title: RNA-sequencing of primary human glioblastoma resected tissue before and after a personalized, neoantigen-based vaccine plus poly-ICLC (NeoVax); tissues from a Phase 1 clinical trial

city: Columbus

first: Katherine

last: Miller

address: 700 Children's Dr.

department: Institute for Genomic Medicine

institute: Nationwide Children's Hospital

summary: Glioblastoma (GBM) patient outcomes remain low despite surgery, radiation, and chemotherapy. There are few immunotherapy options due to the lack of tumor immunogenicity. Several clinical trials have reported promising results with cancer vaccines, although most trials have used single tumor region sequencing to identify targetable antigens. This approach limits the antigen pool due to the heterogeneity of GBM tumors. We have implemented multisector sequencing to identify targetable neoantigens across the GBM tumor landscape that were then incorporated into personalized peptide vaccines called NeoVax. Here, we report the findings of four subjects enrolled on the trial. Following NeoVax, we observed a clonal expansion of CD8+ T cells in the peripheral blood that, when expanded in vitro, were reactive to, on average, half of the NeoVax neoantigens tested by IFN-g ELISPOT, including BRAF V600E in one subject. A post-NeoVax tumor biopsy was resected from one subject and also showed evidence of infiltrating CD8+ T cell clonal expansion. Although the patients enrolled in this trial did not achieve remission, this pilot study demonstrates the feasibility of NeoVax and the importance of multisector sampling for neoantigen screening.

overall_design: Gene expression analysis of primary human glioblastoma resected from Subjects 1-4 on the Phase 1 clinical trial before administration of a personalized, neoantigen-based vaccine plus poly-ICLC (NeoVax) and one post-NeoVax (subject completed the priming phase and two cycles of booster vaccinations) tumor biopsy from Subject 3

first: Gavin

last: Dunn

first: Tanner

last: Johanns

first: Elizabeth

last: Garfinkle


title: Integration of multisector molecular characterization into a personalized peptide vaccine approach for patients with newly diagnosed glioblastoma; Gene expression using NanoString CAR-T Characterization Panel

status: Public on Apr 12 2024

middle: E

state: OH

country: USA

summary: Incorporation of multisector sampling in glioblastoma increases the candidate pool of potentially targetable neoantigens that can be successfully developed into a peptide-based vaccine

overall_design: Here, we report the results of the CAR-T Characterization Panel from three regions of primary human glioblastoma resected from Subject 3 before administration of a personalized, neoantigen-based vaccine plus poly-ICLC (NeoVax) and one post-NeoVax (subject completed the priming phase and two cycles of booster vaccinations) tumor biopsy.

middle: P

middle: M

middle: A


title: Integrating multisector molecular characterization into personalized peptide vaccine design for patients with newly diagnosed glioblastoma.

type: Expression profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Tlr9 deficiency in B cells leads to obesity by modulating the immune system and gut microbiota

status: Public on Apr 11 2024

city: New Haven

first: Pai

last: Wang

address: 333 cedar

institute: Yale University

summary: We generate B-cell-specific Tlr9-deficient (Tlr9fl/fl/Cd19Cre+/-, KO) B6 mice and model obesity using a high-fat diet. Compared with control mice, B cell Tlr9-deficient mice exhibited increased weight gain, impaired glucose and insulin tolerance, reduced IL-10-producing B cells and increased inflammation in fat tissues. Tlr9 deficiency affected B cell differentiation, immunoglobulin levels, and T cell subsets. Furthermore, altered gut microbiota in B-cell-specific Tlr9-deficient mice led to a pro-inflammatory status in gut associated lymphoid tissues and glucose metabolism dysregulation. Using 16S rRNA sequencing we report altered gut microbial communities in the KO mice. Indeed, a reduction in Lachnospiraceae, may play a key role in the observed metabolic phenotypes in KO mice. Also, We identify an important network involving Tlr9, Irf4 and Il-10.

overall_design: We generate B-cell-specific Tlr9-deficient (Tlr9fl/fl/Cd19Cre+/-, KO) B6 mice and model obesity using a high-fat diet. Compare the body weight and immune phenotype then the gut microbiota between the KO mice and Ctr mice. Then use germ free mice and Rag1-/- mice to confirm the function of Tlr9 deficiency on B cells.Finally we would like to compare the effect of Tlr9 deficiency in B cells on molecular level, then we did RNA-seq.

first: Xin

last: Yang


title: Rapid PTEFb-dependent transcriptional reorganization underpins the glioma adaptive response to radiotherapy

city: AURORA

first: Etienne

last: Danis

state: CO

department: Cancer Center

institute: University of Colorado, Denver - Anschutz Medical Campus

type: Genome binding/occupancy profiling by high throughput sequencing


title: Extreme structural heterogeneity rewires glioblastoma chromosomes to sustain patient-specific transcriptional programs [WGS]

city: Göttingen

first: Argyris

last: Papantonis

state: Goettingen

address: Robert-Koch-Str. 40

department: Institute of Pathology

country: Germany

institute: University Medical Center Göttingen

summary: Glioblastoma multiforme encompasses a range of brain malignancies marked by phenotypic and transcriptional heterogeneity, which is thought to render these tumors aggressive, resistant to therapy, and inevitably recurrent. However, little is known about how the spatial organization of glioblastoma genomes underlies this heterogeneity and its effects. We compiled a cohort of 28 patient-derived glioblastoma stem-like lines (GSCs) known to reflect the properties of their tumor-of-origin; six of these came from primary-relapse tumor pairs in the same patient. We generated and analyzed kbp-resolution whole-genome chromosome conformation capture (Hi-C) data from all GSCs to systematically map >3,100 structural variants (SVs) and >6,300 neoloops. By combining Hi-C, histone modification, and gene expression data with 3D chromatin folding simulations, we explain how the pervasive, uneven, and idiosyncratic occurrence of neoloops sustains tumor-specific transcriptional programs via the formation of new enhancer-promoter contacts. We also exemplify how neoloops, albeit scarcely recurring, can help us infer potential patient-specific vulnerabilities. Thus, our data serve as a resource for dissecting glioblastoma biology and heterogeneity, as well as for informing therapeutic approaches.

overall_design:
Hi-C and RNA-seq are applied to 28 low passage IDH-wild type GSC samples, including pairs of GSCs from primary and recurrent tumors from 3 patients. Cut&tag for H3K27me3 and CTCF are applied to 10 samples and 5 samples, respectively.
WGS for one sample

>>>Raw data are not available due to patient privacy concerns.<<<

first: Ting

last: Xie

first: Adi

last: Danieli-Mackay

first: Mariachiara

last: Buccarelli

first: Q

middle: G

last: D’Alessandris

first: Ioanna

last: Papadionysiou

first: Nadine

last: Übelmesser

first: Liverana

last: Lauretti

first: Lucia

last: Ricci-Vitiani


title: Rapid PTEFb-dependent transcriptional reorganization underpins the glioma adaptive response to radiotherapy [ATAC-Seq]

summary: Using a combination of ChIP-, ATAC-, RNA-seq, and CUT&RUN in model systems of diffuse intrinsic pontine glioma and other pediatric high-grade gliomas, we observe a rapid genome-wide reorganization of active chromatin within hours of exposure to therapeutic ionizing radiation. This redistribution facilitates PTEFb-mediated pause-release of Pol II to drive nascent transcriptional induction of canonical DNA damage response programs. Concurrent inhibition of PTEFb imparts a transcription elongation defect, disrupting this chromatin reorganization and blunting transcriptional induction. Functionally, concurrent PTEFb inhibition abrogates key adaptive programs such as DNA damage repair and cell cycle regulation. This combination demonstrates a potent, synergistic therapeutic potential agnostic of glioma subtype, leading to a marked induction of tumor cell apoptosis and prolongation of xenograft survival.

overall_design: ATAC-seq was conducted in the SU-DIPG4 cell line after 6 Gy radiation in the presence or absence of AZD4573.

first: Lays

last: Martin Sobral

first: Faye

last: Walker

first: Nathan

last: Dahl


title: Nucleoporin Nup358 drives the differentiation of myeloid-biased multipotent progenitors by modulating HDAC3 nuclear translocation

city: La Jolla

first: Maximiliano

last: D'Angelo

state: CA

address: 10901 North Torrey Pines Road

department: Cell and Molecular Biology of Cancer Program, NCI-Designated Cancer Center

institute: Sanford Burnham Prebys Medical Discovery Institute

summary: We report that Nup358, a nucleoporin linked to acute myeloid leukemia and myeloproliferative neoplasms, is required for the developmental progression of early myeloid progenitors. We found that loss of Nup358 in mice is associated with the accumulation of myeloid-primed multipotent progenitors (MPPs) in bone marrow and the loss of myeloid-committed progenitors and mature myeloid cells. Accumulated MPPs in Nup358 knockout mice are greatly restricted to megakaryocyte/erythrocyte biased MPP2 cells and fail to progress into committed myeloid progenitors.

overall_design: Bone marrow cells were stained for flow cytometry with Lin antibody cocktail and with antibodies against Sca1 and cKit  and gene expression analyzed using 10X Genomic scRNA-seq protocol.

first: Valeria

last: Guglielmi

last: D’Angelo


title: Elevated expression of wildtype RhoC promotes ErbB2- and Pik3ca-induced mammary tumor formation

city: Toronto

first: Sean

last: Egan

state: Ontario

address: 686 Bay St.

country: Canada

institute: Hospital for Sick Children

summary: Copy number gains in genes coding for Rho activating exchange factors as well as losses affecting genes coding for RhoGAP proteins are common in breast cancer, suggesting that elevated Rho signaling may play an important role. Extra copies and overexpression of RhoC also occur, although a role for RhoC overexpression in driving tumor formation has not been assessed in vivo. To this end, we report on the development of a Rosa26 (R26)-targeted Cre-conditional RhoC overexpression mouse (R26-RhoC). This mouse was crossed to two models for ERBB2/NEU+ breast cancer: one based on expression of an oncogenic ErbB2/Neu cDNA downstream of the endogenous ErbB2 promoter (FloxNeoNeuNT), the other, a metastatic model that is based on high-level expression from MMTV regulatory elements (NIC). RhoC overexpression dramatically enhanced mammary tumor formation in FloxNeoNeuNT mice but showed a more subtle effect in the NIC line, which forms multiple mammary tumors after a very short latency. Many mammary tumors that form in FloxNeoNeuNT mice show selection for increased NeuNT expression linked to high-level amplification of the ErbB2/NeuNT locus. In contrast, NeuNT overexpression did not require high-level amplification of ErbB2/NeuNT in RhoC-FloxNeoNeuNT lesions. RhoC overexpression also enhanced mammary tumor formation in a model for breast cancer induced by Pik3ca(H1047R). The transforming effect of RhoC was associated with epithelial/mesenchymal transition (EMT) in ErbB2/NeuNT and Pik3ca-H1047R systems. Thus, RhoC copy number gains with resultant overexpression contribute to breast tumor formation and/or progression.

overall_design: To identify transcriptional changes linked to RhoC-mediated accelerated mammary tumor formation, we performed bulk RNA-sequencing on mammary tumors from R26(RhoC/+);FloxNeoNeuNT;MMTV-CreNLST and control (FloxNeoNeuNT;MMTV-CreNLST and Gata4(loxP/+);FloxNeoNeuNT;MMTV-CreNLST) cohorts. Differential gene expression analysis and pathway enrichment analyses were then performed.

first: Nandini

last: Raghuram

first: Eda

middle: I

last: Temel

first: Toshihiro

last: Kawamata

first: Katelyn

middle: J

last: Kozma

first: Amanda

first: Wei

last: Wang

first: Jessica

middle: R

last: Adams

first: William

last: Muller

last: EGAN


title: Organization of Corticocortical and Thalamocortical Top-down Inputs in the Primary Visual Cortex

city: Shanghai

first: Guofen

last: Ma

state: Shanghai

address: 280 Chongqing South Road,Huangpu District,

department: School of Medicine

country: China

institute: Shanghai Jiao Tong University

summary: Unified visual perception relies on the integration of bottom-up and top-down inputs in the primary visual cortex (V1), with top-down inputs known to provide behavior-related modulation on visual processing. However, the organization of top-down inputs in V1 remains unclear. Here, using optogenetics-assisted circuit mapping, we characterized how multiple top-down inputs from higher-order cortical and thalamic areas engage excitatory and inhibitory neurons in V1. Systematic layer- and cell-type-specific profiling of the innervation properties of top-down inputs ultimately revealed that each top-down input employs a unique laminar profile when innervating V1. These profiles partially overlap in superficial layers, bypass layer 4 (L4), and clearly segregate upon reaching deep layers. Specifically, inputs from the medial secondary visual cortex (V2M) and anterior cingulate cortex (ACA) preferentially activate L6 neurons, while inputs from the ventrolateral orbitofrontal cortex (ORBvl) and lateral posterior thalamic nucleus (LP) activate L5 neurons. Having defined the inputs, we conducted independent optogenetic activation studies and discovered that ORBvl and LP inputs selectively activate two types of pyramidal neurons (Pyrs) in L5: Pyr <-- ORBvl and Pyr <-- LP neurons, each with specific electrophysiological properties and gene expression profiles. Retrograde mapping subsequently revealed that Pyr <-- ORBvl neurons preferentially innervate subcortical areas and Pyr <-- LP neurons innervate cortical areas, indicating parallel processing of ORBvl and LP inputs in Pyr-type-specific subnetworks. Further, we found mutual inhibition between these two subnetworks, as LP inputs indirectly inhibit Pyr <-- ORBvl neurons and ORBvl inputs inhibit Pyr <-- LP neurons through local inhibitory neurons. Our study thus deepens understanding of the neuronal mechanisms involved in top-down modulation of visual processing by providing a valuable resource characterizing the layer- and cell-type-specific organization of top-down inputs in V1 and by revealing that L5 Pyr-type-specific subnetworks engage in parallel processing of corticocortical and thalamocortical top-down inputs.

overall_design: To acquire the eletropysiological and transcriptomical profiles of Pyr <-- ORBvl and Pyr <-- LP  neuron subtypes, we applied patch-seq to 91 Pyr neurons. After classifying the Pyr subtypes based on their electrophysiological properties, we then performed gene expression analysis and identified the differentially expressed genes in Pyr <-- ORBvl and Pyr <-- LP  neuron subtypes.

first: Jiahe

last: Zhang

first: Siyu


title: Dolutegravir induces FOLR1 expression during brain organoid development.

status: Public on Apr 10 2024

city: Houston

first: Robert

last: Cabrera

state: TX

address: 1 Baylor Plaza

department: CPEH

institute: Baylor College of Medicine

summary: The brain and spinal cord are formed during neurulation, a developmental process that occurs during the first month of pregnancy and can be modified by low serum levels of folic acid (FA), environmental factors, or genetic predispositions. In 2018, data from a surveillance study in Botswana, a country with one of the highest incidences of human immunodeficiency virus (HIV) and lacking mandatory food folate fortification programs, revealed an increased risk of neural tube defects (NTDs) among newborns whose mothers were taking Dolutegravir (DTG) during the first trimester of pregnancy. Subsequently, the World Health Organization and the U.S. Food and Drug Administration issued recommendations and guidelines emphasizing the potential risks associated with the use of DTG-based antiretroviral therapies during pregnancy. To elucidate the potential mechanisms underlying the DTG-induced NTDs, we sought to employ a rapid experimental system to test specific hypotheses.  Brain organoids are three-dimensional cellular structures that can be used to study early neural development, and to assess the potential neurotoxicity of drugs and other compounds. To understand the development of brain organoids in the presence of DTG, we analyzed gene expression from RNA sequencing together with biomechanical information from Optical Coherence Tomography, Optical Coherence Elastography and Brillouin microscopy. Our sequencing data indicates that DTG induces the expression of folate receptor (FOLR1) and modulates the expression of genes required for neurogenesis. The Brillouin frequency shift observed at the surface of DTG-exposed brain organoids indicates an increase in superficial tissue stiffness, while reverberant Optical Coherence Elastography measurements indicate decreased organoid volumes and an internal stiffness decrease.

overall_design: One biological replicate was analyzed for each of 5 conditions (one control; four treatments)

first: Daniel

last: Kraushaar

first: Carlo

last: Donato Caiaffa


title: An amphioxus neurula stage cell atlas supports a complex scenario for the emergence of vertebrate head mesoderm

city: Barcelona

first: Arnau

last: Sebe-Pedros

address: Dr. Aiguader 88

department: Systems Biology

country: Spain

institute: Centre for Genomic Regulation

summary: We performed single-cell transcriptome analysis (using MARS-seq) and accessible chromatin profiling (ATAC-seq) in Branchiostoma lanceolatum embryos (21 hours post-fertilization)

overall_design: Embryos were dissociated and pooled before sorting single-cells for scRNAseq.

first: Xavier

last: Grau-Bove

first: Stéphanie

last: Bertrand

first: Hector

last: Escriva


title: The  transpeptidase sortase A, binds nucleic acids, which can be used and to mediates their cell labeling

city: Hangzhou

first: Lijia

state: Zhejiang

address: Shilongshan Road No.18, Cloud Town, Xihu District

department: School of Life Sciences

institute: Westlake university

summary: We found the transpeptidase Sortase A also binds nucleic acids and mediates mammalian cell labeling. We confiremed the labeling reaction  using . Furthermore, a variety of additional classes of wild-type sortase and engineered mutants of sortase A exhibited the ability to label the surface of mammalian cells with nucleic acids. Heparin on the cell surface appears to be involved, and sortase might serve as an anchor for the nucleic acids. Interestingly, the bacterial sortases of Staphylococcus aureus spontaneously bind extracellular oligonucleotides, which seems to be related to the involvement of sortase and the participation of extracellular nucleic acids in the formation of bacterial biofilms. Finally, we utilized the cell-labeling capability of mgSrtA to enable highly efficient and ready-to-use multiplexed cell labeling for single-cell RNA-seq (scRNA-seq).

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: In SMART-seq experiment, cells were labeled with oligos at first and the oligo were captured by SMART-seq kit. In CRISPR screening experiment, cells were labeled with the oligo modified by fluorescents and sorted by flow cytometry before prepared library. In scRNA-seq experiment, cells from different samples were labeled separately and mixed before loaded to the 10× microfluidics lane.

first: Yingzheng

last: Liu

first: Zhike

last: Lu

first: Panfeng

last: Wu


title: Angiogenesis is uncoupled from osteogenesis during calvarial bone regeneration

city: Muenster

first: Hyun-Woo

last: Jeong

state: NRW

department: Dept. Tissue Morphogenesis

institute: Max-Planck-Institute for Molecular Biomedicine

summary: Gene expression profile at single cell level of calvarial bone stromal cells after drill hole lesion injury (1 mm) at post-lesion day 14 (PLD14) from injured calvarial bones and corresponding unijured controls. Adult wild type animals and endothelial-specific Fbxw7 mutants to study Notch gain-of-function were used.

overall_design: Bone stromal cell preparation: Skulls with calvarial lesions (diameter: 1 mm) at PLD14 and their corresponding age-matched controls were harvested, cut into small pieces and digested with collagenase. Single cell suspensions were lineage depleted to enrich bone stromal cells.

first: M

last: Bixel

first: Kishor

middle: K

last: Sivaraj

first: Kai

last: Kruse

first: Ralf

middle: H


title: A human STAT3 gain of function variant drives Th17 expansion and IL-22 dependent skin inflammation in a model of psoriasiform dermatitis

status: Public on Apr 09 2024

city: SAINT LOUIS

first: Megan

last: Cooper

state: MISSOURI

address: MPRB, 6th floor, 4905 Childrens Place

department: Pediatrics

institute: Washington University in St Louis

summary: Germline gain-of-function (GOF) variants in STAT3 cause early-onset poly-autoimmunity and immune dysregulation. STAT3 is a pleiotropic transcription factor that affects the induction and regulation of immune responses, with diverse effects on the immune response. Using a mouse model of STAT3 GOF (p.G421R), we observed spontaneous and imiquimod (IMQ)-induced skin inflammation with increased cell-intrinsic local Th17 responses. CD4+ T cells were required and sufficient to drive skin inflammation, and upregulated Il22 expression in expanded clones. However, certain aspects of disease, including epidermal thickness, required the presence of STAT3 GOF in epithelial cells. Treatment with a JAK inhibitor improved skin disease, without affecting local Th17 recruitment and cytokine production. Collectively, these data support a role for a Th17 response and in the development of organ-specific immune dysregulation in STAT3 GOF, and also suggest that the presence of STAT3 GOF in tissues is important for disease and can be targeted with JAK inhibition.

overall_design: We examined transcriptional differences and clonal expansion in T cells of STAT3 GOF mice following imiquimod (IMQ) treatment. Bulk CD3+ T cells were isolated from cervical lymph nodes of untreated and IMQ-treated WT and STAT3 GOF mice. T cells were labeled with CITE-seq antibodies. We then generated 5’ GEX v2 libraries for scRNA sequencing, dual indexed with VDJ and surface immune receptor sequencing.

first: Kelsey

last: Toth

first: Erica

last: Schmitt

first: Ana

last: Kolicheski


title: Next Generation Sequencing Facilitates Quantitative Analysis of distal part of the auricle from three Prrx1-CreERT;Bmpr1af/f and age- and gender-matched control mice.

city: shanghai

first: hongshang

last: chu

address: 800 Dongchuan RD. Minhang District, Shanghai, China

summary: We had one goal to perform this study: Analysis of distal part of the auricle from three Prrx1-CreERT;Bmpr1af/f and age- and gender-matched control mice.

overall_design: Comparison of distal part of the auricle from three Prrx1-CreERT;Bmpr1af/f and age- and gender-matched control mice at adult age.

first: Ruichen

last: Yang

first: Hongshang

last: Chu


title: RNA-seq of intestinal epithelia of Candida albicans treated mice

city: Beijing

first: Chuan

last: Ye

address: 38 Xueyuan Road, Haidian District

institute: Peking University

summary: Purpose: To explore the influence of Candida albicans treatment in mice intestinal epithelia. Method: 108 CFU C. albicans or PBS was gavaged to Apoe-/- mice every 3 days for 2 weeks. mRNA profiles were generated by deep sequencing.Results: Several signaling pathways were changed after C. albicans treatment, including response to hypoxia pathway.

overall_design: Expression profiling by RNA-seq, profiling of intestinal epithelia after C. albicans treatment.

first: Xuemei

last: Wang

first: Changtao

last: Jiang

first: Yanli

last: Pang


title: Activation of Nrf2/HO-1 signaling pathway exacerbates cholestatic liver injury

status: Public on Apr 08 2024

city: Zunyi

first: Shaoyu

last: Zhou

state: Guizhou

address: Xinpu Campus, No.6 West Xuefu Road, Xinpu District, Zunyi City, Guizhou, China

institute: Zunyi Medical University

summary: Nrf2 antioxidant signaling is involved in liver protection, but this generalization overlooks conflicting studies indicating that Nrf2 effects are not necessarily hepatoprotective. The role of Nrf2/HO-1 in cholestatic liver injury (CLI) remains poorly defined. Here, we report that Nrf2/HO-1 activation exacerbates liver injury rather than exerts a protective effect in CLI. Inhibiting HO-1 or ameliorating bilirubin transport alleviates liver injury in CLI models. Nrf2 knockout confers hepatoprotection in CLI mice, whereas in non-CLI mice, Nrf2 knockout aggravates liver damage. In the CLI setting, oxidative stress activates Nrf2/HO-1, leads to bilirubin accumulation, and impairs mitochondrial function. High levels of bilirubin reciprocally upregulate the activation of Nrf2 and HO-1, while antioxidant and mitochondria-targeted SOD2 overexpression attenuate the toxicity of bilirubin. Additionally, the expression of Nrf2 and HO-1 is significantly elevated in serum of patients with CLI. These results reveal an unrecognized function of Nrf2 signaling in exacerbating liver injury in cholestatic disease.

overall_design: To investigate the effects of HO-1 in cholestatic liver injury, we performed transcriptome sequencing (RNA seq) analysis on mouse liver tissues treated with OA and ZnPP+OA.

first: Yuanfu

last: Lu

first: Yi

first: Xiaolong

last: Fu


title: Molecular Basis for Differential Igk Versus Igh V(D)J Joining Mechanisms [3C-HTGTS data]

city: Boston

first: Frederick

last: Alt

state: MA

address: 1 Blackfan Circle

department: Program in Cellular and Molecular Medicine

institute: Boston Children's Hospital / Harvard Medical School

summary: In developing B lymphocytes, V(D)J recombination assembles IgH and Igk variable region exons from hundreds of gene segments clustered across mega-base Igh and Igk loci. V, D, and J segments are flanked by conserved recombination signal sequences (RSSs) that target RAG endonuclease. RAG orchestrates Igh V(D)J recombination upon capturing a JH-RSS within the JH-RSS-based recombination center (RC). JH-RSS orientation programs RAG to scan upstream D- and VH-containing chromatin linearly presented by cohesin-mediated loop extrusion. During Igh scanning, RAG robustly utilizes only D- or VH-RSSs in convergent ("deletional") orientation with JH-RSSs. However, for Vk-to-Jk joining, RAG utilizes Vk-RSSs from deletional- and inversional-oriented clusters, inconsistent with linear scanning. Here, we elucidate the Vk-to-Jk joining mechanism. Igk undergoes robust primary and secondary rearrangements, which confounds scanning assays. Thus, we engineered cells to undergo only primary Vk-to-Jk rearrangements and found that RAG-scanning from the primary Jk-RC terminates just 8kb upstream within the CTCF-Site-based Sis element. While Sis and the Jk-RC barely interacted with the Vk locus, the CTCF-site-based Cer element, 4kb upstream of Sis, interacted with various loop-extrusion impediments across the locus.  Like VH-locus inversion, DJH inversion abrogated VH-to-DJH joining; yet Vk-locus or Jk inversion allowed robust Vk-to-Jk joining. Together, these experiments implicated loop extrusion in bringing Vks near Cer for short-range diffusion-mediated capture by RC-based RAG. To elucidate key mechanistic elements for diffusional V(D)J recombination in Igk versus Igh, we assayed Vk-to-JH and D-to-Jk rearrangements in hybrid Igh-Igk loci generated by targeted chromosomal translocations, and pinpointed remarkably strong Vk and Jk RSSs. Indeed, RSS replacements in hybrid or normal Igk and Igh loci confirmed ability of Igk versus Igh RSSs to promote robust diffusional joining. We propose that Igk evolved strong RSSs to mediate diffusional Vk-to-Jk joining; while Igh evolved weaker RSSs requisite for modulating VH joining by RAG scanning impediments.

overall_design: We performed 3C-HTGTS in BM pre-B cells and v- Abl transformed pro-B cells and its various mutant derivatives to study the differential molecular mechanisms of Igk versus Igh V(D)J recombination.

middle: W

first: Hongli

last: Hu

first: Yiwen

last: Zhang

first: Xiang

last: Li


title: Molecular Basis for Differential Igk Versus Igh V(D)J Joining Mechanisms


title: Effect of Orb2 depletion on mRNA expression in Drosophila larval brains

city: Durham

first: Don

last: Fox

state: NC

address: DUMC Box 3813  C315 LSRC  Duke University

institute: Duke University

summary: Regulation of codon optimality is an increasingly appreciated layer of cell- and tissue-specific protein expression control. Here, we use codon-modified reporters to show that differentiation of Drosophila neural stem cells into neurons enables protein expression from rare-codon-enriched genes. From a candidate screen, we identify the cytoplasmic polyadenylation element binding (CPEB) protein Orb2 as a positive regulator of rare-codon-dependent expression in neurons. Using RNA sequencing, we reveal that Orb2-upregulated mRNAs in the brain with abundant Orb2 binding sites have a rare-codon bias. From these Orb2-regulated mRNAs, we demonstrate that rare-codon enrichment is important for expression control and social behavior function of the metabotropic glutamate receptor (mGluR). Our findings reveal a molecular mechanism by which neural stem cell differentiation shifts genetic code regulation to enable critical mRNA and protein expression.

overall_design: Brains were dissected from larvae expressing inscuteable GAL4 driving white RNAi or orb2 RNAi. Total mRNA was extracted from two replicates of 60 brains each condition and submitted for RNA sequencing.

first: Rebeccah

last: Stewart


title: The effect of miR-200 overexpression in lung cancer-associated fibroblasts

city: Seoul

first: Young-Ho

last: Ahn

address: 25 Magokdong-ro 2-gil, Gangseo-gu

department: College of Medicine

country: South Korea

institute: Ewha Womans University

summary: To investigate the roles of miR-200 family memebrs in the regulation of lung cancer-associated fibroblasts (CAFs), we overexpressed miR-200b/a/429 cluster in murine lung CAFs through lentiviral infection. We then performed gene expression profiling analysis using data obtained from RNA-seq of control and miR-200-overexpressed CAFs.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for control CAFs (CAF-vec) and miR-200-overexpressed CAFs (CAF-200).

first: Inyoung

last: Cheon


title: Translation initiation factor eIF1.2 orchestrates Toxoplasma gondii differentiation by regulating stage-specific gene expression

city: Indianapolis

first: Michael

middle: James

last: Holmes

state: IN

address: 635 Barnhill Drive

department: Pharmacology & Toxicology

institute: Indiana University School of Medicine

summary: Toxoplasma gondii persists in humans by converting from actively replicating acute stage tachyzoites to slow-growing chronic stage bradyzoites. The molecular mechanisms that mediate T. gondii differentiation remain poorly understood. Through a chemical mutagenesis screen, we identified translation initiation factor eIF1.2 as being critical for T. gondii differentiation. The presence of an F97L mutation in eIF1.2 identified in the screen or the complete lack eIF1.2 (∆eIF1.2) markedly impeded bradyzoite cyst formation in culture and in the brains of infected mice. ∆eIF1.2 parasites were defective in the upregulation of bradyzoite differentiation regulators BFD1 and BFD2 during stress-induced differentiation. Conditional overexpression of BFD1 or BFD2 rescued differentiation in ∆eIF1.2 parasites. We further show that eiF1.2 interacted with the yeast 40S ribosome and directed the scanning of a model 5’ untranslated region. Together, our findings imply that eIF1.2 functions by regulating the translation of key differentiation factors necessary to establish T. gondii chronic infection.

overall_design: expression changes at the transcriptional and translational levels compared between parental and eIF1.2 knockout Toxoplasma gondii tachyzoites and 48h alkaline stressed parasites

first: William

last: Sullivan Jr


title: MYCT1 Controls Environmental Sensing in human Hematopoietic Stem Cells.

city: Los Angeles

first: Júlia

last: Aguadé-Gorgorió

state: California

address: 615 Charles E Young Drive South

department: Molecular, Cell and Developmental Biology

institute: University of California Los Angeles

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

overall_design: Refer to individual Series


title: Obesogenic high-fat diet and MYC cooperate to promote lactate accumulation and tumor microenvironment remodeling in prostate cancer

city: Montreal

first: David

last: Labbé

state: QC

address: 1001 Decarie Boulevard

department: Urology

institute: Research Institute of McGill University Center

summary: In this study, we investigated the effect of high-fat diet on prostate cancer progression. We specifically analyzed the metabolome, transcriptome, and tumor microenvironment of MYC transgenic and wild-type mice subjected to high-fat diet (enriched in saturated fat) and control diet for 21 weeks.

overall_design: Comparative gene expression profiling analysis of RNA-seq data from prostates (dorsolateral lobes) collected from MYC-transgenic and wild-type FVB mice fed high-fat diet (HFD) or control diet (CTD) for 21 weeks (1. CTD_WT, 2. HFD_WT, 3. CTD_MYC, HFD_MYC). Five mice/group were analyzed.

first: Nadia

last: Boufaied

first: Giorgia

last: Zadra


title: MYCT1 Controls Environmental Sensing in human Hematopoietic Stem Cells [scRNA-seq]

summary: We identified a novel HSC regulatory gene, MYCT1 (MYC target 1), that is selectively expressed in undifferentiated human HSPC and is critical for human HSC expansion and engraftment. MYCT1 knockdown (KD) in human fetal liver and cord blood (CB) HSPCs impaired expansion ex vivo and engraftment after transplantation, whereas restoring the compromised MYCT1 expression via lentiviral overexpression (OE) in cultured human CB HSPCs improved ex vivo expansion of the most undifferentiated human HSPCs (CD34+CD38-CD90+CD45RA-EPCR+ITGA3+) and enhanced their engraftment ability. We performed single cell RNAseq of uncultured human CB HSPCs, as well as 72 hours after MYCT1 KD and OE to identify MYCT1-regulated programs in HSC and understand the role of MYCT1 in governing human HSPC expansion and engraftment ability. Functional enrichment analysis linked the loss of MYCT1 in HLF+ HSCs to a profound dysregulation of multiple cellular programs essential for HSC stemness, such as oxidative phosphorylation or proteostasis, while MYCT1 OE had the opposite effect and showed closer similarity to uncultured HLF+ HSCs. These data indicate that the loss or gain of MYCT1 expression elicits major effects on programs regulating human HSC functional competence.

overall_design: Single cell RNAseq of sorted human cord blood HSPCs (CD34+CD38-CD90+) either uncultured (day 0) or transduced with control, MYCT1 knockdown, and MYCT1 overexpression lentiviral vectors and resorted (CD34+CD38-CD90+GFP+) 72 hours after transduction (day 4).

first: Feiyang

last: Ma

first: Hanna

last: Mikkola


title: MYCT1 Controls Environmental Sensing in human Hematopoietic Stem Cells [RNA-seq]

summary: We identified a novel HSC (hematopoietic stem cell) regulatory gene, MYCT1 (MYC target 1), that is selectively expressed in undifferentiated human HSPC and is critical for human HSC expansion and engraftment. To define the molecular function of MYCT1 we performed cell fractionation, immunofluorescence, and immunoprecipitation coupled with high sensitivity mass spectrometry of V5-tagged MYCT1 in KG1 hematopoietic cells and human endothelial cells (E4-immortalized HUVEC, E4EC (Butler et al. 2012)). We found that MYCT1 localizes in the endosomal membrane and interacts with vesicle trafficking regulators and signalling machinery essential for HSC and endothelial cell function. We performed RNAseq to asses the expression of the identified MYCT1 interactors in KG1 and E4EC cell lines.

overall_design: RNAseq of KG1 (acute myeloid leukemia) and E4EC (E4-immortalized HUVEC) cell lines.


title: RNA Sequencing Analysis of bone marrow derived macrophages from Wild Type and TIGAR-/- mice

status: Public on Apr 06 2024

city: Nanjing

first: dong

middle: dong

last: wang

state: jiangsu

address: 101Longmian Avenue, Jiangning District,

institute: Nanjing Medical University

summary: We identified that global TIGAR deficiency reduced the AS plaque formation and the systemic inflammatory response. The purpose of this experiment was to identify how TIGAR induces macrophage inflammation.We then performed gene expression profiling analysis using data obtained from RNA-seq of bone marrow derived macrophages from WT and TIGAR KO mice stimulated with LPS.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for bone marrow derived macrophages from WT and TIGAR KO mice stimulated with LPS.

first: Dongdong

last: Wang

first: Yanxia

first: Bin

last: Jiang

first: Chenyu


title: Colorado potato beetle transcriptional response to aegerolysin treatment

city: Ljubljana

first: Marko

last: Petek

address: Večna pot 111

department: Department of Biotechnology and Systems Biology

country: Slovenia

institute: National Institute of Biology

summary: Protein complexes of aegerolysins pleurotolysin A2 (PlyA2) and pleurotolysin B (PlyB) from oyster mushrooms Pleurotus sp. display targeted toxicity against Colorado potato beetle (CPB; Leptinotarsa decemlineata) larvae. This selective toxicity is achieved through aegerolysins' interaction with insect-specific membrane sphingolipids. This study explores the potential adaptive response of CPB larvae to the aegerolysin complex.

overall_design: Leptinotarsa decemlineata larvae fed on potato leaves soaked in PlyA2/PlyB/buffer solution vs control larvae fed on leaves soaked in buffer; two sampling time points - 1 and 5 days post treatment

first: Kristina

last: Gruden

first: Primož

last: Žigon

first: Jaka

last: Razinger

first: Katja

last: Stare

first: Karmen

last: Pogačar

first: Maja

last: Grundner

first: Anna

last: Coll

first: Anastasija

last: Panevska

last: Sepčić


title: Fatty acid beta oxidation-dependent H3K18ac in the Drosophila melanogaster wing imaginal disc (CUT&RUN)

city: Heidelberg

first: Philipp

last: Willnow

address: Im Neuenheimer Feld 280

institute: Deutsches Krebsforschungszentrum (DKFZ)

summary: The  distribtion of H3K18ac in the fruit fly genome was analyzed in wing imaginal discs treated for 2 h with etomoxir and compared to control discs.

overall_design: H3K18ac in wing imaginal discs treated for 2 h with etomoxir and compared to control discs

first: Aurelio

last: Teleman


title: DEAD-box helicase 17 (DDX17) protects cardiac function by promoting mitochondrial homeostasis in heart failure

status: Public on Apr 07 2024

city: Shanghai

first: Yuxuan

last: Zheng

address: 825 Zhangheng Rd.

institute: Fudan University

type: Methylation profiling by high throughput sequencing

summary: DEAD box 17 (DDX17) is a typical member of the DEAD box family with transcriptional cofactor activity. Although DDX17 is abundantly expressed in the myocardium, its role in the heart is not fully understood. We generated cardiomyocyte-specific Ddx17-knockout mice and cardiomyocyte-specific Ddx17 transgenic mice and explored the function of DDX17 using various cardiomyocyte injury and heart failure (HF) models. We also validated the correlation between DDX17 expression and cardiac function in myocardial biopsy samples from HF patients. DDX17 was downregulated in myocardial samples from HF mouse models and cardiomyocyte injury models. We found that the cardiomyocyte-specific knockout of DDX17 promotes autophagic flux blockage and cardiomyocyte apoptosis in pathological conditions, resulting in progressive heart dysfunction, thereby leading to maladaptive remodeling and progression of HF. The replenishment of DDX17 in cardiomyocytes protected heart function in pathological conditions. Further studies showed that DDX17 can bind to the transcriptional repressor BCL6 and inhibit the expression of the mitochondrial fission protein DRP1. When DDX17 expression was decreased, the transcriptional repression of BCL6 was reduced, resulting in increased DRP1 expression and mitochondrial fission, which caused autophagy flux blockage and apoptosis in cardiomyocytes, leading to impaired mitochondrial homeostasis and HF. We also verified the clinical correlation of DDX17 expression with cardiac function and DRP1 expression in endomyocardial biopsies from patients with HF. These findings suggest that DDX17 protects cardiac function by promoting mitochondrial homeostasis through the BCL6-DRP1 pathway during HF.

overall_design: We collected NMVMs (neonatal mouse ventricular myocytes) from two groups of mouse hearts (three biological replications in each group). These two groups were labelled as 'TG-minus (Con)' (which meant the wild-type littermates of Ddx17-transgenic C57BL/6 male mice were used in the research, NTGM1-3) and 'TG-plus (Ddx-17 TG)' (which meant cardiac-specific Ddx17-transgenic overexpression mice generated by standard CRISPR/Cas9 techniques, NTGP1-3). To explore the molecular mechanisms through which DDX17 inhibits mitochondrial fission and promotes mitochondrial fusion, we applied the nucleosome occupancy and methylome sequencing (NOMe-seq) in these samples.


title: The thioredoxin system determines CHK1 inhibitor sensitivity via redox-mediated regulation of ribonucleotide reductase activity

status: Public on Apr 04 2024

city: Columbus

first: Chandra

middle: Bhushan

last: Prasad

state: OH

address: 460 W. 12th Ave. Biomedical Research Tower, Room 460G

department: radiation Oncology

institute: OSU Wexner Medical Center

summary: Checkpoint kinase 1 (CHK1) is critical for cell survival under replication stress (RS). CHK1 inhibitors (CHK1i’s) in combination with chemotherapy have shown promising results in preclinical studies but have displayed minimal efficacy with substantial toxicity in clinical trials. To explore combinatorial strategies that can overcome these limitations, we perform an unbiased high-throughput screen in a non-small cell lung cancer (NSCLC) cell line and identify thioredoxin1 (Trx1), a major component of the mammalian antioxidant-system, as a determinant of CHK1i sensitivity. We establish a role for redox recycling of RRM1, the larger subunit of ribonucleotide reductase (RNR), and a depletion of the deoxynucleotide pool in this Trx1-mediated CHK1i sensitivity. Further, the TrxR inhibitor auranofin, an approved anti-rheumatoid arthritis drug, shows a synergistic interaction with CHK1i via interruption of the deoxynucleotide pool. Together, we show a pharmacological combination to treat NSCLC that relies on a redox regulatory link between the Trx system and mammalian RNR activity.

overall_design: To investigate the biomerker for CHK1i sensitivity in NSCLC, we established H1299 and MCF7 cell lines in which each target gene has been knocked down by shRNA.

first: Zhaojun

last: Qiu

first: Junran

last: Zhang


title: Transcriptome analyses of intestine-specific Slc39a8-KO mice

status: Public on Apr 05 2024

city: Ann Arbor

first: Shigeki

last: Iwase

address: 1241 E. Catherine St.

institute: University of Michigan

summary: Purpose: To identify genes whose expression was altered in the intestinal epithelial cells (IECs) of Slc39a8-KO mice.  Methods: Poly(A) RNA-seq  Results: We identified four genes that showed changes in their expression in the Slc39a8-KO IECs.

overall_design: RNAs were isolated from the illeum and colon of control and intestine-specific Slc39a8-KO mice.

first: Eun-Kyung

last: Choi

first: Luisa

last: Aring

first: Young-Ah

last: Seo


title: RNA transcriptome sequencing analysis of SNU-668 Erastin-resistant cells and SNU-668 parental cells, SNU-484 RSL3-resistant cells and SNU-484 parental cells

first: Ming

middle: zhe

last: Ma

email: mmz666@163.com, mmz666@shca.org.cn

address: Dongan Road 270

department: Department of gastric surgery

institute: Fudan university shanghai cancer center

summary: RNA transcriptome sequencing analysis was performed in SNU-668 Erastin-resistant cells and SNU-668 parental cells, SNU-484 RSL3-resistant cells and SNU-484 parental cells

overall_design: mRNA profiles of  SNU-668 Erastin-resistant cells and SNU-668 parental cells, SNU-484 RSL3-resistant cells and SNU-484 parental cells

first: Mingzhe


title: Gene expression profiling of multiregional primary colorectal cancer tissue samples

city: Oslo

first: Anita

last: Sveen

address: P.O.Box 4953, Nydalen

department: Department of Molecular Oncology

country: Norway

institute: Institute for Cancer Research, Oslo University Hospital

summary: We have analyzed transcriptomic intra-tumor heterogeneity among 2-4 multiregional samples from each of 98 primary colorectal cancers. We investigated the level and prognostic value of intra-tumor heterogeniety of the consensus molecular subtypes, and identified subtypes more robust to heterogeneity based on genes with uniform expression levels across tumor regions.

overall_design: This series consists of 286 multiregional samples of 98 primary colorectal tumors from 96 patients (two patients with two synchronous tumors). All samples were analyzed with Affymetrix Human Transcriptome 2.0 Arrays.

first: Jonas

last: Langerud

first: Ragnhild

last: Lothe


title: Transcriptome map of human temporopolar and 2ary Auditory Anterior cortex

status: Public on Apr 03 2024

city: Lund

first: Ilia

last: Kurochkin

state: Skane

address: BMC A12, Sölvegatan 17

country: Sweden

institute: Lund University

summary: RNA sequencing was performed for 35 human brain regions representing a subset of 75 brain regions used in lipid analysis in the same four human individuals. Combining lipid composition and mRNA expression data highlighted functional connectivity association.

overall_design:
8 polyA-selected RNA-seq samples were obtained from 2 brain regions from four adult cognitively healthy human individuals.
This submission includes only 8 of the healthy samples because the other 132 were submitted previously in GSE127898 (GSM3678700-GSM3678831).

first: I

first: E

last: Khrameeva

first: P

last: Khaitovich


title: Molecular Basis for Differential Igk Versus Igh V(D)J Joining Mechanisms [HTGTS-V(D)J-seq data]

first: Frederick

last: Alt

summary: In developing B lymphocytes, V(D)J recombination assembles IgH and Igk variable region exons from hundreds of gene segments clustered across mega-base Igh and Igk loci. V, D, and J segments are flanked by conserved recombination signal sequences (RSSs) that target RAG endonuclease. RAG orchestrates Igh V(D)J recombination upon capturing a JH-RSS within the JH-RSS-based recombination center (RC). JH-RSS orientation programs RAG to scan upstream D- and VH-containing chromatin linearly presented by cohesin-mediated loop extrusion. During Igh scanning, RAG robustly utilizes only D- or VH-RSSs in convergent ("deletional") orientation with JH-RSSs. However, for Vk-to-Jk joining, RAG utilizes Vk-RSSs from deletional- and inversional-oriented clusters, inconsistent with linear scanning. Here, we elucidate the Vk-to-Jk joining mechanism. Igk undergoes robust primary and secondary rearrangements, which confounds scanning assays. Thus, we engineered cells to undergo only primary Vk-to-Jk rearrangements and found that RAG-scanning from the primary Jk-RC terminates just 8kb upstream within the CTCF-Site-based Sis element. While Sis and the Jk-RC barely interacted with the Vk locus, the CTCF-site-based Cer element, 4kb upstream of Sis, interacted with various loop-extrusion impediments across the locus.  Like VH-locus inversion, DJH inversion abrogated VH-to-DJH joining; yet Vk-locus or Jk inversion allowed robust Vk-to-Jk joining. Together, these experiments implicated loop extrusion in bringing Vks near Cer for short-range diffusion-mediated capture by RC-based RAG. To elucidate key mechanistic elements for diffusional V(D)J recombination in Igk versus Igh, we assayed Vk-to-JH and D-to-Jk rearrangements in hybrid Igh-Igk loci generated by targeted chromosomal translocations, and pinpointed remarkably strong Vk and Jk RSSs. Indeed, RSS replacements in hybrid or normal Igk and Igh loci confirmed ability of Igk versus Igh RSSs to promote robust diffusional joining. We propose that Igk evolved strong RSSs to mediate diffusional Vk-to-Jk joining; while Igh evolved weaker RSSs requisite for modulating VH joining by RAG scanning impediments.

overall_design: We performed HTGTS-V(D)J-seq in BM pre-B cells and v- Abl transformed pro-B cells and its various mutant derivatives to study the differential molecular mechanisms of Igk versus Igh V(D)J recombination.

first: Hongli

last: Hu

first: Yiwen

first: Xiang

last: Li


title: A set of Sox2-dependent OSN enhancers mediates the immediate response to neural induction and default neural differentiation of embryonic stem cells

city: Berlin

first: Pavel

last: Tsaytler

address: Ihnestrasse  63-73

institute: MPI Molecular Genetics

summary: Sox2 is a transcription factor involved in the regulatory network maintaining the pluripotency of embryonic stem cells in culture as well as in early embryos. In addition, Sox2 plays a pivotal role in neural stem cell formation and neurogenesis. How Sox2 can serve both processes has remained elusive. Here we identified a set of Sox2-dependent neural associated enhancers (SONAE) activated immediately following neural induction and required in neural gene activation. SONAE are a distinct subgroup (1898) of 8531 Oct4/Sox2/Nanog (OSN) bound enhancers, characterized by enhanced Sox2 binding and chromatin accessibility. SONAE activation is triggered by neural induction via Retinoic Acid treatment of wild type cells and is antagonized by mesodermal transcription factors repressing Sox2 following mesoderm induction. Neural differentiation is also set off by default in Smad4 ablated cells resistant to mesodermal induction.  Our data provide mechanistic insight into the regulation of early neural versus mesodermal induction in ESCs and embryos.

overall_design: Transcriptomes (RNA-Seq) and chromatin accessibility (ATAC-Seq) from non-differentiated or neural differentiated (RA) Sox2 wild-type (DMSO) and Sox2 knock-out (dTAG13) mouse ESCs.

first: Gaby

last: Blaess

first: Manuela

last: Scholze-Wittler

first: Frederic

last: Koch

first: Bernhard

last: Herrmann


title: miRNA expression profile of human GMSC-derived EVs

status: Public on Apr 02 2024

first: Jingrong

last: Chen

address: 768 Zhongshan Middle Road, Shanghai

institute: Shanghai Jiaotong University

type: Non-coding RNA profiling by high throughput sequencing

summary: The transcriptome of extracellular vesicles (EVs) from human gingival mesenchymal stem cells (GMSC) hasn't been compenhensively profiled. We performed the RNA-SEQ transcriptomic analysis of EVs from GMSC or Fibroblasts. Guman gingiva samples  were collected following routine dental procedures. The primary cultured human dermal fibroblasts were used as a control since them share similar morphologies but lack the functional activities of GMSCs. Primary human dermal fibroblasts were isolated from the foreskin dermis of children aged between 6 and 8 years who underwent surgery.

overall_design: Using small RNA-seq to investigate the miRNA expression of human gingival mesenchymal stem cells-derived EVs, fibroblast-derived EVs as the control. GMSCs or Fibroblasts that were freshly prepared and derived from passages three to five.


title: Nuclear factor 90 isoform of vascular smooth muscle cells accelerates diabetic arteriosclerotic calcification by mediating FBXW7-AGER1-AGEs axis

city: Jinan

first: Fei

last: Xie

address: 107 Wenhua West Road, Lixia District, Jinan City, Shandong Province

institute: Qilu Hospital of Shandong University

summary: Hyperglycemia accelerates calcification of atherosclerotic plaques in diabetic patients, and the prolonged accumulation of advanced glycation end products (AGEs) induced by hyperglycemia may be closely related to the pathogenesis of aortic calcification. However, the mechanisms underlying this association remain unclear. In the current study, we investigated the role of vascular smooth muscle cell nuclear factor 90/110 (NF90/110) in mediating AGEs accumulation and accelerating diabetic atherosclerotic calcification. Using vascular smooth muscle cells (VSMCs), human samples, and mouse models, we found that hyperglycemia-mediated AGEs markedly increased VSMC NF90/110 activation both in human and mouse atherosclerotic calcified tissues with diabetes. Silencing of NF90/110 in vitro and genetic deletion of VSMC NF90/110 in mice decreased obviously AGEs-induced arteriosclerotic calcification. Mechanistically, AGEs increased the activity of NF90, which then enhanced ubiquitination and degradation of AGE receptor 1 (AGER1) by stabilizing the mRNA of E3 ubiquitin ligase, F-box, and WD repeat domain 7 (FBXW7), thus causing the accumulation of more AGEs. Furthermore, AGEs accumulation accelerated diabetic atherosclerotic calcification by inducing VSMC phenotypic changes to osteoblast-like cells, apoptosis, and matrix vesicle release. Collectively, our study demonstrated the effects of VSMC NF90 in mediating the metabolic imbalance of AGEs to accelerate diabetic arteriosclerotic calcification. These novel findings elucidate a pivotal mechanism underlying AGE-induced diabetic atherosclerotic calcification and provide a framework for potential interventions against diabetic vascular complications.

overall_design: To confirm that NF90 interacts with FBXW7 mRNA, RIP assays were performed using NF90 antibodies and IgG in HAVSMCs with or without stimulation by AGE.


title: 3D genomics analysis identifies enhancer hijacking caused by structural alterations and extrachromosomal circular DNA formation [HiChIP]

city: Salt Lake City

first: Xiaoyang

last: Zhang

state: Utah

address: 1950 Circle of Hope Drive

department: Oncological Sciences

institute: University of Utah

summary: Enhancer hijacking, caused by structural alterations, is a common cancer driver event that causes aberrant expression of oncogenes. Unfortunately, enhancer hijacking is difficult to detect due to the complexity of the cancer genome. Here we propose a simple yet robust strategy HAPI (Highly Active Promoter Interactions) to identify and characterize such events by following two rules: 1) oncogenes subject to enhancer hijacking should be potentially highly regulated by enhancers, 2) the hijacked enhancers should contribute an appreciable proportion of an oncogene’s overall enhancer activity. Applying this strategy to HiChIP data we and others generated in 34 cancer cell lines, we identified known enhancer hijacking events and uncovered novel enhancers hijacked by known or potentially novel oncogenes such as CCND1, ETV1, ID4, and NKX2-5, which we validated using CRISPRi assays and RNA-seq analysis. Furthermore, we found that complex enhancer hijacking events connecting genes and enhancers from multiple chromosomal segments are often caused by the formation of extrachromosomal circular DNA (ecDNA). Focusing on ecDNAs harboring the MYC oncogene, one of the most common gene targets of ecDNA, we found that these ecDNAs often stitch additional genes such as CDX2, ERBB2, and NFIB from other chromosomes to the MYC locus. These genes heavily hijack MYC enhancers for their activation, a novel insight into ecDNA biology, suggesting alternative therapeutic targets for MYC ecDNAs. Our study provides an efficient strategy to detect enhancer hijacking events, and more importantly reveals novel mechanisms underlying oncogene activation caused by simple or complex structural alterations.

overall_design: H3K27ac HiChIP assays were performed to map enhancer-promoter interactions in cell lines of different cancer types

first: Katelyn

middle: L

last: Mortenson

first: Katherine

last: Varley


title: 3D genomics analysis identifies enhancer hijacking caused by structural alterations and extrachromosomal circular DNA formation [RNA-Seq]

overall_design: Bulk RNA-seq on cells treated with MYC promoter CRISPRi to assess role of MYC's promoter on enhancer hijacking regulatory network on ecDNA strcutures


title: 3D genomics analysis identifies enhancer hijacking caused by structural alterations and extrachromosomal circular DNA formation [ChIP-Seq]

overall_design: ChIP-seq of H3K27ac in LNCaP cells were performed to measure on-target effects of CRISPRi. d98 and NT1, two guides with no targets in the human genome, serve as negative controls


title: Single cell transcriptomics on zebrafish gliovascular niche

city: New York

first: Caghan

last: Kizil

state: NY

institute: Columbia University Irving Medical Center

summary: To investigate the cell types involved in the gliovascular niche of the zebrafish brain, we performed single cell sequencing on cells sorted from fli1:gfp and her4: GFP transgenic animals in control and amyloid toxicity conditions.

overall_design: Tg(her4.1:dRED) and Tg(fli1a:GFP) lines were injected with Amyloid-beta 42 peptide as descibed (Bhatatrai et al. 2016) and cells were dissociated sorted by FACS as described (Cosacak et al. 2019). The single-cell encapsulation, cDNA synthesis by 10X Genomics. The reads were aligned to zebrafish genom (GRChZ 11) and ensemble version 105 was used for gene annotation and assigning reads to genes. Genomics Kit, and Sequencing done by Illumina.

first: Prabesh

last: Bhattarai

first: Mehmet

last: Cosacak

first: Elanur

last: Yilmaz


title: Expression data of mRNA from twins with tuberculosis infection or not

status: Public on Apr 01 2024

city: Tongliao

first: Batudeligen

last: Batudeligen

address: 536 West Hollin River Street

institute: Affiliated Hospital of Inner Mongolia University for the Nationalities

summary:
To search the genes which were associated with tuberculosis infecion
We used the microarrays to detail the global programme of gene exprssion between with the tuberculosis infecion and without the tuberculosis in monozygous twins

overall_design: Total blood RNA was extracted using the PAXgene RNA Blood Mini Kit (Qiagen, Venlo, The Netherlands), according to the manufacturer’s instructions. The amount of RNA was quantified by the spectrophotometric A260 to A280 ratio of 1.8, whereas quality was assessed using denaturing gel electrophoresis using the Agilent 2100 system (Agilent Technologies, Santa Clara, CA, USA).  Gene expression profiles were analyzed by PrimeView Human Gene Expression Array (Affymetrix, Santa Clara, CA, USA) according to the manufacturer’s instructions.

first: Zhi

last: Liu

first: Batu

last: Deligen


title: Orthogonal proteogenomic approaches identify the druggable PA2G4-MYC axis in 3q26 AML [RNA-Seq]

city: Parma

first: Andrea

last: Gherli

address: Via Gramsci 14

department: Medicine and Surgery

country: Italy

institute: University of Parma

summary: The overexpression of the ecotropic viral integration site-1 gene (EVI1/MECOM) marks the most lethal acute myeloid leukemia (AML) subgroup carrying chromosome 3q26 abnormalities. By taking advantage of the intersectionality of high-throughput cell-based and gene expression screens, we identified selective and pan-histone deacetylase inhibitors (HDACis) as potent repressors of EVI1. To understand the mechanism driving on-target anti-leukemia activity of this compound class, we dissected the expression dynamics of the bone marrow leukemia cells of patients treated with HDACi and reconstituted the EVI1 chromatin-associated co-transcriptional complex merging on the role of proliferation-associated 2G4 protein. PA2G4 overexpression rescues AML cells from the inhibitory effects of HDACis, while genetic and small molecule inhibition of PA2G4 abrogated EVI1 in 3q26 AML cells, including in patient-derived leukemia xenografts. This study positions PA2G4 at the crosstalk of the EVI1 leukemogenic signal for developing new therapeutics and urges the use of HDACi-based combination therapies in patients with 3q26 AML.

overall_design: Comparative gene expression profiling in HNT34 leukemia cell line after genetic knockout with two different shRNA targeting EVI1 gene (#16 and #87) and a control non targeting shRNA (NT)

first: G

last: Roti

first: M

last: Marchesini

first: A

last: Montanaro

first: L

last: Moron dalla Tor


title: Orthogonal proteogenomic approaches identify the druggable PA2G4-MYC axis in 3q26 AML [ChIP-seq]

overall_design: Genome binding/occupancy profiling by high throughput sequencing


title: Orthogonal proteogenomic approaches identify the druggable PA2G4-MYC axis in 3q26 AML [scRNA-Seq]

overall_design: Longitudinal scRNA profiling of PR#002 bone marrow cells before (Pre) and after (Post) four cycles of therapy with Azacitidine and Entinostat.

first: E

last: Simoncini

first: S

last: Colla

first: N

last: Thongon


title: Orthogonal proteogenomic approaches identify the druggable PA2G4-MYC axis in 3q26 AML

city: Meldola

first: Matteo

state: FC

address: via Piero Maroncelli 40

institute: IRCSS Istituto Romagnolo per lo Studio dei Tumori "Dino Amadori" (IRST)

overall_design: Human CD45+ leukemia cells were isolated through cell sorting from PDLX mice model after 4 weeks of treatment with Vehicle, azacitidine (Aza), entinostat (Ent), or a combination of azacitidine and entinostat (Ent_Aza)


title: C57BL/6 mice: skeletal muscle-specific GDE5 (Gpcpd1)-deficient mice (Gde5 skKO mice)

city: Higashi-Hiroshima

first: Yanaka

last: Noriyuki

address: Kagamiyama 1-4-4

department: Graduate School of Biosphere Science

country: Japan

institute: Hiroshima University

summary: Transcriptional profiling of mouse skeletal muscles. The objective of this study is to explore gene expression profiles of skeletal muscle in response to Gpcpd1 (GDE5) deficiency by DNA microarray analysis.

overall_design: Two-condition experiment, gastrocnemius (GAS) muscles of five WT and Gde5 skKO mice at 15 weeks of age

first: Noriyuki

last: Yanaka


first: Anna

department: Department of Medicine and Surgery

overall_design: Human CD45+ leukemia cells were isolated through cell sorting from PDLX mice model after 2 weeks of treatment with WS6 or Vehicle

last: Moron Dalla Tor


title: 3D genomic analysis reveals novel enhancer-hijacking caused by complex structural alterations that drive oncogene overexpression

first: Xiaoyang

last: Zhang

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

overall_design: Refer to individual Series


title: Effects of fat body microRNA miR-305-5p knockdown on brain and fat body gene expression in Apis mellifera

city: Urbana

first: Gene

middle: E.

last: Robinson

state: IL

address: 1206 W. Gregory Dr. M/C 195

department: Carl R. Woese Institute for Genomic Biology

institute: University of Illinois at Urbana-Champaign

summary: RNA sequencing of Apis mellifera abdominal fat body and matched whole brain following a knockdown in fat body ame-miR-305-5p expression

overall_design: mRNA analsys of 5-day-old workers bees 48h post oral antagomir treatment

first: Sarai

last: Stuart

first: Amy

middle: C

last: Cash Ahmed


title: Glucocorticoids-triggered CXCR4 signaling activates the PLC signalosome and promotes tumor resistance in B cell acute lymphoblastic leukemia

city: Rouen

first: Souleymane

last: Abdoul-Azize

state: State...

address: 22 Bd Gambetta

country: France

institute: Rouen normandy Univ

summary:
Approximately 20-25% of patients with acute lymphoblastic leukemia (ALL) relapse after initial chemotherapy and become resistant to glucocorticoids (GC). Although GC signaling act on well-defined transcriptional responses via GC receptors (GCR), there is emerging evidence that GC also mediate transcription-independent signaling events that appear to be very rapid. We previously established that rapid calcium-mediated cell survival signals, triggered by GC stimulation, are involved in the resistance to GC anti-leukemic effects. Furthermore, we demonstrated that chelating intracellular calcium with Bapta-AM or inhibiting downstream ERK1/2 signaling with PD98059 overcame GC resistance in ALL samples.
Here, we investigate the potential role of PLC in GC resistance. We identified that Dexamethasone (Dex) induced cytosolic calcium release via a mechanism involving the cell surface receptor of CXCL12 (CXCR4) internalization and subsequent activation of PLC signalosome. We found that PLC inhibition was able to induce cell death in GC-resistant B-ALL cells by compromising the expression of genes in several metabolic pathways including cell cycle checkpoints, glycolysis and mitochondrial function. These results establish a role for nongenomic signaling in ALL resistance to GC and support identifying novel therapeutics targeting Ca2+ signaling to restore GC sensitivity.

overall_design: We generated GC-resistant cell line by in vitro long-term dexamethasone treatment. GC resistant Nalm-6 cells were treated with or without PLC inhibitor, U73122 (NU), or DMSO (NC) for 16 hours. Samples were then hybrized to RNA sequencing in duplicate.


title: MYCT1 Controls Environmental Sensing in human Hematopoietic Stem Cells [scRNA-seq Dextran]

city: Los Angeles

first: Júlia

last: Aguadé-Gorgorió

address: 615 Charles E Young Drive South

institute: University of California Los Angeles

summary: We identified a novel HSC regulatory gene, MYCT1 (MYC target 1), that is selectively expressed in undifferentiated human HSPC and is critical for human HSC expansion and engraftment. MYCT1 knockdown (KD) in human fetal liver and cord blood (CB) HSPCs impaired expansion ex vivo and engraftment after transplantation, whereas restoring the compromised MYCT1 expression via lentiviral overexpression (OE) in cultured human CB HSPCs improved ex vivo expansion of the most undifferentiated human HSPCs (CD34+CD38-CD90+CD45RA-EPCR+ITGA3+) and enhanced their engraftment ability. We performed single cell RNAseq of uncultured human CB HSPCs, as well as 72 hours after MYCT1 KD and OE to identify MYCT1-regulated programs in HSC and understand the role of MYCT1 in governing human HSPC expansion and engraftment ability. Functional enrichment analysis linked the loss of MYCT1 in HLF+ HSCs to a profound dysregulation of multiple cellular programs essential for HSC stemness, such as oxidative phosphorylation or proteostasis, while MYCT1 OE had the opposite effect and showed closer similarity to uncultured HLF+ HSCs. These data indicate that the loss or gain of MYCT1 expression elicits major effects on programs regulating human HSC functional competence. Mechanistically, we found that MYCT1 governs endocytosis. Therefore, we performed scRNAseq of cultured HSPCs (CD34+CD38-CD90+EPCR+) with low, medium or high levels of endocytosis and found that low endocytosis HSCs had higher MYCT1 expression and favorable transcriptomic profiles of HSC functional competence.

overall_design: Single cell RNAseq of cultured human cord blood HSPCs day 4 (96h) sorted  for HSPCs surface phenotype (CD34+CD38-CD90+EPCR+) and low, medium, or high levels of endocytosis (measured as internalization of fluorescent 10KDa dextran).

first: Feiyang

last: Ma

first: Hanna

last: Mikkola


title: Monoubiquitination of histone H2B is a master regulator of the transcriptome during memory formation

status: Public on Mar 31 2024

city: Blacksburg

first: Timothy

last: Jarome

state: VA

address: 175 W Campus Dr

institute: Virginia Polytechnic Institute and State University

summary: We report a vital role for H2Bubi in active and repressive gene transcription during long-term memory formation. We found that fear conditioning increased or decreased the expression of 86 genes in the hippocampus one hour after training. Remarkably, siRNA knockdown of the H2Bubi ligase, Rnf20, abolished changes in all but one of these genes, Per1, suggesting a critical role for the H2Bubi epigenetic mark in active and repressive gene transcription during memory consolidation.

overall_design: Male Sprague-Dawley rats were injected into the dorsal hippocampus CA1 region with Accell siRNA against Rnf20 or control (Scr-siRNA). Five days later animals were trained to contextual fear conditioning and the CA1 region (dorsal) was dissected, RNA collected via columns and used for RNA-seq analysis

first: Shaghayegh

last: Navabpour

first: Kayla

last: Farrell

first: Nour

last: Omar

first: Madeline

last: Musaus

first: Yu

last: Lin

first: Hehunag

last: Xie


title: TDG orchestrates ATF4-dependent gene transcription during retinoic acid-induced cell lineage committment

city: Nantes

first: Christine

last: Chauveau

address: 30 Boulevard Jean Monnet

institute: CR2TI UMR 1064

summary: Cell differentiation during development is associated to large-scale modifications in the methylome that require the engagement of an active DNA demethylation machinery including Ten-Eleven-Translocation enzymes for oxidation of 5-methylcytosine and the T:G mismatch DNA glycosylase (TDG) for removal of the oxidized bases. Despite this well-defined molecular function, the biological output of TDG activity remains elusive. Here we combined transcriptomic and epigenomic approaches in TDG knock-out embryonal carcinoma cells, an epiblast stem-like cell model, to decipher TDG function in pluripotent cells and their retinoic acid-induced differentiated progeny. We determined that TDG activity is balancing differentiation in favor of a neural fate at the expense of a cardiac mesoderm fate. This process is associated with a sustained activity of a large set of ATF4-dependent genes in relation with TDG engagement at the Atf4 gene promoter and in conjunction with a TDG-dependent regulation of the mammalian target of rapamycin complex 1. These observations highlight the central role of TDG in cell differentiation and support a model linking metabolic reprogramming to cell fate acquisition.

overall_design: To investigate the role of TDG during cell differentiation, we invalidated the Tdg gene by CRISPR-Cas9 in P19 embryonal carcinoma cells. wt and Tdg-null cells were then grown as monolayers or aggregates in the presence or not of retinoic acid. RNA-seq was then applied to the different samples in order to establish the transcriptome associated with these different cell culture conditions.

first: Jeremie

last: Poschmann

first: Gilles

last: Salbert

first: Martin

last: Braud

first: Cynthia

last: Fourgeux

first: Thierry

last: Madigou

first: Marion

last: Turpin


title: In-depth organic mass cytometry reveals differential contents of 3-hydroxybutanoic acid at the single-cell level

city: Beijing

first: Shao Jie

last: Qin

state: Beijing

address: No.5 Summer Palace Road, Yanyuan Street, Haidian District

institute: Peking University

summary:
In-depth organic mass cytometry reveals differential contents of 3-hydroxybutanoic acid on single cell level. Single-cell transcriptome indicates the expression difference of BHB downstream anti-oxidative stress proteins such as MT2A while Fluorescence Activated Cell Sorting (FACS) assay validates positive relationship between BHB and target proteins, which suggests that contents heterogeneity of BHB may endow cancer cells with variant ability to resist surrounding oxidative stress. Our ID-organic cytoMS paves the way for deep single-cell metabolome profiling and the investigation of cancer physiological and pathological processes.
Comprehensive single-cell metabolic profiling is critical for revealing phenotypic heterogeneity and elucidating molecular mechanisms of biological processes. However, single-cell metabolomics remains challenging because of the limited metabolites coverage and disability of isomer discrimination. Herein, we establish a novel single-cell metabolomics platform of in-depth organic mass cytometry (ID-organic cytoMS). Extended single-cell analysis time guarantees sufficient MS/MS acquisition for metabolites identification and the isomers discrimination with online high-throughput analysis, achieving the largest number of about 600 metabolites identified in single cells. Fine sub-typing of MCF-7 cells are first demonstrated by differential contents of 3-hydroxybutanoic acid (BHB) among clusters. Single-cell transcriptome indicates the expression difference of BHB downstream anti-oxidative stress proteins such as MT2A while Fluorescence Activated Cell Sorting (FACS) assay validates positive relationship between BHB and target proteins, which suggests that contents heterogeneity of BHB may endow cancer cells with variant ability to resist surrounding oxidative stress. Our ID-organic cytoMS paves the way for deep single-cell metabolome profiling and the investigation of cancer physiological and pathological processes.

overall_design: 10ⅹGenomics single cell transcriptome sequencing project was undertook by BGI sequencing platform. The distribution of the fragments size was checked by the Agilent 2100 bioanalyzer and the final products were sequenced using the DNBSEQTM platform.

first: Shaojie

first: Jiansen

last: Lu

first: Hongmin

last: Yu


city: Trieste

first: Silvano

last: Piazza

address: localita' Padriciano 99

institute: ICGEB

summary: Selective and pan-histone deacetylase inhibitors (HDACis) emerged at the intersection of these approaches. HDACis suppress EVI1 expression and preferentially impair leukemia proliferation in 3q26 AML compared to other leukemia subtypes in multiple preclinical models. To understand this mechanism of action, we dissected the expression dynamics of the bone marrow leukemia cells of patients treated with hit list compounds, reconstituted with the chromatin-associated co-transcriptional complex of EVI1 by rapid immunoprecipitation mass spectrometry (RIME) in human 3q26 AML models; we focused on the role of the proliferation-associated 2G4 (PA2G4) protein. We demonstrated that genetic and HDACi-mediated suppression of EVI1 modulates MYC and that PA2G4 overexpression rescues AML cells from the inhibitory effects of HDACis. Genetic and small molecule inhibition of PA2G4 abrogated EVI1 and, consequently, MYC in 3q26 AML models, including in patient-derived leukemia xenografts. In conclusion, our work positions PA2G4 at the crosstalk of the EVI1 leukemogenic signal for developing new therapeutics and, at this time, urges the use of upfront HDACi-based combination therapies in patients with 3q26 AML.

overall_design: In this work, we challenged the ecotropic viral integration site-1 gene (EVI1/MECOM), the most lethal oncogenic TF in AML carrying chromosome 3q26 abnormalities, by crossing cell-based screens, gene expression tools, and mass spectrometry–based approaches to identify modulators of EVI1.

first: Matteo

last: Marchesini

first: Andrea

last: Gherli

first: Lucas

last: Moron Dalla Tor

first: Elisa

last: Simoncini

first: Anna

last: Montanaro

last: D’Antuono

first: Luca

last: Pagliaro

first: Raffaella

last: Zamponi

first: Angela

last: Falco

first: Bruno

last: Lorusso

first: Federica

last: Mezzasoma

first: Costanza

last: Anna Maria Lagrasta

first: Federico

last: Quaini

first: Giovanni

last: Roti


title: Minor splicing factor 65K/RNPC3 interacts with ANKRD11 and mediates HDAC3-regulated histone deacetylation and transcription

city: Wuhan

first: Shaobo

last: Liang

state: Hubei Sheng

address: BaYi Road, LUOJIA HILL

institute: Wuhan University

summary: Pre-mRNA splicing is coupled with DNA- and other RNA-processing machineries in the nucleus to form multilayer networks for accurate regulation of gene expression. However, those established couplings are major spliceosome-related, whether the minor spliceosome is involved remains unclear. Here, we first identified a direct interaction between a minor-specific protein 65K/RNPC3 and the histone deacetylase HDAC3 cofactor ANKRD11 through affinity co-purification using Drosophila lysate. In vivo assays revealed that the head of both DmAnkrd11Δ/Δ and Dm65KΔ/Δ mutants exhibited increased histone acetylation at H3K9 and H4K5, consistent with the high level of ANKRD11 in the central nervous systems. We then found that the 65K-ANKRD11 interaction is conserved in human, in which the HsANKRD11 middle-stretched domain mediates Hs65K association with HDAC3. CUT&Tag assays demonstrated that HDAC3 and Hs65K synergistically bind on chromatin, and the knockdown of ANKRD11 simultaneously decreased their bindings at the same chromatin locations and also increased nearby acetylation of H3K9. Sequencing of mRNAs revealed that the deficiency of HsANKRD11 or Hs65K caused changes in gene expression correlated with their chromatin-binding changes of HDAC3 and Hs65K and the levels of H3K9ac. This study provides a novel strategy for the regulation of histone deacetylation and gene expression through a minor spliceosome-specific component.

sample_organism:
Drosophila melanogaster
Homo sapiens

platform_organism:
Drosophila melanogaster
Homo sapiens

overall_design: To investigate the cooperative function 65K/ANKRD11/HDAC3 complex in the regulation of histone acetylation and gene transcription, we established the Dm65K-deletion and DmAnkrd11-deletion Drosophila strain, RNA was from Proximal heads or adults from 100 flies (1 days old), then libraries was prepared for transcriptome sequencing. Comparative gene expression profiling analysis of RNA-seq data for WT, Dm65K-deletion and DmAnkrd11-deletion Drosophila strain. 293T cell lines in which each target gene has been knocked down by shRNA. Comparative gene expression profiling analysis of RNA-seq data for 293T cells and its KD derivatives (sh65K and shANKRD11). The CUT&Tag assay was performed as described (Kaya-Okur et al., 2019). CUT&Tag assay using 65K or HDAC3 antibody to find out binding sites in 293T cells or its KD derivatives (ctrl and shANKRD11). The ChIP assay was performed as described (Gorkin et al., 2020). Briefly, the HEK293T KD derivatives (ctrl, sh65K and shANKRD11) cells were cross-linked by 1% formaldehyde for 10 min and then the cell lysate was prepared by removing cytoplasm followed by sonication-based chromatin fragmentation. Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for Histone 3 lysine 9 acetylation DNAs were subjected to library preparation with the Scale ssDNA-seq Lib Prep Kit for Illumina V2 (#RK20228, Abclonal).

first: Chenhui

last: Li

first: Qiwei

last: Huang

first: Yongzhen

last: Xu


title: Profiling of transcriptional targets of Jak1 and Stat1 in proliferating and differentiating myoblasts

city: Hong Kong

first: Xianwei

last: Chen

address: Clearwater bay, Kowloon

institute: Hong Kong University of Science and Techanology

summary:
Styxl2, a poorly characterized pseudophosphatase, was identified as a transcriptional target of the Jak1-Stat1 pathway during myoblast differentiation in culture. Styxl2 is specifically expressed in vertebrate striated muscles. By gene knockdown in zebrafish or genetic knockout in mice, we found that Styxl2 plays an essential role in maintaining sarcomere integrity in developing muscles. To further reveal the functions of Styxl2 in adult muscles, we generated two inducible knockout mouse models: one with Styxl2 being deleted in mature myofibers to assess its role in sarcomere maintenance, and the other in adult muscle satellite cells (MuSCs) to assess its role in de novo sarcomere assembly.
We find that Styxl2 is not required for sarcomere maintenance but functions in de novo sarcomere assembly during injury-induced muscle regeneration. Mechanistically, Styxl2 interacts with non-muscle myosin IIs, enhances their ubiquitination, and targets them for autophagy-dependent degradation. Without Styxl2, the degradation of non-muscle myosin IIs is delayed, which leads to defective sarcomere assembly and force generation. Thus, Styxl2 promotes de novo sarcomere assembly by interacting with non-muscle myosin IIs and facilitating their autophagic degradation.

overall_design: We previously reported that the Jak1-Stat1 pathway plays an important role in myogenic differentiation in cultured myoblasts. To examine the transcriptomes regulated by Jak1 and Stat1, we conducted a microarray analysis in C2C12 myoblasts: cells were first transfected with either a control (GFP) siRNA or specific siRNAs targeting Jak1 or Stat1 and the cells were harvested at two different time points: (1) after 24 hours of growth in the growth medium (GM 24h, the proliferation stage); (2) after 12 hours in the differentiation medium following 24 hours of growth in the GM (DM 12h, the early differentiation stage).

first: Yanfeng


title: Transcriptional profiles in lesional skin of Postn deficient FADS mice

city: Saga

first: Satoshi

last: Nunomura

address: 5-1-1 Nabeshima

department: Department of Biomolecular Sciences

institute: Saga Medical School

summary: We recently generated FADS mouse (Nestincre-mediated conditional knockout mouse of Ikk2 gene), a mouse model of atopic dermatitis.  To examine the functions of periostin in dermatitis, periostin KO FADS mice were prepared. We then performed Cap analysis of gene expression (CAGE) to elucidate transcriptional profiles in lesional skin of periostin KO FADS mice.

overall_design: Four lesional facial skins were prepared from 4 week-old periostin KO FADS mice.

first: Kenji

last: Izuhara

first: Isao

last: Kitajima


title: Growth retardation in a mouse model of Kabuki syndrome 2 bears mechanistic similarities to Kabuki syndrome 1.

status: Public on Mar 30 2024

city: Baltimore

first: Christine

last: Gao

state: MD

address: 733 N. Broadway, Rm 406

institute: Johns Hopkins University School of Medicine

summary: Growth deficiency is a characteristic feature of both Kabuki syndrome 1 (KS1) and Kabuki syndrome 2 (KS2), Mendelian disorders of the epigenetic machinery with similar phenotypes but distinct genetic etiologies. We previously described skeletal growth deficiency in a mouse model of KS1 and further established that a Kmt2d -/- chondrocyte model of KS1 exhibits precocious differentiation. Here we characterized growth deficiency in a mouse model of KS2, Kdm6a tm1d/+. We show that Kdm6a tm1d/+ mice have decreased femur and tibia length compared to controls and exhibit abnormalities in cortical and trabecular bone structure. Kdm6a tm1d/+ growth plates are also shorter, due to decreases in hypertrophic chondrocyte size and hypertrophic zone height. Given these disturbances in the growth plate, we generated Kdm6a -/- chondrogenic cell lines. Similar to our prior in vitro model of KS1, we found that Kdm6a -/- cells undergo premature, enhanced differentiation towards chondrocytes compared to Kdm6a +/+ controls. RNA-seq showed that Kdm6a -/- cells have a distinct transcriptomic profile that indicates dysregulation of cartilage development. Finally, we performed RNA-seq simultaneously on Kmt2d -/-, Kdm6a -/-, and control lines at Days 7 and 14 of differentiation. This revealed surprising resemblance in gene expression between Kmt2d -/- and Kdm6a -/- at both time points and indicates that the similarity in phenotype between KS1 and KS2 also exists at the transcriptional level.

overall_design: We generated Kdm6a -/- and Kdm6a +/+ cell lines from the murine teratocarcinoma parental cell line ATDC5 using CRISPR-Cas9. For the first experiment, we differentiated Kdm6a -/- and Kdm6a +/+ cells towards chondrocytes for 14 days. We then harvested RNA and performed library prep for RNA-seq. For the second experiment, we performed chondrogenic differentiation on Kdm6a -/- and Kdm6a +/+ cells as well as Kmt2d -/- and Kmt2d +/+ cell lines, previously generated from the same parental ATDC5 cell line. We collected samples for RNA-seq at both Day 7 and Day 14 of differentiation for all cell lines.

first: Kasper

middle: D

last: Hansen

first: Jill

last: Fahrner


title: Single-cell RNA-sequencing of  E2f7 knockout and wild-type littermate control murine tracheal epithelial cells cultured at air-liquid interace for seven days.

city: San Francisco

first: Semil

last: Choksi

email: semilc@gmail.com, semil.choksi@ucsf.edu

state: CA

address: 555 Mission Bay Blvd. South

institute: UCSF

summary: The canonical mitotic cell cycle coordinates cell growth, DNA replication, centriole duplication and cytokinesis to generate two cells from one. In certain contexts, such as in mammalian trophoblast giant cells or hepatocytes, cells employ cell cycle variants like the endocycle or endomitosis to increase DNA content without undergoing cytokinesis. Multiciliated cells, found in the mammalian airway, brain ventricles and reproductive tracts, generate hundreds of centrioles during differentiation, each of which extend a motile cilium. We found that multiciliated cells utilize a variant of the cell cycle, which we refer to as the multiciliation cycle. The multiciliation cycle redeploys much of the mitotic cell cycle regulatory framework, including cell division kinases (CDKs) and cyclins, to generate hundreds of centrioles without undergoing DNA synthesis or cytokinesis. Unlike the mitotic cycle, a transcriptional repressor, E2F7, is upregulated during the multiciliation cycle. E2F7 directly represses genes encoding DNA synthesis machinery during the multiciliation cycle. Loss of E2F7 results in aberrant DNA synthesis and disruption of centriole biogenesis in differentiating multiciliated cells. Thus, multiciliation is coordinated by a variant cell cycle that uses E2F7 to uncouple centriole synthesis from DNA replication.

overall_design: Mouse tracheal epithelial cells grown at air-liquid interface for seven days from either wildtype or E2f7 knockout mice were analyzed using scRNAseq.

first: Lauren

last: Byrnes

first: Mia

last: Konjikusic

first: Rachel

last: Deleon

first: Quanlong

last: Lu

first: Christopher

last: Westlake

first: Jeremy

middle: F

last: Reiter


title: Single-cell RNA-sequencing of Ribociclib treated mTECs

overall_design: Mouse tracheal epithelial cells grown at air-liquid interface for three days after treatment with DMSO or Ribociclib were analyzed using scRNAseq.


title: Expression data from Arabidopsis suspension cells overexpressing VND1-GFP

city: tokyo

first: Kyoko

last: Ohashi-Ito

address: 7-3-1 hongo bunkyo-ku

department: biological sciences

institute: the university of tokyo

summary: VND1 is a member of NAC-type family transcription factors and has a important role in xylem differentiation. However, how VND1 regulates vascular differentiation is not well known.  Here, we identified specific genes downstream of  VND1 with transformed suspension culture cells in microarray experiments.

overall_design: To identify genes downstream of VND1, we collected three samples at 24h with or without estrogen with triplicates.

first: Kuninori

last: Iwamoto

first: Hiroo

last: Fukuda


title: Gene expression profile at single cell level of ovarian clear cell carcinoma (OCCC) [snRNA-seq OCCC]

city: Tokyo

first: Yutaro

last: Mori

address: 2-21-1 Kaga, Itabashi-ku

department: Advanced Comprehensive Research Organization

institute: Teikyo University

summary: The diversity of chemoresistant OCCC is unknown. We performed single nucleus RNA sequencing (snRNA-seq) to analyze the diversity of OCCC.

overall_design: Nuclei were isolated from frozen OCCC tumor samples and analyzed using snRNAseq.

first: Koji

last: Okamoto


title: Biochemical-free enrichment or depletion of RNA classes in real-time during direct RNA sequencing with RISER (whole blood)

status: Public on Mar 29 2024

city: Canberra

first: Alexandra

last: Sneddon

state: ACT

address: 131 Garran Rd Acton

country: Australia

institute: The Australian National University

summary: The heterogeneous composition of cellular transcriptomes poses a major challenge for detecting weakly expressed RNA classes, as they can be obscured by abundant RNAs. Although biochemical protocols can enrich or deplete specified RNAs, they are time-consuming, expensive and can compromise RNA integrity. Here we introduce RISER, a biochemical-free technology for the real-time enrichment or depletion of RNA classes. RISER performs selective rejection of molecules during direct RNA sequencing by identifying RNA classes directly from nanopore signals with deep learning and communicating with the sequencing hardware in real time. By targeting the dominant messenger and mitochondrial RNA classes for depletion, RISER reduced their respective read counts by more than 85%, resulting in an increase in sequencing depth of up to 93% for long non-coding RNAs. We also applied RISER for the depletion of globin mRNA in whole blood, achieving a decrease in globin reads by more than 90% as well as a significant increase in non-globin reads. Furthermore, using a GPU or a CPU, RISER is faster than GPU-accelerated basecalling and mapping. RISER’s modular and retrainable software and intuitive command-line interface allow easy adaptation to other RNA classes. RISER is available at https://github.com/comprna/riser.

overall_design: Nanopore direct RNA sequencing was performed using 2 blood samples from human donors for training RISER, and 1 blood sample from another human donor for testing RISER during live MinION sequencing.

first: Agin

last: Ravindran

first: Somasundhari

last: Shanmuganandam

first: Madhu

last: Kanchi

first: Nadine

last: Hein

first: Simon

last: Jiang

first: Nikolay

last: Shirokikh

first: Eduardo

last: Eyras


title: Visium spatial gene expression analyses of ovarian clear cell carcinoma (OCCC) [visium OCCC]

summary: We attempted to determine histological localization of chemoresistant cancer subpopulation by performing spatial transcriptomics.

overall_design: OCCC tumor samples embedded in OCT compound were frozen and analyzed using visium spatial transcriptomics.


title: Targeting PDGF signaling of cancer-associated fibroblasts blocks feedback activation of HIF-1α and overcomes chemoresistance in ovarian clear cell carcinoma

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

overall_design: Refer to individual Series


title: Mitotic gene regulation by the N-MYC-WDR5-PDPK1 nexus

city: Nashville

first: Jing

last: Wang

state: TN

address: 2220 Pierce Avenue

institute: Vanderbilt University Medical Center

summary:
Background: During mitosis the cell depends on proper attachment and segregation of replicated chromosomes to generate two identical progeny. In cancers defined by overexpression or dysregulation of the MYC oncogene this process becomes impaired, leading to genomic instability and tumor evolution. Recently it was discovered that the chromatin regulator WDR5—a critical MYC cofactor—regulates expression of genes needed in mitosis through a direct interaction with the master kinase PDPK1. However, whether PDPK1 and WDR5 contribute to similar mitotic gene regulation in MYC-overexpressing cancers remains unclear. Therefore, to characterize the influence of WDR5 and PDPK1 on mitotic gene expression in cells with high MYC levels, we performed a comparative transcriptomic analysis in neuroblastoma cell lines defined by MYCN-amplification, which results in high cellular levels of the N-MYC protein.
Results: Using RNA-seq analysis, we identify the genes regulated by N-MYC and PDPK1 in multiple engineered CHP-134 neuroblastoma cell lines and compare them to previously published gene expression data collected in CHP-134 cells following inhibition of WDR5. We find that as expected N-MYC regulates a multitude of genes, including those related to mitosis, but that PDPK1 regulates specific sets of genes involved in development, signaling, and mitosis. Analysis of N-MYC- and PDPK1-regulated genes reveals a small group of commonly controlled genes associated with spindle pole formation and chromosome segregation, which overlap with genes that are also regulated by WDR5. We also find that N-MYC physically interacts with PDPK1 through the WDR5-PDPK1 interaction suggesting regulation of mitotic gene expression may be achieved through a N-MYC-WDR5-PDPK1 nexus.
Conclusions: Overall, we identify a small group of genes highly enriched within functional gene categories related to mitotic processes that are commonly regulated by N-MYC, WDR5, and PDPK1 and suggest that a tripartite interaction between the three regulators may be responsible for setting the level of mitotic gene regulation in N-MYC amplified cell lines. This study provides a foundation for future studies to determine the exact mechanism by which N-MYC, WDR5, and PDPK1 converge on cell cycle related processes.

overall_design: For one experiment, we induced the genetic inhibitor of MYC, called OmoMYC, for three days in engineered MYCN-amplified CHP-134 cells and compared gene expression changes to enhanced green fluorescent protein induction. We also treated parental CHP-134 cells with 500 nM dTAG47 for 18 hr as a control experiment in this study.

first: April

last: Weissmiller

first: William

middle: P

last: Tansey

first: Qi

last: Liu


title: Acute adenoviral cardiac infection elicits an arrhythmogenic substrate prior to inflammatory myocardial remodeling and myocarditis

city: Roanoke

first: James

last: Smyth

address: 2 Riverside Circle

institute: Fralin Biomedical Research Institute

summary: Viral cardiac infection represents a significant clinical challenge encompassing several etiological agents, disease stages, complex  presentation, and a resulting lack of mechanistic understanding. Myocarditis is a leading cause of sudden cardiac death in young adults, where current knowledge in the field is dominated by later disease phases, and pathological host immune responses. However, little is known regarding how viral infection can acutely induce an arrhythmogenic substrate prior to significant immune responses. Adenovirus is a leading cause of myocarditis, but due to species-specificity, models of infection are lacking and it is not understood how adenoviral infection may underlie sudden cardiac arrest. Mouse Adenovirus Type-3 (MAdV-3) was previously reported as cardiotropic, yet has not been utilized to understand mechanisms of cardiac infection and pathology. We have developed MAdV-3 infection as a model to investigate acute cardiac infection and molecular alterations to the infected heart prior to an appreciable immune response or gross cardiomyopathy. By optical mapping of infected hearts we find decreases in conduction velocity concomitant with increased Cx43Ser368 phosphorylation, a residue known to regulate gap junction function. Hearts from animals harboring a phospho-null mutation at Cx43Ser368 are protected against MAdV-3 induced conduction velocity slowing. Additional to gap junction alterations, patch clamping of MAdV-3-infected adult mouse ventricular cardiomyocytes reveals prolonged action potential duration as a result of decreased IK1 and IKs current density. Turning to human systems, we find human adenovirus type-5 (HAdV-5) increases phosphorylation of Cx43Ser368 and disrupts synchrony in human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs), indicating common mechanisms with our mouse whole heart and adult cardiomyocyte data. Together, these findings demonstrate that adenoviral infection creates an arrhythmogenic substrate through direct targeting of gap junction and ion channel function in the heart. Such alterations are known to precipitate arrhythmias and likely contribute to sudden cardiac death in acutely infected patients.

overall_design: To determine transcriptomic changes in the cardiac ventricles following mouse adenovirus type-3 (MAdV-3) infection, cardiac ventricles were collected from 3 mock-infected mice and 3 MAdV-3-infected mice at 7 days post infection. Snap-frozen ventricles were sent to GeneWiz for RNA-seq processing. Gene expression in MAdV-3-infected ventricles were compared to that of mock-infected ventricles.

first: Rachel

last: Padget

first: Michael

last: Zeitz

first: Grace

middle: A

last: Blair

first: Xiaobo

last: Wu

last: North

first: Mira

middle: T

last: Tanenbaum

first: Kari

middle: E

last: Stanley

first: Chelsea

middle: M

last: Phillips

first: David

last: King

first: Samy

last: Lamouille

first: Robert

middle: G

last: Gourdie

first: Gregory

middle: S

last: Hoeker

first: Sharon

last: Swanger

first: Steven

last: Poelzing


title: Multiome analysis (ATAC + Gene Expression (GEX)) of SUN1 knockout and control CT26 tumors grown in syngeneic mouse model

city: Calgary

first: Edwin

last: Wang

state: Alberta

address: 2500 University Drive N.W.

institute: University of Calgary

summary: T cell infiltration is essential for immune checkpoint inhibitors to be effective in treating solid cancers. Through a bioinformatic pipeline, we identified a target gene SUN1 that might relate to modulating immune cell infiltration and immune response. Thus, we generated one Sun1_knockout CT26 cell line (Sun1_KO) using CRISPR-Cas9. By performing multiome single nuclei sequencing using tumors grown in syngeneic model, we set out to understand how mouse Sun1 can affect the regulatory network in tumor cells in vivo.

overall_design: Sun1_KO and Sun1_WT cells were subcutaneously injected in the right flank. Tumors were collected at day 14 post-inoculation, and digested into single cells, which were applied to the preparation of 10x multiome libraries according to the instruction. The resulted GEX library and ATAC library were sequenced separately.

first: Sahar

last: Nasr

first: Lin

last: Li