

address: 1510 Clifton Rd, Suite 3086A

department: Microbiology and Immunology

institute: Emory University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: To further understand the chromatin accessibility changes between early and late CD8 T cell populations responding to acute and chronic infections we performed ATAC-seq on early Stem-like, effector and late Stem-like and effector populations.

overall_design: 2-3 samples per time point, infection model, and cell type were profiled by ATAC-seq

first: Christopher

middle: D

first: Rafi

last: Ahmed

first: Daniel

middle: T

last: McManus


title: RNA Sequencing of Mrp8-Cre;TGFbR2-fl/fl Lung Neutrophils

city: Singapore

first: Nicholas

last: Ang

address: 8a Biomedical Grove, Singapore 138648

country: Singapore

institute: Singapore Immunology Network (SIgN)

summary: Purpose: The goal of this study is to examine how the specific loss of TGFbR2 (under Mrp8 promoter for Cre recombinase) changes the transcriptome of neutrophils in the lung compared to their wildtype littermates.

overall_design: Lung neutrophils from Mrp8-Cre;TGFbR2-fl/fl (mutant) or Cre-negative TGFbR2-fl/fl littermate control mice (wt) [B6.Cg-Tg(S100A8-cre,-EGFP)1Ilw/J x B6;129-Tgfbr2tm1Karl/J] (N=3 per group) were purified by flow cytometry-assisted cell sorting, then submitted for bulk RNA sequencing.

first: LiangYao

middle: J

first: Andres

last: Hidalgo

first: Lai

middle: G

last: Ng

first: Ming

middle: Y

last: Chooi

first: Shu

middle: Z

last: Chong


title: Effect of peptide ADSCP2 on gene expression of primary hypertrophic scar fibroblasts

city: Nanjing

first: Jingyun

address: 123rd Tianfei Street, Mochou Road, Nanjing 210004, China.

institute: Women’s Hospital of Nanjing Medical University (Nanjing Maternity and Child Health Care Hospital)

summary: In order to find the downstream mechanism of peptide ADSCP2, we performed RNA-sequencing (RNA-seq) to examine the transcriptome change induced by ADSCP2. Differentially expressed genes were obtained by RNA-seq. Compared with the scramble control, 160 genes were up-regulated (fold change ≥ 2, P < 0.05) and 170 genes were down-regulated (fold change ≤ 0.5, P < 0.05) in the ADSCP2 group.

overall_design: Primary hypertrophic scar fibroblasts were treated with 25 μM Sc or ADSCP2 in triplicates for 24 h and collected for transcriptome library preparation and sequencing (Illumina NovaSeq6000, in PE150 mode) by ShanghaiBio Corporation. Differentially expressed genes (DEGs) were found using edgeR and corrected with false discovery rate (FDR). Genes with a fold change of ≥2 and q value ≤ 0.05 between two groups were identified as differentially expressed genes.

first: Jun


title: The optimum concentration ratio of zinc and magnesium for co-production of bioethanol and pigments for Monascus purpureus

city: Changzhou

first: Xia

last: Yi

address: Gehu Middle Road #21

institute: Changzhou University

summary: Zinc and magnesium were the essectial activators for bioenzyme. Here, we assayed single factor assays for the effect of zinc and magnesium for Monascus purpureus.Then, we carried out transcriptome assays to uncover molecular mechanism of the enhanced fermentability of bioethanol and pigments for M. purpureus under the optimal metal ion concentration.This work would provide the fermentation process for biofuels and biochemicals.

overall_design: We performed gene expression profiling analysis using from RNA-seq data from two different mycelia at one time point for Monascus purpureus.


title: Two parallel neural ectoderm populations contribute to the developing brain [scRNA-seq]

status: Public on Jan 10 2025

city: Stanford

first: Arjun

last: Rajan

state: California

address: 265 Campus Drive

department: Developmental Biology

institute: Stanford University

summary: Stem cell research strives to create diverse types of human brain cells. Given plentiful successes in converting human pluripotent stem cells (hPSCs) into forebrain and midbrain cells, here we focus on generating cells of the human hindbrain, a life-sustaining brain region. This is, in turn, predicated on a developmental roadmap of when and how brain progenitors diversify from one another. In one model, a common brain progenitor can generate all brain regions. Here our studies of mouse embryos and hPSCs support a different model that as early as gastrulation, two parallel brain progenitors emerge simultaneously: anterior neural ectoderm (forebrain/midbrain progenitor) and posterior neural ectoderm (which forms most of the hindbrain). Expanding on classical studies, we suggest anterior and posterior neural ectoderm are already lineage-committed in vitro and cannot transdifferentiate. Anterior and posterior neural ectoderm have divergent chromatin landscapes, respectively foreshadowing their forebrain versus hindbrain potentials. We differentiated hPSCs into posterior neural ectoderm, and subsequently, hindbrain rhombomere 5/6-specific motor neurons. Anterior and posterior neural ectoderm thus represent two parallel developmental routes to generate forebrain/midbrain vs. hindbrain neurons. Finally, anterior and posterior ectoderm are likely conserved from vertebrates to hemichordates, intimating an ancient neural lineage bifurcation that predated chordates, with implications for nervous system evolution.

overall_design: Single cell gene expression was profiled using the Chromium Single Cell Gene Expression Flex kit (10x Genomics) for all progenitor and neuronal cell-types generated in this manuscript. Two separate rounds of experiment were performed, one for D0-D2 differentiated cell populations, and one for D4-D14 cell populations.

first: Carolyn

middle: E

last: Dundes

first: Rayyan

last: Jokhai

first: Hadia

last: Ahsan

first: Rachel

middle: S

last: Kang

last: Salomon-Shulman

first: Yoon

last: Seok Kim

first: Liam

last: Stanton

first: Christine

last: Xu

first: Stephanie

last: Do

first: Brennan

last: McDonald

first: José

last: Miguel Andrande López

first: Hugo

middle: A

last: Urrutia

first: Hannah

last: Greenfeld

first: Alicia

last: Wong

first: Yimiao

last: Qu

first: Andrew

last: Petkovic

last: Miao

last: Garcia

first: Michelle

last: Monje

last: Wagner

first: Marianne

last: Bronner

first: Cristopher

last: Lowe

first: Kyle

middle: M

last: Loh


title: Gene expression in signaling subsets of AML blasts induced by G-CSF

first: Yury

last: Goltsev

address: 3220 CCRS, Baxter lab,

department: Department of Microbiology and Immunology

institute: Stanford Medical School

summary: While the response to G-CSF is a significant predictor of AML outcome, the nature of cells responding differentially to this and other cytokines remains unclear. Based on gene expression signatures observed in G-CSF Responsive Cells (RC) and non-G-CSF Responsive Cells (NRC) a cohort of AML samples were segregated into two types. In Type1 both cell subsets displayed an undifferentiated phenotype.  In the Type2 AML the RC showed a very similar undifferentiated progenitor-like phenotype as in Type1 RC, whereas the NRC exhibited a more differentiated hematopoietic phenotype. The frequency of RC per sample was a determinant of whether a patient belonged to Type1 or Type2. Type2 RC had higher expression of CD321, higher colony formation capacity and greater responsiveness to cytokines secreted by NRC. Further analysis of clinical mRNA databases revealed that CD321 was a prognostic indicator for AML and a potential marker of clonogenic activity associated with this AML cancer lineage.

overall_design: Cryopreserved PBMC from 11 AML patinets were stimulated with G-CSF. Following stimulation cells were fixed, permeabilized and stained for selected surface markers (CD3, CD33, CD45) and and phospho-Stat3/5. After staining subsets positive and negative for phospo-Stat3/5 were sorted out for further microarray analysis. T-cells were sorted from each sample to control for the RNA quality by comparisons to RNA prepared from live T-cells sorted form healthy donors.


title: Generation of self-renewing neuromesodermal progenitors with neuronal and skeletal muscle bipotential from human embryonic stem cells

city: Shanghai

first: Pingxin

last: Sun

address: 800 Xiangyin Road, Yangpu District

department: Department of Cell Biology

institute: Naval Medical University

summary: Progress has been made in generating spinal cord and trunk derivatives from neuromesodermal progenitors (NMPs). However, maintaining the self-renewal of NMPs in vitro remains a challenge. In this study, we developed a cocktail of small molecules and growth factors that induces human embryonic stem cells to produce self-renewing NMPs (srNMPs) under chemically defined conditions. These srNMPs maintain the state of thoracic neuromesodermal progenitors in prolonged culture and have the bipotential to generate mesodermal cells and neurons, even at the single-cell level. Additionally, suspended srNMP aggregates could partially mimic the structural properties of early embryonic trunks. Furthermore, transplanted srNMP-derived muscle satellite cells or progenitors of motor neurons were integrated into skeletal muscle or the spinal cord, respectively, and contributed to regeneration in mouse models. In summary, srNMPs hold great promise for applications in developmental biology and as renewable cell sources for cell therapy for trunk and spinal cord injuries.

overall_design: Differentiate NMP induced by H1 and HUES9 cell lines into neurons and skeletal muscles, respectively


title: Endogenous fine-mapping of functional regulatory elements in complex genetic loci in HAP1 [scMulti]

city: Tianjin

first: Ke

last: Zhao

address: Qixiangtai Road, No.22

institute: Tianjin Medical University

summary: The majority of genetic loci linked to polygenic complex traits are found in non-coding regions of the human genome. These loci often exhibit complex gene regulatory relationships and linkage disequilibrium (LD) configurations, making it challenging to accurately identify causal variants and their target genes. We used multiplexed single-cell CRISPR interference and activation perturbations to investigate cis-regulatory element (CRE) and target gene expression relationships within tight LD in the endogenous chromatin context. We demonstrated the prevalence of multiple causality in perfect LD (pLD) for independent expression quantitative trait locus (eQTL) and uncovered fine-grained genetic effects on gene expression within pLD, which are difficult to decipher using traditional eQTL fine-mapping or existing computational methods. We found that over one third of the causal CREs lack classical epigenetic markers, and we functionally validated one of these hidden regulatory mechanisms. Leveraging Multiome single-cell epigenetic and sequence perturbations, we highlighted the regulatory plasticity of the human genome. Our study will guide the exploration of missing causal mechanisms in molecular trait formation and disease development.

overall_design: Nine scRNA-seq datasets under varying multiplicity-of-infection (MOI) conditions, three single-cell multiome datasets (ATAC + Gene Expression) from modified HAP1 clones(dCas9-KRAB-blast, dCAS9-VP64-blast and Cas9-blast stably expressed HAP1 clones), four ATAC-seq datasets from wild-type and CRISPR-edited HAP1 cells(64bp deletion, 17bp deletion and CRISPRi around rs73156934-tagged cis-regulate element(CRE)), six CUT&TAG histone modification datasets and 1 Micro-C dataset from wild-type HAP1 cells, and three 4C-seq datasets for validating CRE-gene regulatory relationships in HAP1 cells.

first: Yao

last: Zhou

first: Mulin

last: JunLi


title: YAP1 Activation Rescues Nagashima-type Palmoplantar Keratosis Pathological Features in Skin Organoid Disease Models

first: Huiting

last: Liu

address: Lujing road NO.2

institute: Dermatology Hospital, Southern Medical University

summary: YAP1 activation rescues NPPK pathological features Abstract Nagashima-type palmoplantar keratosis (NPPK), a recessive genetic skin disorder, is characterized by the presence of keratosis on the palmar and plantar regions, resulting from SERPINB7 mutation. However, the detail pathogenesis of NPPK remains elusive. This study revealed that SERPINB7 does not interact with proteolytic enzymes directly. Instead, SERPINB7 mutants disrupt keratinocyte proteolysis by down-regulating the expression of KLKs, and MMPs in NPPK epidermis. Mechanistically, SERPINB7 mutants inhibit keratinocyte glycolytic metabolism by reducing HK1 kinase activity, which subsequently impedes the activation, and nuclear translocation of transcription factor YAP1. This cascade ultimately leads to decreased expression of proteolytic enzymes within the KLKs, and MMPs families in keratinocytes. Furthermore, we successfully established a skin organoid disease model harboring the SERPINB7 c.796C>T mutation, faithfully recapitulating the clinical, and molecular features of NPPK lesions. Notably, treatment of NPPK skin organoid models with YAP1 agonists significantly improved the pathological changes of the disease. In summary, our findings offer novel insights into the molecular mechanisms underlying NPPK pathogenesis and identify YAP1 as a promising therapeutic target for this disease.

overall_design: To further unravel NPPK mechanisms, we generated a clinically relevant SERPINB7 mutant (c.796C>T) in NHEK cells using CRISPR/Cas9-mediated homologous recombination

first: Jun

first: Wen

last: Zheng

first: Yingying

last: Luo

first: Tao

last: Yang

first: Qiuping

last: Zhang

first: Qiuyu

last: Chen

first: Bin


title: Spaceflight alters protein levels and gene expression associated with cell survival and metabolic characteristics in human cardiac spheroids

first: Gregory

middle: K

last: Tharp

address: 954 Gatewood Dr

department: Developmental and Cognitive Neuroscience

institute: Yerkes National Primate Research Center

summary: Cardiomyocytes derived from human induced pluripotent stem cells (hiPSC-CMs) possess tremendous advantage for cardiac regeneration. However, cell survival is challenging upon cell transplantation. Since microgravity can profoundly affect cellular properties, we investigated the effect of spaceflight on hiPSC-CMs. Cardiac spheroids derived from hiPSCs were transported to the International Space Station (ISS) via the SpaceX Crew-8 mission and cultured under space microgravity for 8 days. Beating cardiac spheroids were observed on the ISS and upon successful experimentation by the astronauts in space, the live cultures were returned to Earth. These cells had normal displacement (an indicator of contraction) and Ca2+ transient parameters in 3D live cell imaging. Proteomics analysis revealed that spaceflight upregulated many proteins involved in metabolism (n=90), cellular component of mitochondrion (n=62) and regulation of proliferation (n=10). Specific metabolic pathways upregulated by spaceflight included glutathione metabolism, biosynthesis of amino acids, and pyruvate metabolism. In addition, spaceflight upregulated cellular stress response- and survival-related proteins and the AMPK signaling pathway, a master regulator of metabolism. Transcriptomic profiles indicated that spaceflight upregulated genes associated with cardiomyocyte development, and cellular components of cardiac structure and mitochondrion. Furthermore, spaceflight upregulated genes in metabolic pathways associated with cell survival such as glycerophospholipid metabolism and glycerolipid metabolism. These findings indicate that short-term exposure of the space environment to 3D hiPSC-CMs led to significant changes in protein levels and gene expression involved in cell survival and metabolism.

overall_design: 3 replicates of Cardiac spheroids derived from hiPSCs were transported to the International Space Station (ISS) via the SpaceX Crew-8 mission and cultured under space microgravity for 8 days are compared to 3 replicates of the same type cultured on the ground using the same cell culture device.

first: Parvin

last: Forghani

first: Wenhao

first: Zeyu

last: Wang

first: Zhi

last: Ling

first: Felipe

last: Takaesu

first: Evan

first: Sheila

last: Nielsen

first: Shankini

last: Doraisingam

first: Stefanie

last: Countryman

first: Michael

last: Davis

first: Ronghu

last: Wu

first: Shu

last: Jia

first: Chunhui

last: Xu


title: Two parallel neural ectoderm populations contribute to the developing brain [RNA-seq]

first: Arjun

last: Rajan

overall_design: Bulk RNA-seq on pure populations of hPSCs, aNE, pNE, and fore/mid/hindbrain differentiation of aNE/pNE. Total RNA was extracted  for all cell types in 2 distinct biological replicates.

first: Carolyn

last: Dundes

first: Rayyan

last: Jokhai

first: Hadia

last: Ahsan

first: Rachel

last: Kang

last: Salomon-Shulman

first: Yoon

last: Seok Kim

first: Liam

last: Stanton

first: Christine

first: Stephanie

last: Do

first: Brennan

last: McDonald

first: José

last: Miguel Andrande López

first: Hugo

last: Urrutia

first: Hannah

last: Greenfeld

first: Alicia

last: Wong

first: Yimiao

last: Qu

first: Andrew

last: Petkovic

first: Yi

last: Miao

first: Christopher

last: Garcia

first: Michelle

last: Monje

first: Daniel

last: Wagner

first: Marianne

last: Bronner

first: Cristopher

last: Lowe

first: Kyle

last: Loh


title: Dysregulated ac4C modification of mRNA in a mouse model of early stage Alzheimer's disease [RNA-seq]

city: shanghai

first: danhong

last: wu

address: 3rd Floor, Building 2, 801 Heqing Road, Shanghai

department: Department of Neurology

institute: Shanghai Fifth People’s Hospital, Fudan University

summary: In this study, ac4C modifications in mRNA were investigated using six-month-old 5×FAD transgenic mice, a widely recognized model of Alzheimer's disease, along with age- and sex-matched wild-type (WT) controls. To elucidate the role of ac4C-modified mRNA in AD, we employed three analytical techniques: acetylated RNA immunoprecipitation sequencing (ac4C-RIP-seq), RNA sequencing (RNA-seq), and proteomic analysis. The first two were used to identify mRNAs carrying ac4C modifications and to quantify mRNA abundance, respectively.

overall_design: mRNA-seq was used to determine the abundance of mRNA in the hippocampus from 6 month mice (3 5xFAD mice and 3 WT mice).

first: Haonan

last: Ji

first: Haiqian

last: Zhou

first: Jingbo

last: Qie

first: Wenmei

last: Lu

first: Haitao

last: Gao

first: Danhong

last: Wu


title: A comparative analysis of tissue-based and liquid-based next-generation sequencing in patients with anaplastic thyroid carcinoma

city: Taoyuan

first: Chiao-Ping

last: Chen

address: No.259, Wenhua 1st Rd., Guishan Dist.

country: Taiwan

institute: Chang Gung University

summary: Ongoing progress in high-throughput next-generation sequencing (NGS) has enabled clinicians to comprehensively characterize the genomic landscape of tumors, guide treatment decisions, and facilitate clinical trial enrollment. The role of liquid NGS in anaplastic thyroid carcinoma (ATC) remains unclear, particularly in cases where tissue NGS is not feasible or yields inadequate results. Among these, 26 patients had adequate tissue for commercially available tissue NGS (ACTOnco®+, 440 genes), 15 had access to a commercially available liquid NGS platform (ACTMonitor®+, 50 genes), and 13 patients underwent both tissue and liquid NGS. The genetic alterations observed in ATC exhibited a high degree of heterogeneity, involving several pathways, including RAS/RAF/MEK/ERK, PI3K/AKT/mTOR, cell cycle regulation, other receptor tyrosine kinases, DNA damage response, DNA mismatch repair (MMR, 34.6%, including MLH1, MSH6, MSH2, and PMS1), and chromatin remodeling. The most frequently mutated genes in tissue NGS were TP53 (17/26, 65.4%) and BRAF (8/26, 30.8%). Among the 13 pairs analyzed on both platforms, the concordance rates were 84.6% and 69.2% for BRAF and TP53, respectively. Among two patients without sufficient tissue for NGS, liquid NGS provided additional information on genetic alterations. NGS platforms, whether applied to tissue or liquid samples, can empower clinicians to identify targetable oncogenic events in ATC. Liquid biopsy provides supplementary information when the tissue is insufficient for NGS.

overall_design: ATC patients provided adequate tissue samples for analysis using commercially available tissue-based, liquid-based NGS, or both. The study aimed to assess the frequency of established genetic events in tumorigenesis, identify potential novel driver mutations, and characterize candidates for targeted therapy. Additionally, it examined the concordance between tissue and liquid NGS results in ATC cases.

first: Chun Nan

last: Yeh

first: Shu Fu

last: Lin

first: Chia Ling

first: Miaw Jene

last: Liou

first: I Wen

first: Chiao Ping

first: Ching Fu

last: Chang

first: Qi An

last: Wang

first: Chiao En


title: Two parallel neural ectoderm populations contribute to the developing brain [ATAC-seq]

first: Arjun

last: Rajan

overall_design: Bulk ATAC-seq on pure cell populations of hPSCs, DE, aNE, pNE, and fore/mid/hindbrain differentiation of aNE/pNE. OmniATAC-seq protocol was performed for all cell types in 3 distinct biological replicates.

first: Carolyn

last: Dundes

first: Rayyan

last: Jokhai

first: Hadia

last: Ahsan

first: Rachel

last: Kang

last: Salomon-Shulman

first: Yoon

last: Seok Kim

first: Liam

last: Stanton

first: Christine

last: Xu

first: Stephanie

last: Do

first: Brennan

last: McDonald

first: José

last: Miguel Andrande López

first: Hugo

last: Urrutia

first: Hannah

last: Greenfeld

first: Alicia

last: Wong

first: Yimiao

last: Qu

first: Andrew

last: Petkovic

first: Yi

last: Miao

first: Christopher

last: Garcia

first: Michelle

last: Monje

first: Daniel

last: Wagner

first: Marianne

first: Cristopher

first: Kyle


title: Hepatic Lpcat3 Accelerates Nonalcoholic Fatty Liver Disease by slut1e1-related promoting fatty acid oxidation and reducing fatty acid synthesis [HFHS RNA-seq]

city: 上海

first: 艳群

last: 罗

state: --- Select One ---

address: Cailun Road 1200

institute: shanghai university of traditional Chinese medicine

summary: Non-alcoholic steatohepatitis (NASH) is a common chronic liver disease associated with metabolic disorders such as obesity, diabetes, and high cholesterol. It results in inflammation and fibrosis of liver tissue, eventually leading to cirrhosis and liver cancer. While LPCAT3 has been linked to the formation of fat cells, its role in the development of NASH is not yet fully understood. The purpose of this study was to gain insights into the mechanisms that accelerate NASH induced by LPCAT3. We utilized a genetic engineering rodent model and transcriptomics sequencing approach to explore the role and regulatory mechanism of LPCAT3 in the development of NAFLD. Approach & Results: We studied the impact of Lpcat3 deficiency on NASH progression using three distinct Lpcat3 liver-specific knockout mouse (LKO) models and conducted RNA sequencing, lipidomics, and metabolomics studies on liver samples. Human samples were obtained and analyzed for LPCAT3 expression in patients with NASH to determine its correlation with disease severity. The HepG2 and Huh-7 cell lines were utilized for in vitro analyses. We discovered that LPCAT3 expression was elevated in human NASH livers, and its expression correlated with NAFLD activity score and fibrosis stage. Lpcat3 deficiency in the mouse liver slows the development of diet-induced NASH. Lpcat3 deficiency mechanistically reduces lipid production by inhibiting lipid metabolic pathwaysDeletion of Lpcat3 significantly improved lipid accumulation in NASH mice, possibly due to the involvement of sult1e1 in lipogenesis and oxidation regulation. Moreover, the hepatic overexpression of Lpcat3 significantly intensified lipid buildup in steatotic cells, indicating that LPCAT3 plays a crucial role in the progression of NASH.  Conclusions: The findings indicate that LPCAT3 regulates the participation of sult1e1 in both adipogenesis and oxidation, leading to enhanced NASH outcomes.

overall_design: To investigate the role and mechanistic pathways of LPCAT3 in the context of non-alcoholic fatty liver disease (NAFLD), we induced the disease in C57BL/6 mice through the administration of a high-fat, high-fructose(HFHS) diet.

first: Tao

last: Wu

first: Yanqun

last: Luo

first: Caiyun

last: Zhang

first: Guang

last: Ji


title: Characterization of E2F8-mediated transcriptional regulation in lethal prostate cancer

city: Durham

first: Qianben

last: Wang

address: 905 S. Lasalle Street

institute: Duke University

summary: To identify the genomic bindings and transcriptional targets of E2F8 in lethal prostate cancer cells. To examine the potential off-target effects by CRISPR/CasRx-mediated E2F8 knockdown in 293FT cells. To analyze differentially expressed genes upon TT3 LLN-CRISPR/CasRx-mediated E2F8 knockdown in DU145 xenografts.

overall_design: We performed E2F8 ChIP-seq to map genome-wide E2F8 binding sites in PC3 and DU145 cells. To identify E2F8-regulated genes, RNA-seq was performed in PC3 and DU145 cells transfected with control siRNA or siRNA targeting E2F8 for 72h. To evaluate the off-target effects of CasRx-mediated E2F8 silencing, RNA-seq was conducted in 293FT cells transfected with either a CasRx expression plasmid along with a non-targeting pre-gRNA control(pre-gCtrl), or a pre-gRNA targeting E2F8(pre-gE2F8) plasmid for 24h. To elucidate the molecular mechanisms by which TT3 lipid-like nanoparticles(LLNs) carrying CasRx/pre-gE2F8 inhibit DU145 xenografts growth, RNA-seq was performed to identify the differentially expressed genes between TT3 LLN-CasRx/pre-gCtrl and TT3 LLN-CasRx/pre-gE2F8 treated groups.

first: Furong

last: Huang

first: Kexin

last: Li

first: Zhong

last: Chen

first: Kun

last: Fang


title: Inhibition of PCSK9 with evolocumab modulates lipoproteins and monocyte activation in high-risk ASCVD subjects

first: Sascha

last: Goonewardena

state: Michigan

address: 1500 E Medical Center Dr

department: Department of Internal Medicine

institute: University of Michigan

summary: Abstract: BACKGROUND: Mechanistic studies suggest that proprotein convertase subtilisin/kexin type 9 monoclonal antibody inhibitors (PCSK9mAb) not only reduce low density lipoprotein cholesterol (LDL-C) but can also modulate inflammation. OBJECTIVES: This trial was designed to define the impact of evolocumab on circulating immune cell properties in high-risk ASCVD subjects. METHODS: Double-blind, placebo-controlled trial randomized 41 ASCVD subjects with type 2 diabetes with microalbuminuria and LDL-C level of >70 mg/dL on maximum tolerated statin therapy to receive either subcutaneous evolocumab 420 mg every 4 weeks or matching placebo. The primary outcomes were the change in lipoproteins, blood viscosity, and circulating immune cell transcriptional response at 2 weeks and 12 weeks with evolocumab compared with matching placebo. Safety was assessed in all subjects who received at least one dose of assigned treatment and analyses were conducted in the intention-to-treat population. RESULTS: All 41 randomized subjects completed the 2-week visit. Six subjects (4 evolocumab, 2 placebo) did not receive study medication consistently after the 2-week visit due to suspension of research activities during the COVID-19 pandemic. The groups were well-matched with respect to age, comorbidities, baseline LDL-C, white blood cell counts, and markers of systemic inflammation.  Evolocumab reduced LDL-C by -68.8% (p<0.0001) and -52.8% (p<0.0001) at 2 and 12 weeks respectively. There were no differences in blood viscosity at baseline nor at 2 and 12 weeks. RNA-seq was performed on peripheral blood mononuclear cells (PBMC) with and without TLR4 stimulation (“Stress” transcriptomics). There were no group differences in transcriptional profiles in unstimulated PBMC; however, “stress” transcriptomics unmasked immune cell phenotypic differences between evolocumab and placebo groups at 2 and 12 weeks. CONCLUSIONS: This trial is the first to demonstrate that PCSK9 mAB with evolocumab can modulate circulating immune cell properties and highlights the importance of “stress” profiling of circulating immune cells that more clearly define immune contributions to ASCVD. (Funded by Amgen Inc.; ClinicalTrials.gov number, NCT03829046).

first: Robert

last: Rosenson

first: Ashley

last: Tate

first: Phyu

last: Mar

first: Olga

last: Grushko

first: Qinzhong

middle: N


title: The miR-290 and miR-302 clusters are essential for reprogramming of fibroblasts to induced pluripotent stem cells

city: San Francisco

last: Blelloch

address: 35 Medical Center Way

department: Dept. of Urology

institute: UC, San Francisco

summary: The miR-290 and miR-302 clusters of microRNAs are highly expressed in naïve and primed pluripotent stem cells, respectively. Ectopic expression of the embryonic stem cell-specific cell cycle regulating (ESCC) family of microRNAs arising from these two clusters dramatically enhances the reprogramming of both mouse and human somatic cells to induced pluripotency. Here, we used genetic knockouts to dissect the requirement for the miR-290 and miR-302 clusters during the reprogramming of mouse fibroblasts into induced pluripotent stem cells (iPSCs) with retrovirally introduced Oct4, Sox2, and Klf4. Knockout of either cluster alone did not negatively impact the efficiency of reprogramming. Resulting cells appeared identical to their embryonic stem cell counterparts. Notably miR-290 knockout cells showed a compensatory increase in miR-302 expression and the combined loss of both clusters blocked the formation of iPSCs. While rare double knockout clones could be isolated, they showed a dramatically reduced proliferation rate, a persistent inability to fully silence the exogenously introduced pluripotency factors, and a transcriptome distinct from individual miR-290 or miR-302 mutant ESC and iPSCs. Taken together, our data show that miR-290 and miR-302 are essential yet interchangeable in reprograming to the induced pluripotent state.

overall_design: Comparative gene expression analysis of miRNA knockout cells either derived as ESCs or made by reprogramming MEFs with Oct4/Sox2/Klf4. Except for WT iPSC controls, each biological replicate (e.g. colony 1 or 2) was independently derived, cultured, collected, and prepared for RNA-seq. All data provided come from cells cultured in LIF+FBS media conditions. For iPSC or iPSC-like cells, samples were collected more than 18 passages after reprogramming.

first: Ryan

last: Boileau


title: CXCL12 produced from Foxl1high mesenchymal cells modulates epithelial cell metabolism and prevents intestinal neoplasia (RNA-Seq)

city: Suita

first: Daisuke

last: Motooka

state: Osaka

address: 3-1, Yamadaoka

country: Japan

institute: NGS core facility, Research Institute for Microbial Diseases, Osaka University

summary: Several mesenchymal cell populations are involved in the maintenance and differentiation of intestinal stem cells (ISCs) by secreting Wnts, R-spondins, and bone morphogenetic proteins. However, the influences of signaling mediators derived from mesenchymal cells other than ISC niche factors on epithelial homeostasis remain poorly understood. Here, we revealed that CXCL12 produced from Foxl1high sub-epithelial mesenchymal cells regulates epithelial cell proliferation through modulation of the mevalonate pathway, which contributes to prevention of tumorigenesis. Foxl1-cre; Cxcl12f/f mice showed increase in the number of Ki67+ proliferative cells in the colonic epithelium, which was decreased by treatment with mevalonate biogenesis inhibitor simvastatin. Moreover, Cxcl12 deficiency in Foxl1high mesenchymal cells promoted adenoma development in the colon and ileum of ApcMin/+ mice. Collectively, these results demonstrate the critical role of CXCL12 derived from Foxl1high sub-epithelial cells in prevention of intestinal neoplasia through modulation of cellular metabolism in epithelial cells.

overall_design:
CD45- EpCAM- CD31- gp38+ PDGFRα high Ly6C neg mesenchymal cells were isoloated from colon of  Foxl1cre; Cxcl12 flox/flox  and  Cxcl12 flox/flox mouse and gene expression analysis using data from RNA-seq of the mesenchymal cells were performed.
Comparative gene expression analysis of RNA-Seq analysis for CD45- EpCAM- CD31- gp38+ PDGFRα high Ly6C neg mesenchymal cells from C57BL/6 mice treated with 1mM GABA and 20mM taurine at two time points(0hr and 6hr)

first: Mayu

last: Yagita-Sakamaki

first: Tkayoshi

last: Ito


title: Inhibiting hepatic LPCAT3 ameliorates alcoholic liver injury through regulating NAMPT and PTLP signaling

first: 艳群

last: 罗

summary: Background and Aims: Lysophosphatidylcholine acyltransferase 3 (LPCAT3), a key molecule regulating intracellular phospholoesters has abundantly expression level in metabolic tissues including liver, intestine and adipose. And it is considered to be directly related to the biosynthesis of inflammatory lipid mediators in humans. However, the mechanism of LPCAT3 in alcoholic liver disease (ALD) is unclear. Approach and Results: Liver-specific LPCAT3 liver knockout (LPCAT3-LKO) mice were established using CRISPR/Cas9, and then an ALD model followed by NIAAA modele was established. After LPCAT3-specific knockout of liver tissue, ALT, AST, TG, TC and liver TG, TC in ALD mice were reduced. Our group found that three downstream targets of LPCAT3, namely phospholipid transfer protein (PLTP) and nicotinamide phosphoribosyltransferase (NAMPT) and elongation of long-chain fatty acids family member 5 (ELOVL5) through RNA sequencing and the unbiased systematic investigative approach. In addition, our experimental data indicated that LPC and PC metabolism were altered in ALD mice by using targeted metabolomics. PCR and WB showed that PLTP and NAMPT protein expression levels were ascended in the LKO_x005F_x0002_EtOH group than Flox-EtOH group in the ALD model group. At the same time, in ethanol-induced cell models, LPCAT3 overexpression upregulated ALT and TG levels, aggravated hepatocytes, damage and lipid production, and reduced the expression of PLTP and NAMPT. Conclusions: The results provide novel views on the significance of LPCAT3 in regulating liver phospholipid synthesis in the pathogenesis of ALD. Reduction of LPCAT3 in liver tissue is able to mitigate alcohol-induced hepatocyte damage and lipid production by regulating PLTP and NAMPT.

overall_design: To investigate the mechanistic role of LPCAT3 in alcoholic liver injury, we established an NIAAA mouse model.

first: Tao

last: Wu

first: Ying

last: Gao

first: Junmin


title: CXCL12 produced from Foxl1_high mesenchymal cells modulates epithelial cell metabolism and suppresses intestinal neoplasia in Apc_Min/+ mice (16S rRNA-seq)

summary: Several mesenchymal cell populations are involved in the maintenance and differentiation of intestinal stem cells (ISCs) by secreting Wnts, R-spondins, and bone morphogenetic proteins. However, the influences of signaling mediators derived from mesenchymal cells other than ISC niche factors on epithelial homeostasis remain poorly understood. Here, we revealed that CXCL12 produced from Foxl1_high sub-epithelial mesenchymal cells regulates epithelial cell proliferation through modulation of the mevalonate pathway, which contributes to prevention of tumorigenesis. Foxl1-cre; Cxcl12_f/f mice showed increase in the number of Ki67+ proliferative cells in the colonic epithelium, which was decreased by treatment with mevalonate biogenesis inhibitor simvastatin. Moreover, Cxcl12 deficiency in Foxl1_high mesenchymal cells promoted adenoma development in the colon and ileum of Apc_Min/+ mice. Collectively, these results demonstrate the critical role of CXCL12 derived from Foxl1_high sub-epithelial cells in prevention of intestinal neoplasia through modulation of cellular metabolism in epithelial cells.

overall_design: We analyzed the average frequencies of relative abundance of bacteria at the family level by fecal DNA sequencing of 16S ribosomal DNA in untreated Cxcl12f/f and Foxl1-cre; Cxcl12f/f mice.


title: TrAEL-seq for WRN inhibitors

city: Cambridge

first: Laura

last: Biggins

address: Babraham Research Campus

department: Bioinformatics

country: United Kingdom

institute: The Babraham Institute

summary: Cancers with microsatellite instability (MSI) depend on the WRN helicase enzyme to manage issues during DNA replication caused by long stretches of (TA) repeats in the DNA. Targeting WRN is a promising strategy for treating MSI cancers, and drugs that inhibit WRN are being developed. Through a process called fragment-based screening, we developed powerful and specific drugs that block WRN's action. These drugs effectively slowed down the growth of MSI cancer models in lab and animal studies by acting like WRN is absent, leading to DNA breaks at the long TA repeats and causing DNA damage. The development of these potent and specific drugs targeting WRN in MSI cancers proves that this approach can work and also helps us understand more about WRN's role in biology.

overall_design: We employed Transferase-Activated End Ligation sequencing (TrAEL-seq), a method that captures single-stranded DNA 3′ ends genome-wide and with base pair resolution, to identify the genomic sites of DNA double-strand breaks (DSBs) in MSI cells treated with WRN inhibitors. We used HCT116 cells engineered with an inducible sgRNA system for WRN targeting (i-WRN) as a positive control, along with GSK4532858A-treated HCT116 cells and GSK4532858A-treated KM12 cells

first: Kieron

last: May

first: Gabriele

last: Picco

first: Shriram

last: Bhosle

first: Angham

last: Al Saedi

first: Mathew

last: Garnet

first: Jonathan

last: Houseley


title: A Metabolic Switch Controlling Liver Cancer Evolution from Senescent NASH Hepatocytes

city: La Jolla

first: Masafumi

last: Ohira

address: 9500 Gilman Drive

department: Department of Pharmacology

institute: University of California San Diego

summary: Hepatocellular carcinoma (HCC) originates from differentiated hepatocytes undergoing compensatory proliferation in livers damaged by viruses or nonalcoholic steatohepatitis (NASH). While increasing HCC risk, NASH triggers TP53-dependent hepatocyte senescence, which we found to parallel hypernutrition-induced DNA breaks. How this tumor-suppressive response is bypassed to license accumulation of oncogenic mutations and enable HCC progression was previously unknown. We identified the gluconeogenic enzyme fructose-1,6-bisphosphatase 1 (FBP1) as a TP53 target that is elevated in senescent-like NASH hepatocytes but suppressed through promoter hypermethylation and proteasomal degradation in most human HCCs. FBP1 first declines in metabolically-stressed premalignant disease-associated hepatocytes and HCC progenitor cells, paralleling the protumorigenic activation of AKT and NRF2. By accelerating FBP1 and TP53 degradation AKT and NRF2 enhance the proliferation and metabolic activity of previously senescent HCC progenitors. The senescence-reversing NRF2-FBP1-AKT-TP53 metabolic switch, operative in mice and humans, also enhances proliferation-enabled accumulation of DNA damage-induced somatic mutations that drive NASH to HCC progression.

overall_design: MUP-uPA and Mup-uPA/Fbp1ΔHep mice were fed with high fructose diet and live freshly livers were collected at 40 weeks. liver nuclei were extracted and  analyzed using snRNA-seq.

first: Li

last: Gu

first: Yahui

last: Zhu

first: Shuvro

middle: P

last: Nandi

first: Maiya

last: Lee

first: Kosuke

last: Watari

first: Breanna

last: Bareng

first: Yuxiao

last: Liu

first: Sadatsugu

last: Sakane

first: Rodrigo

last: Carlessi

first: Consuelo

last: Sauceda

first: Debanjan

last: Dhar

first: Souradipta

last: Ganguly

first: Marcos

last: Teneche

first: Peter

last: Adams

first: David

last: Gonzalez

first: Mojgan

last: Hosseini

first: Tatiana

last: Kisseleva

first: Janina

last: Tirnitz-Parker

first: M

last: Simon

first: Ludmil

middle: B

last: Alexandrov

first: Michael

last: Karin


title: Population dynamics modeling reveals myeloid bias involves both HSC differentiation and progenitor proliferation biases

city: Los Angeles

first: Apeksha

last: Singh

state: CA

address: 570 Boyer Hall

institute: University of California, Los Angeles

summary: Aging and chronic inflammation are associated with overabundant myeloid-primed multipotent progenitors (MPPs) amongst hematopoietic stem and progenitor cells (HSPCs).  While HSC differentiation bias has been considered a primary cause of myeloid bias, whether it is sufficient has not been quantitatively evaluated.  Here, we analyzed bone marrow data from the IκB– (Nfkbia+/-Nfkbib-/-Nfkbie-/-) mouse model of inflammation with elevated NFκB activity, which shows increased myeloid-biased MPPs.  We interpreted these data with differential equations models of population dynamics to identify alterations of HSPC proliferation and differentiation rates.  This analysis revealed that short-term (ST) HSC differentiation bias alone is likely insufficient to account for the increase in myeloid-biased MPPs.  To explore additional mechanisms, we used single-cell RNA sequencing (scRNA-seq) measurements of IκB– and wild-type HSPCs to track the continuous differentiation-trajectories from HSCs to erythrocyte/megakaryocyte, myeloid, and lymphoid primed progenitors.  Fitting a partial differential equations model of population dynamics to these data revealed not only less lymphoid-fate specification amongst HSCs, but also increased expansion of early myeloid-primed progenitors.  Differentially expressed genes along the differentiation-trajectories supported increased proliferation amongst these progenitors.  These findings were conserved when wild-type HSPCs were transplanted into IκB– recipients, indicating that an inflamed bone marrow microenvironment is a sufficient driver.  We then applied our analysis pipeline to published scRNA-seq measurements of HSPCs isolated from aged mice, as well as human myeloid neoplasm patients.  These analyses identified the same myeloid-primed progenitor expansion as in the IκB– models, suggesting that it is a common feature across different settings of myeloid bias.

overall_design: Lin- Sca1+ cKitHi (LSK) HSPCs were isolated from the bone marrow of the bilateral femur and tibia by flow sorting, from young WT and IκB– mice, and analyzed by scRNA-seq.

first: Jennifer

last: Chia

first: Alexander

last: Hoffmann


title: CXCL12 produced from Foxl1_high mesenchymal cells modulates epithelial cell metabolism and suppresses intestinal neoplasia in Apc_Min/+ mice (3' scRNA-seq)

first: Daisuke

last: Motooka

overall_design: CD45- EpCAM- CD31- gp38+ mesenchymal cells were isolated from colon of adult(>8-week-old) mice and Germ Free mice

first: Bo

last: Li

first: Hisako

last: Kayama


title: The cationic liposome of  trehalose dimycolate (TDM) inducees antitumor immunity in bladder.

city: Tsukuba city

first: Kozaburo

last: Tanuma

state: Ibaraki

address: Tennodai 2-1-1

department: Urology

institute: University of Tsukuba

summary: BCG therapy is the most effective immunotherapy for bladder cancer, but it sometimes causes serious adverse events because of infection with live BCG bacteria. To develop more effective and less toxic immunotherapeutic agents, we have developed formulations using BCG cell wall components. Trehalose dimycolate (TDM) is a predominant glycolipid that constitutes the cell wall of BCG, and reported to show activation of innate immunity and have adjuvant effect. We already established hydrophilic cationic liposomes incorporating TDM, and reported that local administration of the cationic liposome TDM (Lip-TDM) exerted antitumor effect to subcutaneously inoculated tumors by inducing activation of CD8+ T cells via dendritic cell maturation. In present study, we demonstrated that intraperitoneal administration of Lip-TDM exerted antitumor effects in orthotopic bladder carcinogenesis mouse model induced by N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN). In this model, intraperitoneal administration of Lip-TDM enhanced the systemic activation of dendric cells and macrophages, and increased infiltration of adaptive immune cells such as CD8+ T cells. Furthermore, Lip-TDM treatment induced Natural Killer (NK) cells to be cytotoxic NK cells in tumors and regulatory NK cells in lymphoid tissues. Lip-TDM treatment induced activation of DCs and cytotoxic CD8+ T cells and exerted antitumor effect in WT mice, but not in NK cell-depleted mice. These results indicate NK cells are essential for Lip-TDM to induce sufficient acquired immune response. Our findings elucidate the mechanism of the antitumor effect by Lip-TDM and suggest that Lip-TDM is a non-infectious agent which can be administered intraperitoneally and has potential to be an alternative to BCG.

overall_design: RNA sequencing was performed using the bladders from the wild-type C57BL/6 mice treated with Lip-TDM or Lip-CON. Bladders were extracted from sacrificed animals at baseline and after 16 weeks of treatment.

first: Jun

last: Miyazaki

first: Masanobu

last: Shiga

first: Keisuke

last: Sano

first: Yoshiyuki

last: Nagumo


title: Analysis of the developmental function of the PAIRED-Like homeobox transcription factor Drgx in T4/T5 neurons  of Drosophila melanogaster [RNA-seq]

city: Regensburg

first: Mathias

last: Rass

address: Universitätsstraße 31

country: Germany

institute: University of Regensburg

summary: Targeted DamID was performed to identify binding targets of Drgx in T4/T5 neurons. RNA-seq analysis of differentially expressed genes  following the knockdown of Drgx in T4/T5 neurons of late third instar larvae, compared with control.

overall_design: RNAseq of FACS sorted T4/T5 neurons from wandering third instar larvae (control: R23G12 x cherry-IR, experiment: R23G12 x DRGX-IR)

first: Laura

last: Gizler

first: Stephan

last: Schneuwly


title: CXCL12 produced from Foxl1_high mesenchymal cells modulates epithelial cell metabolism and suppresses intestinal neoplasia in Apc_Min/+ mice (5' scRNA-seq)

overall_design:
CD45- EpCAM+ epithelial  cells were isolated from colon of Foxl1cre; Cxcl12flox/flox  and  Cxcl12flox/flox mouse and analyzed using scRNAseq.
CD45- EpCAM- CD31- gp38+ mesenchymal cells were isolated from colon of embryo, neonate, 3-week-old and 4-week-old mice and analyzed using scRNAseq.

first: Mayu

last: Yagita-Sakamaki


title: Expression profiling of HPV+ Patient Derived Xenografts (PDXs) by high throughput sequencing

city: PHILADELPHIA

first: Devraj

last: Basu

state: PENNSYLVANIA

address: 3400 SPRUCE STREET, 5 RAVDIN

department: Otorhinolaryngology

institute: Perelman School of Medicine, University of Pennsylvania

summary: The incidence of HPV-related oropharyngeal squamous cell carcinoma (OPSCC) has increased in recent years. RNAseq was performed for systems-based analysis of cellular pathways involved in HPV+ve OPSCCs.

overall_design: We sought to analyze transcriptional profiling of 7 HPV+ OPSCC PDXs

first: Basu

last: Devraj

first: Sannigrahi

last: Malay

first: Rajagopalan

last: Pavithra


title: Analysis of the developmental function of the PAIRED-Like homeobox transcription factor Drgx in T4/T5 neurons  of Drosophila melanogaster [DamID-Seq]

overall_design: DamID-Seq on T4/T5 neurons from wandering third instar Drosophila larvae expressing Dam control or Dam-Drgx transgenes driven by R23G12-Gal4 (three replicates each).


title: The impact of WDR77 on the AR cistrome in prostate cancer cells

city: Buffalo

first: Tao

last: Liu

state: NY

address: Elm and Carlton St

department: Biostatistics and Bioinformatics

institute: Roswell Park Comprehensive Cancer Institute

summary: Androgen receptor (AR) drives prostate cancer (CaP) even after androgen deprivation therapy (ADT) failed. Inhibiting AR’s transcription factor function to overcome acquired resistance to ADT is an attractive therapeutic avenue. However, which AR-coregulator interaction should be targeted is not clear, AR-coregulator complex higher order structures and stoichiometry are unknown, and how manipulating AR-coregulator interactions impacts the AR cistrome is not understood. Here, we examine interactions between AR and WD repeat 77 (WDR77, non-catalytic component of the methylosome complex), which are enriched in ADT-resistant CaP and mediate CaP cell survival. We performed CUT&RUN assays defined AR and WDR77 cistromes, the WDR77-dependence of the AR cistrome and the overlap between the AR and WDR77 cistromes in CaP cells.

overall_design: In a first experiment, LNCaP cells were seeded in RPMI1640 medium supplemented with 9% FBS without added antibiotics. The next day, cells were transfected with siRNAs targeting WDR77 or non-targeting control siRNA. The next day, medium was changed to medium supplemented with 9% charcoal-stripped FBS. After 80hours, medium was refreshed and cells were treated with 5nM R1881 or vehicle (ethanol) for another 16h. CUT&RUN assays were performed using an antibody targeting AR. In a second experiment, LNCaP cells were seeded in RPMI1640 medium supplemented with 9% charcoal-stripped FBS.  Two days later, medium was refreshed and cells were treated with 5nM R1881 or vehicle (ethanol) for another 16h. CUT&RUN assays were performed using an antibody targeting WDR77.

first: Ujjwal

middle: R

last: Dahiya

first: Hannelore

last: Heemers

first: Song


title: ITK-SYK induced peripheral T-cell lymphomas induce granulocyte activation

city: Freiburg

first: Geoffroy

last: Andrieux

address: Breisacherstr 153

institute: University clinics Freiburg

summary: ITK-SYK or control vector were overexpressed in Balb/c mice and ITK-SYK mice developed peripheral T-cell lymphomas within 5 weeks. CD4+ T-cells and granulocytes were harvested from the spleens of control and ITK-SYK mice and compared.

overall_design: Comparison of ITK-SYK+ and control CD4+ T-cells and granulocytes

first: Melanie

last: Boerries

first: Amelie

last: Jäger

first: Christine

last: Dierks


title: RNAseq analysis for transcriptomes of  DAC treated myeloid malignant cell lines expressing p53-R282W

first: Jiale

last: Wu

address: Room 707, Science and Education Plaza  197 Rui Jin Er Road

institute: State Key Laboratory of Medical Genomics, Shanghai Institute of Hematology, Rui Jin Hospital, Shanghai Jiaotong University

summary: To globally evaluate transcriptome treated by decitabine (DAC) in THP-1 and U937 cells expressing p53-R282W, cells were treated with DMSO or DAC. mRNA was isolated and then subject to deep sequencing, using Illumina HiSeq Xten. The sequence reads that passed quality filters were analyzed using htseq. The expressions had been validated using qRT–PCR using SYBR Green assays.    Results and conclusions: Global transcriptional change of THP-1 expressing p53-R282W treated with 5 μM DAC was analyzed. Protein interaction network and Enrichment assay was analyzed with the 216 upregulated genes exemplified by ISG15 and IFI27) (Gfold ≥0.4, (RPKM+1 fold change > 1.5), the type I interferon signaling was hit .Similar observations were achieved in U937 cells.

overall_design: RNAseq to evaluate transcriptional activity upon DAC treatment.

first: Min

last: Lu


title: Single-Cell RNA Sequencing of PBMCs from a Patient with Recurrent Anti-Synthetase Syndrome-Associated Interstitial Lung Disease: Pre- and Post-Regimen Adjustment

status: Public on Jan 09 2025

city: Beijing

first: Qingya

last: Shi

address: 30 Shuangqing Road, Haidian District

department: Tsinghua Medicine

institute: Tsinghua University

summary:
Background: A 58-year-old male with anti-synthetase syndrome-associated interstitial lung disease (ASS-ILD) underwent bilateral lung transplantation (LTx) but experienced ILD progression and myositis worsening 38 weeks post-transplant, despite glucocorticoids, tacrolimus, and mycophenolic acid treatment. High-dose glucocorticoids led to adverse effects and ILD exacerbation upon tapering.
Objective: To identify persistent inflammatory pathways despite anti-rejection therapy.
Methods: Two sets of single-cell RNA-seq (scRNA-seq) analyses were conducted on peripheral blood mononuclear cells (PBMCs) from our patient who experienced recurrent ASS-ILD following bilateral LTx: one before regimen adjustment and one after. The goal these analyses is to investigate systemic inflammatory profiles.
Results: In comparison to five ASS-ILD patients without LTx, our patient following bilateral LTx (before regimen adjustment) displayed a higher proportion of CD14⁺ and CD16⁺ monocytes, alongside reduced T, B, and NK cells. Notably, interferon- and JAK-STAT–associated pathways were significantly enriched, as evidenced by marked up-regulation of key genes (e.g. JAK1, JAK2, STAT1, STAT2, STAT3, IL4R, IL6R) in the patient’s monocytes. Elevated serum ferritin levels and the enrichment of macrophage and neutrophil activation pathways further underscored hyperactivation of the mononuclear phagocyte system.
Due to poor tolerance of high-dose steroids, limited efficacy of tacrolimus and MPA, and the need to manage post-transplant rejection while controlling ASS, the patient’s regimen was modified. Considering the activation of the mononuclear phagocyte system in the patient, tacrolimus was replaced with cyclosporine A, which is more widely used in macrophage activation syndrome (target trough concentration 120-150 ng/ml). Given the significant activation of the interferon and JAK-STAT pathways in the patient’s activated monocytes, we replaced EC-MPS with the Janus kinase inhibitor (JAKi) tofacitinib at 5 mg twice daily. Short-term high-dose glucocorticoids (methylprednisolone 120 mg/d for 3 days) were also administered before rapid tapering. After regimen adjustment, scRNA-seq was conducted again, which indicated a reduction in CD14⁺ and CD16⁺ monocytes, with T, B, and NK cells rebounding. Neutrophils, characterized by high S100A8, S100A9, and DEFA3 expression, became more prominent. Interferon-related gene expression decreased in monocytes and neutrophils, indicating successful suppression of the interferon pathway.
Conclusions: We presented a complicated case of a patient with early recurrence of underlying ASS-ILD after bilateral lung transplantation. We used two sets of scRNA-seq analyses to investigate the systemic inflammatory responses.

overall_design: scRNA-seq of PBMCs from the patient before and after regimen adjustment

first: Shiyao

last: Wang

first: Huaping

last: Dai

first: Wenhui

last: Chen


title: The effects of differential gut microbiota on hepatic gene expression

city: Cincinnati

first: Joo-Seop

last: Park

address: 3333 Burnet Avenue

institute: Cincinnati Children's Hospital Medical Center

summary: Our preliminary data suggest that differential gut microbiota modulates acetaminophen-induced hepatotoxicity (APAP toxicity) in mice model. The goal of our study is to determine whether commensal gut microbiota modulates the hepatic gene expressions potentially responsible for modulating APAP toxicity.

overall_design:
Method: C57BL/6 male mice from two different vendors [Jackson (JAX) and Taconic (TAC)] were sacrificed and liver tissue collected.  Cecum materials from JAX or TAC mice were inoculated to germ-free (GF) mice. After 4 weeks of conventionalization, mice were overnight fasted (16 h) and sacrificed for liver collection.
Examination of 5 groups; JAX, TAC, GF, GF_JAX and GF_TAC

first: Sungjoon

last: Cho

first: Eunah

last: Chung

first: Hyunyoung

last: Jeong


title: Spatial transcriptomic profiling of the lung tissues from a patient with recurrent anti-synthetase syndrome associated interstitial lung disease after bilateral lung transplantation and one untreated patient

summary: Background: A 58-year-old male with anti-synthetase syndrome-associated interstitial lung disease (ASS-ILD) underwent bilateral lung transplantation (LTx) but experienced ILD progression and myositis worsening 38 weeks post-transplant, despite glucocorticoids, tacrolimus, and mycophenolic acid treatment. High-dose glucocorticoids led to adverse effects and ILD exacerbation upon tapering. Objective: To identify persistent inflammatory pathways despite anti-rejection therapy. Methods: 10X Visium HD spatial transcriptomic analysis was conducted on lung tissues from two distinct patients: one untreated patient with ASS-ILD without LTx serving as a control, and another patient who experienced bilateral LTx. This analysis aims to investigate the inflammatory profiles within the lung tissues and to explore the correlation between pulmonary lesions and systemic inflammation. Results: In comparison to the control patient, the lung tissue from the post-transplant patient appeared to have an increased proportion of macrophages, with a concurrent decrease in the proportions of plasma cells and T cells. Among the macrophages of interest, the post-transplant patient appeared to have an elevated proportion of alveolar macrophages, whereas the control patient seemed to have a larger proportion of monocyte-derived macrophages. Considering that interstitial pneumonia is often driven by alveolar epithelial cell injury, we observed that the post-transplant patient appeared to have a higher proportion of transitional AT2 cells, which were appeared to express both AT2-specific markers (SFTPC, SFTPD) and AT1-specific markers (AGER, RTKN2, GPRC5A). Conclusions: We presented a complicated case of a patient with early recurrence of underlying ASS-ILD after bilateral LTx. We used spatial transcriptomic analysis to investigate the differences between systemic inflammatory responses and lung tissue responses.

overall_design: Visium HD spatial transcriptomics of lung tissues from a patient who underwent bilateral lung transplantation and from an untreated ASS-ILD patient


title: Longistylin A inhibited SaeRS-dependent hemolysin expression and activated Type VII secretory system of Staphylococcus aureus.

city: Guangzhou

first: Xiancai

last: Li

state: None Selected

address: 723 Xingke Road, Tianhe District, Guangzhou, CN 510650

department: South China Botanical Garden

institute: Chinese Academy of Sciences

summary: To maintain cellular homeostasis under the stresses, bacteria always initiate various responses. The transcriptomic profiles of S. aureus treated with longistylin A were assessed by RNA-sequencing. A total of 2585 genes were identified. KEGG analysis enriched differentially expressed genes (DEGs) in different physiological processes. Two-component system and bacterial infection metabolisms, the most significant enrichment pathways, were downregulated by longistry A. The SaeRS two-component system is essential for the hemolytic activity and virulence in S. aureus. SarT is a repressor of alpha-hemolysin (Hla) in S. aureus. Transcriptome data shows that the expression of genes encoding gama-hemolysin components and SaeRS two-component system were inhibited under the treatment of LGA, while the expression of SarT is increased. The Type VII secretory system (T7SS) is responsible for the secretion of virulence factors. Longistry A dramatically upregulated the expression of T7SS related genes, such as essB and esaC. LGA did not affect intracellular Hla accumulation. The downregulation of hemolysin-related genes and upregulation of T7SS related genes may be due to the feedback regulation of inhibited hemolysin secretion caused by longistry A treatment. Overall, longistry A inhibited hemolysin secretion by targeting cell membrane phospholipids, thereby inhibited SaeRS-dependent hemolysin expression and activated Type VII secretory system.

overall_design: S. aureus ATCC 6538 were treated with 0 or 4 μg/mL of longistry A for 4 h and collected for total RNA isolation.


title: Progenitor Effect in the spleen drives early recovery via Universal Hematopoietic Inflation [scMultiome]

city: Minato-ku

first: Takaharu

last: Kimura

state: Tokyo

address: 4-6-1 Shirokanedai

institute: The University of Tokyo

summary: Hematopoietic stem cells (HSC) possess the capacity to regenerate the entire hematopoietic system. However, the precise HSC dynamics in the early post-transplantation phase remains an enigma. Clinically, the initial hematopoiesis in the post-transplantation period is critical, necessitating strategies to accelerate hematopoietic recovery. Here, we uncovered the spatiotemporal dynamics of early active hematopoiesis, “Hematopoietic Inflation”, using a highly sensitive in vivo imaging system and established universal HSPCs capable of immediately supplying vast numbers of blood cells. Hematopoietic Inflation occurs in three peaks in the spleen after transplantation, with common myeloid progenitors (CMPs), notably characterized by HSC-like signatures, playing a central role. Leveraging these findings, we developed expanded CMPs (exCMPs), characterized by a gene expression pattern that selectively proliferates in the spleen and promotes hematopoietic expansion. Moreover, universal exCMPs supported early hematopoiesis in allogeneic transplantation. Human universal exCMPs have the potential to be a viable therapeutic enhancement for all HSC transplant patients.

overall_design: Single-cell multiome analysis : Fresh CMPs, exCMPs and SPaCMPs were collected for single-cell RNA and ATAC sequencing.

first: Takao

last: Yogo

first: Satoshi

last: Yamazaki


title: Two parallel neural ectoderm populations contribute to the developing brain

city: Stanford

first: Arjun

last: Rajan

address: 265 Campus Drive

institute: Stanford University

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

overall_design: Refer to individual Series


title: Progenitor Effect in the spleen drives early recovery via Universal Hematopoietic Inflation [Bulk RNA-Seq]

overall_design: RNA-seq profiling of fresh CMPs from bone marrow and expanded CMPs (exCMPs)


title: CDK4/6 inhibition mitigates chemotherapy-induced expansion of TP53-mutant clonal hematopoiesis

city: SAINT LOUIS

first: Irenaeus

last: Chan

state: Missouri

address: 660 S. Euclid Ave

department: Oncology

institute: Washington University in Saint Louis

summary: Therapy-related myeloid neoplasms (tMN) represent a fatal consequence of exposure to cytotoxic therapy administered in the treatment of cancer. Individuals with pre-existing TP53 clonal hematopoiesis (CH) are at high risk for development of tMN but currently avoidance of therapy is the only strategy to reduce tMN risk. Here, we show in four randomized clinical trials of the CDK4/6 inhibitor trilaciclib given in conjunction with a variety of chemotherapeutic regimens and across diverse cancer patient populations that trilaciclib mitigates expansion of chemotherapy-related CH clones with mutations in DNA damage response genes (including TP53, PPM1D, and CHEK2 mutations). This finding was also observed in a syngeneic murine model of TP53 mutant CH demonstrating that trilaciclib blocks platinum-induced TP53 competitive repopulation through promoting hematopoietic stem and progenitor quiescence and decreasing TP53 mutant stemness advantage. This represents the first demonstration of a pharmacologic strategy to block chemotherapy-induced expansion of pre-leukemic TP53-mutant clones.

overall_design: Following 4 weeks of treatment with either vehicle, carboplatin, trilaciclib, or a combination of carboplatin with trilaciclib, bone marrow cells from competitive transplanted Vav-cre Tp53 WT or Vav-cre Tp53 R172H/WT mice were washed twice with cold PBS before labeling with unique TotalSeqA-compatible HashTags. Subsequently, samples were incubated with mouse Fc receptor-blocking agent and labeled with DAPI for live cell sorting by flow cytometry. After sorting equal numbers of cells were pooled and submitted for scRNA-sequencing

first: Pu

last: Zhang

first: Xiangyu

last: Pan

first: Cynthia

last: Castro

first: Nina

last: Fox

first: Giulia

last: Petrone

first: J Scott

last: Beeler

first: Duc

last: Tran

first: Catrina

last: Fronick

first: Omar

last: Abdel-Wahab

first: Kelly

last: Bolton


title: Q241R mutation of Braf gene causes neurological abnormalities in a mouse model of cardio-facio-cutaneous syndrome independent of developmental malformations

city: Maebashi

first: Hiroshi

last: Ohnishi

state: Gunma

address: 3-39-22 Showa-machi

department: Department of Laboratory Sciences

institute: Gunma University Graduate School of Health Sciences

summary: Constitutively active mutants of Braf cause cardio-facio-cutaneous (CFC) syndrome, which is characterized by growth and developmental defects, cardiac malformations, characteristic facial features, cutaneous manifestations, and mental retardation. An animal model of human CFC syndrome, a systemic BrafQ241R/+ mutant mouse has been reported to exhibit multiple CFC syndrome-like phenotypes. In this study, we analyzed the effects of Braf mutation on neural functions separately from their effects on developmental processes. To this end, we generated Braf mutant mice that express BrafQ241R specifically in mature excitatory neurons (n-BrafQ241R/+). We found no growth retardation or cardiac malformations in n-BrafQ241R/+ mice, indicating normal development of these mice. Behavioral analysis revealed that the mutant mice exhibited decreased locomotor activity and exploratory behavior, enhanced auditory startle response, and impaired hippocampus-dependent learning, many of which were similar to the systemic BrafQ241R/+ mouse. In the hippocampus of n-BrafQ241R/+ mice, the long-term potentiation of synaptic transmission was enhanced, and active forms of ERK1/2 were increased. Transcriptome analysis of hippocampal tissue revealed significant changes in the expression of genes involved in synaptic function and memory learning, as well as increased expression of Dusp6 and Spred3, suppressors of the RAS/mitogen-activated protein kinase (MAPK) signaling pathway. These data suggest that the neuronal dysfunction observed in the systemic CFC mouse model is due to the disruption of homeostasis of RAS/MAPK signaling pathway by the activated Braf mutant after maturation, rather than abnormal development of the brain, and a similar mechanism may be possible in human CFC syndrome.

overall_design: To investigate the effect of hippocampal function on BrafQ241R mutant expression, we generated mice expressing BrafQ241R specifically in mature excitatory neurons.

first: Akira

last: Moriya

first: Shin-ici

last: Inoue

first: Fumihito

last: Saitow

first: Hidenori

last: Suzuki

first: Yoko

last: Aoki


title: Transcriptional profiling of mouse skin upon antibody-mediated neutrophil depletion

city: New Haven

first: Andrés

last: Hidalgo

address: 10 Amistad Street

institute: Yale University

summary: Neutrophils patrol healthy tissues and confer local defense upon perturbations. Recent evidence suggests that they participate in tissue-specific structural programs in addition to their indispensable role in immunity.The goal of this study is to evaluate the effect of neutrophils in healthy mouse skin by comparing the transcriptional landscape of naive skin to neutrophil-depleted skin.

overall_design: Neutrophils were depleted with intraperitoneal injection of anti-Ly6G antibody for 2 consecutive days. Back skin was harvested and processed to obtain single-cell suspensions. Samples were barcoded with Oligo-tagged hashtag antibodies (Biolegend) and incubated with an antibody cocktail of Fixable viability dye eFluor780 (eBioscience), CD45-BUV395, CD11b-BV650, Ly6G-PE, EPCAM-APC, mFESK-4-APC, and CD31-APC. Live neutrophils (CD45+CD11b+Ly6G+), other immune cells (CD45+Ly6G-), and structural cells (CD45-Dump+) were isolated by FACS-based sorting.

first: Alaz

last: Ozcan

first: Andres

first: Andrea

last: Rubio-Ponce


title: Vostok:  A New Looping Factor for the Organization of the Regulatory Genome in the Drosophila Brain

city: Moscow

first: Maksim

last: Erokhin

state: Moscow

address: Vavilov

country: Russia

institute: IGB

summary: The Drosophila regulatory genome is organized in a hierarchical series of topological associating domains (TADs) and intra-TAD regulatory loops.  High-resolution Micro-C maps identified ~645 such loops in the larval brain of Drosophila, spanning distances of 25 kb to 250 kb.  Many of these loops are associated with genes encoding neural adhesion proteins implicated in the formation of specific synapses, including Dprs, Dips, Sides and Beats.  They are formed by interactions of pairs of tethering elements that contain simple repeated sequence motifs, such as GAGAG.  Approximately 10% (64/645) of the loops are lost or diminished in larval brains obtained from homozygous mutants lacking the GAF (GAGA-associated factor) POZ oligomerization domain.  We also present evidence for a second looping factor, the MADF-containing Vostok protein (CG11504), which recognizes a GCAACA motif that is over-represented in brain tethering elements.  There is a loss of 47 of the loops (7%) in larval brains obtained from Vostok mutants. These losses are associated with diminished expression of associated genes, suggesting impaired enhancer-promoter interactions.  Only 9 loops are disrupted in both Vostok and GAF mutants, raising the possibility of a combinatorial code for tether-tether interactions.  Support for this possibility stems from the observation that two previously identified meta-loops spanning 6 Mb rely on both GAF and Vostok. We discuss the prospects of using different combinations of looping factors to produce unlimited 3D associations of genetic loci in animal genomes.

overall_design: ChIP-Seq analyses were performed on the brains of wandering larvae (4-6h before puparium formation). All experiments were performed using wild type or Vostok null mutant larvae.

first: Jie

last: Hu

first: Xiao

last: Li

first: Dmitry

last: Lomaev

first: Nadezhda

last: Vorobyeva

first: Michael

last: Levine

first: Darya

last: Chetverina


title: Chromatin accessibility analysis of human mesenchymal stem cells

city: Pittsburgh

first: Yang

last: Zhang

state: PA

address: 100 Technology Dr.

department: Medicine

institute: University of Pittsburgh

summary: We integrated ATAC-seq analyses of human MSC with CRISPRi screening to identify potential regulatory regions controlling UBE2E2 expression and/or UBE2E1 expression.

overall_design: Duplicate human MSC cell samples were analyzed. Library quality control and sequencing were performed by Novogene (Novogene, CA, USA) on a NovaSeq 6000 sequencer.

first: Silvia

last: Liu

first: Matthew

last: Steinhauser


title: RNA-seq analysis to examine the effect of the Ube2e1 and Ube2e1 knockdown on gene expression profile in 3T3-L1 pre-adipocyte cells

summary: UBE2E2 and UBE2E1 are both members of the same family of E2 conjugating enzymes raising the question of whether they are functionally redundant, particularly given that genetic targeting of both genes inhibited adipogenesis, in vitro. To explore this question, we sought to test for molecular redundancy using transcriptomics analyses. We performed comparative transcriptomics analyses of 3T3-L1 cells in which we targeted either Ube2e2 or Ube2e1.

overall_design: To investigate the cooperative function of Ube2e1 and Ube2e2 in the regulation of adipogenic differentiation, we transduced 3T3-L1 cells with previously validated shRNA retroviruses. We established 3T3-L1 cell lines in which each target gene was knocked down after puromycin selection. Three different stable 3T3-L1 cell lines (shUbe2e1, shUbe2e2, and Scramble) were treated with or without an adipogenic cocktail for 12 hours. We then performed a comparative gene expression profiling analysis using RNA-seq data obtained from six different groups, each with three biological replicates.


title: The Essential Role of ASH2L in Regulating Cell Cycle-Associated Gene Expression to Promote Nephron Progenitor Self-Renewal and Nephron Formation in Mice [RNA-seq]

city: Shanghai

first: Ziyi

last: Zhao

address: 320 Yueyang Road

institute: Shanghai jiaotong university school of medicine

summary: Maintaining adequate nephron number is crucial for normal kidney function, as even slight deficits can predispose individuals to hypertension and other late-onset renal diseases. Congenital anomalies of the kidney and urinary tract arise from abnormal embryonic kidney development, which involves the coordinated reciprocal induction of the ureteric bud and metanephric mesenchyme. Preserving the balance between self-renewal and differentiation of nephron progenitor cells in the metanephric mesenchyme is essential for nephron endowment. Prior work has implicated histone modifications in regulating kidney lineage-specific gene transcription and nephron endowment. Here, we selectively inactivated the H3K4 methyltransferase complex component ASH2L in mouse nephron progenitor cells to elucidate its role in nephron progenitor cell self-renewal and differentiation. The results showed that Ash2l inactivation disrupted H3K4 trimethylation establishment at promoters of genes controlling nephron progenitor cell stemness, differentiation, and cell cycle, leading to premature nephron progenitor cell depletion, cell cycle arrest, and abnormal differentiation of early nephron progenitors, resulting in renal dysplasia at birth. These findings identify ASH2L-mediated H3K4 methylation as a critical epigenetic regulator of kidney development and suggest a novel mechanism contributing to congenital anomalies of the kidney and urinary tract. This knowledge may guide future efforts to unravel the pathogenesis of these conditions and develop targeted therapies.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for FACS-sorted NPCs lineage cells at E15.5 of control and Ash2l cKO mouse embryos.

first: Xuantong

last: Dai

first: Fujun

last: Lin

first: Gengru

last: Jiang


title: The Essential Role of ASH2L in Regulating Cell Cycle-Associated Gene Expression to Promote Nephron Progenitor Self-Renewal and Nephron Formation in Mice [CUT&TAG]

overall_design: Comparative H3K4me3 profiling analysis of CUT&Tag-seq data for FACS-sorted NPCs lineage cells at E15.5 of control and Ash2l cKO mouse embryos.


title: Targeting signaling rewiring and resistant subpopulations in Philadelphia Chromosome-like Acute Lymphoblastic Leukemia [scATAC-seq]

city: Philadelphia

first: Kai

last: Tan

address: 3501 Civic Center Boulevard

department: Pediatrics

institute: Children's Hospital of Philadelphia

summary: Philadelphia chromosome-like acute lymphoblastic leukemia (Ph-like ALL) is a common high-risk leukemia subtype defined by a kinase-activated gene expression pattern and chemoresistance, with low overall survival rates in both children and adults. Clinical responses of patients with Ph-like ALL to tyrosine kinase inhibitor (TKI)-based therapies are incomplete, suggesting partial oncogene addiction and highlighting the need to elucidate alternative biologic dependencies to cure patients. Herein, we report integrated bulk and single-cell multiomics studies of Ph-like ALL cells treated in vitro and in vivo with targeted agents to comprehensively define signaling adaptations and resistant subpopulations. Characterizing residual/resistant human ALL cells harvested from murine spleens revealed transcriptional regulatory network changes and kinase signaling adaptations. We identified key transcription factors including c-MYC and AP1 that mediate transcriptional rewiring associated with therapeutic escape, and demonstrate a specific MYC dependency in Ph-like ALL cells. We found a sub-population of leukemia blasts that have high senescence-associated stem cell-like features that is effectively eradicated only with combined STAT and BCL-2 inhibition. This is clinically significant since both conventional chemotherapy and ruxolitinib are ineffective against senescent stem-like blasts, which may mediate therapeutic tolerance and repopulate relapse. Our findings demonstrate that systems level discovery of key oncogenic dependencies and escape pathways can be exploited for co-targeting or sequential treatment to optimally eradicate blasts and cure this high-risk leukemia subtype.

overall_design: scATAC-seq was performed on Ph-like ALL PDX models treated with control, TKI (ruxolitinib or dasatinib), venetoclax, or combination TKI+venetoclax and harvested at 2 timepoints, 3 days and 28 days.

first: Yang-Yang

last: Ding

first: Jonathan

middle: H

last: Sussman

first: Sarah

last: Tasian


title: Targeting signaling rewiring and resistant subpopulations in Philadelphia Chromosome-like Acute Lymphoblastic Leukemia [Bulk ATAC-seq]

overall_design: Bulk ATAC-seq was performed on Ph-like ALL PDX models treated with control or ruxolitinib at 5 timepoints (0, 3, 7, 28, 56 days).


title: Bcl11b orchestrates subcerebral projection neuron axon development via cell-autonomous, non-cell-autonomous, and subcellular mechanisms

city: Cambridge

first: Yasuhiro

last: Itoh

state: MA

address: 7 Divinity Ave

institute: Harvard University

summary: Both cell-intrinsic competency and extracellular cues regulate axon projection, but mechanisms that coordinate these elements remain poorly understood. Subcerebral projection neurons (SCPN) extend their primary axons from cortex through subcortical structures, including the striatum, targeting the brainstem and spinal cord. We identify that the transcription factor Bcl11b/Ctip2 functions in multiple independent neuron populations to control SCPN axon development. Bcl11b expressed by SCPN is required cell-autonomously for axonal outgrowth and efficient entry into the internal capsule within the striatum, while Bcl11b expressed by medium spiny neurons (MSN) non-cell-autonomously regulates SCPN axon fasciculation within the internal capsule and subsequent pathfinding. Further, integrated investigation of Bcl11b-null SCPN with transcriptomic, immunocytochemical, and in vivo growth cone purification approaches identifies that Cdh13 is localized along axons and on growth cone surfaces of SCPN in vivo, and mediates Bcl11b regulation of SCPN axonal outgrowth. Together, these results demonstrate that Bcl11b controls multiple aspects of SCPN axon development by coordinating intrinsic SCPN cell autonomous subcellular mechanisms and extrinsic MSN non-cell-autonomous mechanisms.

overall_design: RNAseq of in vivo growth cones and somata of wild-type and Bcl11b-null subcerebral projection neurons in developing mouse central nervous system

first: Anne

last: Engmann

first: Dustin

last: Tillman


title: SARS-CoV-2 Variant of Concern Fitness and Adaptation in Primary Human Airway Epithelia

city: St Louis

first: Rita

last: Meganck

address: 660 S Euclid Ave

institute: Washington University in St Louis

summary: Purpose: To identify the cell types change and gene expression change after infection with SARS-CoV-2 variants on primary human airway epithelial cells.

overall_design:
Methods: The primary human large/small airway epithelial cells (LAE/SAE) were cultured on air-liquid interface (ALI) conditions. After 4 weeks, the cells were infected with MOI 0.05 SARS-CoV-2. After 48-96 hours post infection, cells were dissociated with Accutase solution and proceeded to scRNA-seq.
single cell-based mRNA profiles of SARS-CoV-2 D614G, Delta, or Omicron infected HAE cells

first: Cait

last: Edwards

first: Hong

last: Dang

first: Kenichi

last: Okuda


title: PDX models of relapsed pediatric AML preserve global gene expression patterns and reveal therapeutic targets

first: Mark

last: Wunderlich

state: Ohio

address: 3333 Burnet Ave

summary: Patient-derived xenograft (PDX) models are commonly used for preclinical evaluation of targeted therapies. It is important to consider the fidelity with which these systems recapitulate the patient disease state. Currently, there is little information regarding how well pediatric AML PDXs mimic the global gene expression found in patients. Here we analyzed RNAseq data from a diverse series of high-risk pediatric AML PDXs, separately and compared to primary patient data. Unsupervised clustering of PDX data resulted in segregation according to KMT2A (MLL) status. Combined analysis showed PDX samples aligned with patient samples harboring similar genetics. Among KMT2A rearranged samples, we observed strong correlation of expression levels of nearly all expressed transcripts. Furthermore, matched patient/PDX pairs showed strong concordance, suggesting retention of sample-specific gene expression. Interestingly, our analysis uncovered previously unidentified cryptic CBFA2T3-GLIS2 rearrangement in two PDX models. Based on high BCL2 mRNA in these models, we tested the efficacy of venetoclax in combination with chemotherapy. While standard daunorubicin/ara-C treatment failed to produce benefits, CPX-351 decreased disease burden and prolonged survival, particularly when combined with venetoclax. These results validate PDX modeling of high-risk pediatric AML and highlight this system's utility for pre-clinical therapeutic discovery, especially for rare subtypes of disease.

overall_design: We generated a series of PDX models from pediatric AML patient specimens and obtained RNA from BM of mice with high levels of disease. We performed single end RNA sequencing and compared gene expression to samples selected from the NCI TARGET database as well as to residual matched patient samples. PDX models generated in parallel in NSG and NSGS mice were also compared to each other. We identified gene expression profiles associated with MLL and GLIS2 rearrangements in order to identify potential therapeutic strategies.

first: Jing

last: Chen

first: Benjamin

last: Mizukawa


title: Identification of a gene controlling levels of the copper response regulator 1 transcription factor in Chlamydomonas reinhardtii

city: Berkeley

first: Sean

middle: D.

last: Gallaher

address: 176 Stanley Hall, room 408

department: California Institute for Quantitative Biosciences

institute: UC Berkeley

summary: Oxygen prevents hydrogen production in Chlamydomonas (Chlamydomonas reinhardtii), in part by inhibiting the transcription of hydrogenase genes. We developed a screen for mutants showing constitutive accumulation of iron hydrogenase 1 (HYDA1) transcripts in normoxia. A reporter gene required for ciliary motility placed under the control of the HYDA1 promoter conferred motility only in hypoxia. By selecting for mutants able to swim even in normoxia, we obtained strains that constitutively express the reporter gene. One identified mutant was affected in a gene encoding an F-box protein 3 (FBXO3) that participates in ubiquitylation and proteasomal degradation pathways in other eukaryotes. Transcriptome profiles revealed that the mutation, termed cehc1-1 (constitutive expression of hydrogenases and copper-responsive genes), triggers the upregulation of genes known to be targets of copper response regulator 1 (CRR1), a transcription factor involved in the nutritional copper signaling pathway and in the hypoxia response pathway. CRR1 was required for upregulating the HYDA1 reporter gene expression in response to hypoxia and for the constitutive expression of the reporter gene in cehc1-1 mutant cells. The CRR1 protein, normally degraded in Cu-supplemented cells, was stabilized in cehc1-1 cells, supporting the conclusion that CEHC1 facilitates CRR1 degradation. Our results describe a previously unknown pathway for CRR1 inhibition and possibly other pathways leading to complex metabolic changes.

overall_design: RNA-seq was carried out for three strains available at the Chlamydomonas Resource Center (see CC accession numbers in parentheses). The experiment compared the transcriptome of wild-type cells (strain CC-4507) with that of cells carrying a mutation in the CEHC1 gene (JGI ver6.1 Cre01.g005900) (strain CC-4509).  To confirm transcripts regulated by the CEHC1 gene, we also obtained the transcriptome of a strain in which the cehc1-1 mutant phenotype was rescued by transformation with the wild-type CEHC1 gene (strain CC-5386).

first: Xiaoqing

last: Sun

first: Matthew

last: LaVoie

first: Paul

last: Lefebvre

last: Glaesner

first: Daniela

last: Strenkert

first: Radhika

last: Mehta

first: Sabeeha

last: Merchant

first: Carolyn

last: Silflow


title: Media based off the metabolome of tumor interstitial fluid reveals metabolic checkpoints and novel oncometabolites in cancer immunity

first: Greg

last: Delgoffe

address: 5115 Centre Ave.

summary: Cancer evades immunity by establishing an immunosuppressive tumor microenvironment (TME). The metabolic composition of the TME represents a ‘checkpoint’ on cancer immunity as the local milieu, the tumor interstitial fluid (TIF), contains metabolites which may render T cells metabolically deficient and alter immune function. We employed media based on a metabolomic profile of TIF (Tumor Interstitial Fluid Medium; TIFM) to define how T cells behave under TME metabolic stress alone. We found that passage of T cells post-stimulation in TIFM, both limited T cell expansion and imprinted lasting effector dysfunction that persisted even after extended expansion in metabolically replete conditions. And we identified phosphoethanolamine (pEtn), a Kennedy pathway intermediate that accumulates in the TME in many cancers, as a driver of T cell dysfunction. Here, bulk RNA-seq was performed on OT-I CD8 T cells activated with peptide and cultured either in RPMI based media or RPMI based media supplemented with pEtn. We identified several key immunologic genes altered by pEtn treatment. However, there was no notable differentiation pattern associated with pEtn treatment as determined through enrichment analysis of T cell differentiation gene sets, suggesting these cells did not necessarily seem to differentiate into canonical exhaustion or anergic states. We further found in other studies that pEtn, as a precursor for the membrane phospholipid phosphatidylethanolamine (PE), drove elevated PE:PC ratios and modulated TCR signaling, in part by depleting T cells of diacylglycerol required for TCR signal transduction, thus contributing to the observed T cell dysfunction.

overall_design: OT-I CD8 T cells activated with peptide were cultured either in RPMI based media or RPMI based media supplemented with pEtn, and then subjected to bulk RNA sequencing.

first: Yupeng

last: Wang

first: Dayana

last: Rivadeneira


title: Targeting signaling rewiring and resistant subpopulations in Philadelphia Chromosome-like Acute Lymphoblastic Leukemia [scRNA-seq]

first: Kai

last: Tan

overall_design: scRNA-seq was performed on Ph-like ALL PDX models treated with control, TKI (ruxolitinib or dasatinib), venetoclax, or combination TKI+venetoclax and harvested at 2 timepoints, 3 days and 28 days.

first: Yang-Yang

last: Ding

first: Jonathan

last: Sussman

first: Sarah

last: Tasian


title: Prelamin A drives vascular calcification by reorganization of the   epigenetic landscape to promote inflammaging

city: London

first: Catherine

last: Shanahan

address: 125 Coldharbour Lane

institute: King's College London

summary: Background: Vascular calcification is a ubiquitous ageing pathology markedly accelerated in patients with metabolic disorders including diabetes and renal failure.  Prelamin A is a biomarker of vascular ageing which accelerates vascular smooth muscle cell (VSMC) senescence and calcification. Using prelamin A as a model we explored the earliest events in VSMC ageing, focussing on the epigenetic landscape as an environmentally modifiable target.   Methods: Adenoviral expression of prelamin A was used to induce VSMC ageing and epigenetic modifications were monitored using immunofluorescence and targeted PCR-arrays. Epigenetic findings were verified in vitro using drugs targeting epigenetic modifiers and in vivo using immunohistochemistry in human vessels from young, aged and calcified patients.  Transcriptomics, ChIP-seq and antibody arrays were used to identify and verify gene targets impacted by changes in the epigenetic landscape and to monitor effects on inflammation, ageing indices and calcification.  A novel mouse model of VSMC-specific inducible prelamin A expression was used to study epigenetic ageing in vivo and calcification was induced with vitamin D.  Results: Prelamin A accumulation in VSMCs caused rapid global alterations in the repressive heterochromatin marks, H3K9me3 and H3K27me3 which coincided with downregulation of the epigenetic writers EZH2 and SUV39H1.  These same epigenetic changes were recapitulated in primary human VSMCs induced to calcify in response to mineral dysregulation and in the calcified arteries of adults, and young children on dialysis. Global analysis of H3K9me3 and H3K27me3 marks in response to prelamin A expression showed deregulation of cross-talking NF-κB and HIPK2 stress response inflammatory pathways consistent with robust activation of a senescence associated secretory phenotype (SASP). Treatment of VSMCs with EZH2 and SUV39H1 inhibitors amplified the SASP and accelerated senescence and mineral deposition. Induction of prelamin A expression in VSMCs in mice induced rapid loss of heterochromatin which preceded DNA damage and senescence and coincided with early activation of SASP factors creating an environment permissive to calcification.                                                                  Conclusions: Loss of heterochromatin is accelerated by metabolic disease and acts to promote VSMC inflammaging and a tissue microenvironment prone to calcification. Pre-senescent cells with an active SASP may be an early target for tackling vascular ageing pathologies.  tackling vascular ageing pathologies.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for histone modifications H3K9me3 and H3K27me3 in human aortic vascular smooth muscle cells with overexpression of prelamin A.

first: Chin

last: Ho

first: Meng-Ying

last: Wu

first: Sadia

last: Ahmad

first: James

first: Lilia

last: Draganova

first: Grace

last: Anderson

first: Umesh

last: Jonnalagadda

first: Robert

last: Hayward

first: Rukshana

last: Shroff

first: Wilson

middle: L

last: Wen

first: Roger

last: Foo


title: The effect of neutrophil depletion on skin transcriptome

first: Andrés

last: Hidalgo

summary: Neutrophils patrol healthy skin at steady state. Here we evaluate whether the antibody-mediated depletion of neutrophils affect global transcriptome of the skin.

overall_design: scRNAseq of skin from control and antibody-mediated neutrophil depleted mice.

first: Alaz

last: Özcan

first: Andrea

last: Rubio-Ponce