title: A consensus molecular subtypes classification strategy for clinical colorectal cancer tissues

city: Amsterdam

first: Louis

last: Vermeulen

address: Meibergdreef 9

department: LEXOR Department

country: Netherlands

institute: Amsterdam University Medical Center

summary: This work presents a novel Consensus Molecular Subtypes (CMS) classifier for colorectal cancer (CRC), optimized for degraded RNA stemming from clinical formalin-fixed paraffin-embedded tissue samples.

overall_design: The CMSFFPE classifier was developed using genes with a high transcript integrity in FFPE-derived RNA. We evaluated the classification accuracy in two FFPE-RNA datasets with matched fresh-frozen (FF) RNA data, and an FF-derived RNA set. An FFPE-RNA application cohort of metastatic CRC patients was established, partly treated with anti-EGFR therapy. Key characteristics per CMS were assessed.

first: Tim

last: de Back

first: Tan

last: Wu

first: Pascale

last: Schafrat

first: Sanne

last: ten Hoorn

first: Miaomiao

last: Tan

first: Lingli

last: He

first: Sander

last: van Hooff

first: Jan

last: Koster

first: Lisanne

last: Nijman

first: Geraldine

last: Vink

first: Inès

last: Beumer

first: Clara

last: Elbers

first: Kristiaan

last: Lenos

first: Dirkje

last: Sommeijer

first: Xin

last: Wang


title: Transcriptional profiling of CD44hi AT2 cells under homeostasis and after lung injury [I]

status: Public on May 23 2024

city: Chicago

first: Qian

last: Chen

state: IL

address: 909 S Wolcott avenue, COMRB 4160

department: Department of Pharmacology and Regenerative Medicine

country: USA

institute: University of Illinois at Chicago

type: Expression profiling by high throughput sequencing

summary: The epithelium of alveoli is composed of alveoli type I cells (AT1) that cover ~95% of the surface area and alveoli type II cells (AT2) that secret surfactant as well as function as adult stem cells.  To characterize a subgroup of AT2 that express higher levels of hyaluronan receptor CD44 (the CD44high, or CD44hi AT2) and behave as alveoli epithelial stem cells during steady-state homeostasis, we used RNA-sequencing to compare the expression profiles of these cells with the bulk CD44low (or CD44lo ) AT2.  CD44hi AT2 showed gene signatures that indicate partial de-differentiation and a higher potential to proliferate compared with CD44lo AT2.  At steady-state conditions, CD44hi AT2 also showed higher expression of genes activated by NFκB mediated inflammatory responses and transcriptome signatures that resemble early-stage Kras-induced adenocarcinoma.  Furthermore, these cells were more responsive to KrasG12D induction both in 3D tumor organoid culture and in mouse lung cancer cell transplantation models. In summary, CD44hi AT2 plays an essential role in the homeostatic maintenance of the alveolar epithelium and tumor initiation.

overall_design: The goal of this study was to compare the gene expression profiles of CD44hi alveolar type 2 (AT2) cells versus CD44lo AT2 cells. CD44hi and CD44lo AT2 cells were sorted using fluorescence activated cell sorter (FACS) from 8 female SpC-creER/Rosa-mTmG mice at 6-8 weeks of age. Total RNA profiles were generated by deep sequencing using Illumina HiSeq4000. Three biological replicates were included for each cell type.

first: Alessandro

middle: M

last: Chan

first: Yuru

last: Liu


title: Transcriptional profiling of CD44hi AT2 cells under homeostasis and after lung injury [II]

summary: The epithelium of alveoli is composed of alveoli type I cells (AT1) that cover ~95% of the surface area and alveoli type II cells (AT2) that secret surfactant as well as function as adult stem cells.  CD44hi subpopulation of AT2 cells exhibits higher potential to proliferate and differentiate under steady state conditions. To understand the CD44hi subpopulation of AT2 cells after lung injury, we assessed those cells in a Pseudomonas aeruginosa (PA) induced mouse model of lung injury. CD44hi and CD44low AT2 cells were FACS sorted from untreated and 3d PA treated mice. In addition,  since Sca1+ AT2 cells were shown to emerge during the repair phase of the PA induced injury in our earlier studies, we also isolated Sca1+ AT2 cells at 3d post PA injury to compare with the CD44hi subpopulation. RNA-seq was used to compare the expression profile of the different subgroups of AT2 cells.

overall_design: The goal of this study was to (1) compare the gene expression profiles of CD44hi AT2 cells under steady-state conditions and after lung injury and (2) compare the gene expression profile of CD44hi, CD44low, and Sca1+ AT2 cells after lung injury. Total RNA profiles were generated by deep sequencing using Illumina HiSeq4000.

first: Alesandro


title: Gene-expression data for SKH1 mice receiving hindlimb radiation therapy [Glial_Panel]

status: Public on May 22 2024

city: St Paul

first: Davis

last: Seelig

state: MN

address: 1352 Boyd Ave.

department: Department of Veterinary Clinical Sciences

institute: University of Minnesota

summary: NanoString nCounter Glial Profiling Panel

overall_design: In this study, there is gene data from three SKH1 mice brains per timepoint treatment groups

first: Kimberly

last: Demos-Davies

first: Jessica

last: Lawrence

last: Coffey

first: Amy

last: Morgan

last: Ferreira

first: Luke

last: Hoeppner


title: Gene-expression data for SKH1 mice receiving hindlimb radiation therapy [Neuroinflammation Panel]

summary: NanoString nCounter Neuroinflammation Panel


title: CRL4-DCAF15 control of cohesin dynamics sustains cell proliferation [RNA-seq_dTAG]

city: Philadelphia

first: Luca

last: Busino

state: Pennsylvania

address: 421 Curie Boulevard

department: Cancer Biology

institute: University of Pennsylvania

summary: The CRL4-DCAF15 E3 ubiquitin ligase complex is targeted by the aryl-sulfonamide molecular glues, leading to neo-substrate recruitment, ubiquitination, and proteasomal degradation. However, the physiological function of DCAF15 remains unknown. Using a domain-focused genetic screening approach, we reveal DCAF15 as an acute myeloid leukemia (AML)-biased dependency. Loss of DCAF15 results in suppression of AML through compromised replication fork integrity and consequent accumulation of DNA damage. Accordingly, DCAF15 loss sensitizes AML to replication stress-inducing therapeutics. Mechanistically, we discover that DCAF15 directly interacts with the SMC1A protein of the cohesin complex and destabilizes the cohesin regulatory factors PDS5A and CDCA5. Loss of PDS5A and CDCA5 removal precludes cohesin acetylation on chromatin, resulting in uncontrolled chromatin loop extrusion, defective DNA replication, and apoptosis. Collectively, our findings uncover an endogenous, cell autonomous function of DCAF15 in sustaining AML proliferation through post-translational control of cohesin dynamics.

overall_design: RNA-sequencing analysis of OCI-AML3 human AML cell lines after 0h, 24h, or 72h dTAGv1 drug-induced degradation of endogenous DCAF15 protein with N-terminal FLAG-HA-FKBP12(F36V) tag (dTAG).

first: Grant

middle: P

last: Grothusen

first: Renxu

last: Chang


title: Hypoxia promotes histone H3K9 lactylation to enhance LAMC2 transcription in esophageal squamous cell carcinoma

city: tianjin

first: yong

last: zang

address: Heping District qixiangtailu22hao

country: China

institute: Tianjin Medical University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Hypoxia promotes tumorigenesis and lactate accumulation in esophageal squamous cell carcinoma (ESCC). Lactate can induce histone lysine lactylation (Kla, a recently-identified histone marks) to regulate transcription. However, the functional consequence of histone Kla under hypoxia in ESCC remains to be explored. Here, we reveal that hypoxia facilitates histone H3K9la to enhance LAMC2 transcription for proliferation of ESCC. We found that global level of Kla was elevated under hypoxia, and thus identified the landscape of histone Kla in ESCC by quantitative proteomics. Furthermore, we show a significant increase of H3K9la level induced by hypoxia. Next, MNseq ChIP-seq and RNA-seq analysis suggest that H3K9la is enriched at the promoter of cell junction genes. Finally, we demonstrate that the histone H3K9la facilitates the expression of LAMC2 for ESCC invasion by in vivo and in vitro experiments. Briefly, our study reveal a vital role of histone Kla triggered by hypoxia in cancer.

overall_design: To investigate the function of hypoxia  in KYSE30 cells, we cultured KYSE30 cells in hypoxia or normoxia environment.To investigate the function of LAMC2  in KYSE30 cells,we established LAMC2 overexpressed KYSE30 cell lines. MNseq Chromatin immunoprecipitation (MNseq ChIP-seq) for histone 3 lysine 9 lactylation in KYSE30cells cultured in hypoxia or normoxia environment

first: Yong

last: Zang

first: Aiyuan

first: Jianji

last: Zhang

first: Kai


title: CRL4-DCAF15 control of cohesin dynamics sustains cell proliferation [HiC]

overall_design: HiC DNA-sequencing (HiC-seq) analysis in HEL human AML cell line without or with genetic knockout of DCAF15.

first: Phillip

last: Wulfridge

first: Kavitha

last: Sarma


title: Treatment with cultured epidermal autograft from revertant skin for epidermolytic ichthyosis

city: Nagoya

first: Takuya

last: Takeichi

address: 65 Tsurumai-cho, Showa-ku

country: Japan

institute: Nagoya University Graduate of School

summary: We performed clinical trial of treatment with cultured epidermal autograft (CEA) produced from patients’ own revertant epidermal keratinocytes as a proof-of-concept study. We successfully produced CEAs from genetically confirmed revertant skin of the three EI patients and genetically confirmed that CEAs mainly consist of revertant wild-type cells by amplicon sequencing and droplet digital PCR analysis. Single-cell RNA sequencing analysis confirmed normal proliferation and safety profiling of CEAs.

overall_design: To assess the proliferation/differentiation abilities and safety of cultured epidermal autograft generated from the patients’ revertant skin, we performed single-cell RNA sequencing analysis using cells from cultured epidermal autograft and control cultured normal human keratinocytes to see the gene expression profiles.

first: Kana

last: Tanahashi

first: Takenori

last: Yoshikawa

first: Masashi

last: Akiyama


title: CRL4-DCAF15 control of cohesin dynamics sustains cell proliferation [ChIP-seq]

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) analysis for total-SMC3 and acetyl-SMC3 in HEL human AML cell line without or with genetic knockout of DCAF15.

last: Zhou


title: CRL4-DCAF15 control of cohesin dynamics sustains cell proliferation [RNA-seq_CRISPR]

overall_design: RNA-sequencing analysis of MV4-11 and HEL human AML cell lines 4 days after genetic knockout of DCAF15, TP53, both, or neither.


title: Engineering an inducible leukemia-associated transcription factor enables large-scale ex vivo production of functional human phagocytes

city: Planegg

first: Anna

last: Danese

address: Grosshaderner strasse 9

country: Germany

institute: Ludwig Maximilians University

summary: Ex vivo expansion of human CD34+ hematopoietic stem and progenitor cells remains a challenge due to rapid differentiation after detachment from the bone marrow niche. In this study, we assessed the capacity of an inducible transcription factor to enable sustained ex vivo proliferation of hematopoietic precursors and their capacity to differentiate into functional phagocytes. We fused the coding sequences of an FK506-Binding Protein 12 (FKBP12)-derived destabilization domain (DD) to the myeloid/lymphoid lineage leukemia/eleven nineteen leukemia (MLL-ENL) transcription factor to generate the fusion protein DD-MLL-ENL and retrovirally expressed the protein switch in human CD34+ progenitors. Using Shield1, a chemical inhibitor of DD fusion protein degradation, we established large-scale and long-term expansion of late monocytic precursors. Upon Shield1 removal, the cells lost self-renewal capacity and spontaneously differentiated, even after 2.5 years of continuous ex vivo expansion. In the absence of Shield1, stimulation with IFN-γ, LPS, and GM-CSF triggered terminal differentiation. Gene expression analysis of the obtained phagocytes revealed marked similarity with naïve monocytes. In functional assays, the novel phagocytes migrated towards CCL2, attached to VCAM-1 under shear stress, produced reactive oxygen species, and engulfed bacterial particles, cellular particles and apoptotic cells. Finally, we demonstrated Fcγ receptor recognition and phagocytosis of opsonized lymphoma cells in an antibody-dependent manner. Overall, we have established a novel transcription factor that, as a single factor, is useful for large-scale ex vivo production of human phagocytes. Such adjustable transcription factors have the potential to be applied as molecular tools to produce functional immune cells for cell therapy.

overall_design:
RNA expression analysis via NanoString nCounter Analysis System with four different cell types (three biological replicates each) using the myeloid innate immunity panel. Total RNA was isolated with the RNeasy Plus Mini Kit (Qiagen). The four different cell types include:
- Untransduced hematopoietic stem and progenitor cells (HSPCs) from healthy donors (cultivated for 3 days in media with IL-3, IL-6, SCF, TPO, GM-CSF, FLT3-L)
- HSPCs transduced with a retroviral construct DD-MLL-ENL-ires-GFP. DD is a destabilization domain leading to rapid degradation of MLL-ENL. The small molecule Shield-1 prevents degradation. The used cells were cultivated in the presence of Shield-1 for over 50 days to obtain an MLL-ENL expressing, GFP+ progenitor cell population. Cultivation in media with IL-3, IL-6, SCF, TPO, GM-CSF, FLT3-L.
- Macrophages differentiated from the MLL-ENL expressing progenitor cells after withdrawal of Shield-1 and cultivation in media with GM-CSF, LPS and IFN? for 14 days.
- CD14+ monocytes derived from peripheral blood from healthy donors. PBMCs were isolated via density gradient centrifugation. CD14+ monocytes were isolated using the classical monocyte isolation kit from Miltenyi.

****Submitter states "the raw fastq files are not uploaded because they are patient derived cells and under German law we cannot publicly share the raw sequencing file on GEO"***

first: Roland

last: Windisch

first: Alexander

last: Fischer

first: Christian

last: Wichmann


title: Renal papillary cell carcinoma

city: Guangzhou

first: jinhuan

last: wei

address: No.58 zhongshan 2rd Road

institute: SYSU

summary: We used RNA-SEQ technique to detect lncRNAs expression differences in 53 pairs of tumor and adjacent normal tissues of renal papillary cell carcinoma

first: Jin-Huan

last: Wei

first: Kang-Bo

last: Huang

first: Jun-Hang

last: Luo


title: Novel epigenetic based differentiation therapy for Acute Myeloid Leukemia

status: Public on May 21 2024

city: Pamplona

first: Nahia

last: Gomez-Echarte

address: Avenida Pio XII, 55

country: Spain

institute: CIMA

summary: Despite the development of novel therapies for acute myeloid leukemia (AML), outcomes remain poor for most patients, and therapeutic improvements are an urgent unmet need. Although treatment regimens promoting differentiation have succeeded in the treatment of acute promyelocytic leukemia (APL), their role in other AML subtypes needs to be explored. Here we identified and characterized two lysine (K) deacetylase inhibitors (DACi), CM-444 and CM-1758, exhibiting capacity to promote myeloid differentiation in all AML subtypes at low non-cytotoxic doses unlike other commercial HDACi. Analyzing the acetylome after CM-444 and CM-1758 treatment revealed modulation of non-histone proteins involved in the enhancer–promoter chromatin regulatory complex, including bromodomain proteins (BRDs). This acetylation was essential for enhancing the expression of key transcription factors directly involved in the differentiation therapy induced by CM-444/CM-1758 in AML. In summary, these compounds may represent effective differentiation-based therapeutic agents across AML subtypes with a novel mechanism for treatment of AML.

overall_design: Epigenetic small molecules like CM-444, compared to commercial differentiation agents like panobinostat, used for AML differentiation.

first: E

last: San José-Enériz

first: N

last: Gimenez-Camino

first: O

last: Rabal

first: L

last: Garate

last: Miranda

last: Gómez-Echarte

first: F

last: García

first: S

last: Charalampopoulou

last: Sáez

first: A

last: Vilas-Zornoza

first: P

last: San Martín-Uriz

middle: V

last: Valcárcel

last: Barrena

first: D

last: Alignani

last: Tamariz-Amador

last: Pérez-Ruiz

last: Hilscher

first: M

last: Schutkowski

last: Alfonso-Pierola

last: Martinez-Calle

middle: J

last: Larrayoz

first: B

last: Paiva

last: Calasanz

first: J

last: Muñoz

last: Isasa

middle: I

last: Martín-Subero

last: Pineda-Lucena

last: Oyarzabal

first: X

last: Agirre

last: Prosper


title: In vivo interaction screening reveals liver-derived constraints to metastasis [Single Cell Multiome ATAC + Gene Expression]

city: Basel

first: Costanza

last: Borrelli

address: Mattenstrasse 26

country: Switzerland

institute: ETH Zurich

summary: It is estimated that only 0.02% of disseminated tumor cells are able to seed overt metastases1. While this suggests the presence of environmental constraints to metastatic seeding, the landscape of host factors controlling this process remains largely unknown. Combining transposon technology2 and fluorescent niche labeling3, we developed an in vivo CRISPR activation screen to systematically investigate the interactions between hepatocytes and metastatic cells. Our approach enabled the identification of Plexin B2 as a critical host-derived regulator of liver colonization in colorectal, pancreatic cancer and melanoma syngeneic mouse models. We dissect a mechanism by which Plexin B2 interacts with class IV semaphorins on tumor cells, leading to Klf4 upregulation and thereby promoting the acquisition of epithelial traits. Our findings highlight the essential role of signals from the liver parenchyma for the seeding of disseminated tumor cells, prior to the establishment of a growth promoting niche. They further suggest that epithelialization is required for the adaptation of CRC metastases to their new tissue environment. Blocking the Plexin B2-semaphorin axis abolishes metastatic colonization of the liver and thus represents a new therapeutic strategy for the prevention of hepatic metastases. Finally, our screening approach, which evaluates host-derived extrinsic signals rather than tumor-intrinsic factors for their ability to promote metastatic seeding, is broadly applicable and lays a framework for the screening of environmental constraints to metastasis in other organs and cancer types.

overall_design: Single-cell RNA sequencing (10x) of AKPS organoids treated with recombinant mouse Plexin B2 or vehicle

first: Andreas

last: Moor


title: In vivo interaction screening reveals liver-derived constraints to metastasis [scRNA-Seq]


title: Identification of an alternative ligand binding pocket in PPAR-γ and its corelated selective agonist for promoting beige adipocyte differentiation

city: nanjing

first: qiang

last: tian

address: School of Life Sciences, Nanjing University, Nanjing, 210023, China.

department: the school of life sciences

institute: nanjing university

summary: Pharmacological activation of peroxisome proliferator-activated receptor gamma (PPAR-γ) is a convenient and promising tactic for promoting beige adipocyte biogenesis to combat obesity-related metabolic disorders. However, thiazolidinediones (TZDs), the full agonist of PPAR-γ exhibits severe side effects in animal model and clinical uses. Therefore, it is emerging to develop efficient and safe PPAR-γ modulators for metabolic disease treatment. Here, by utilizing comprehensive methods, we report a previously unidentified ligand binding pocket (LBP) in PPAR-γ and link it to beige adipocyte differentiation. Further virtual screening from 4097 natural compounds based on this novel LBP, we discover NJT-2, a terpenoid compound, can bind to PPAR-γ induce co-activator recruitment and effectively activate PPAR-γ mediated transcription of beige adipocyte program. Importantly, in mouse model, NJT-2 administration efficiently promotes beige adipocyte biogenesis and improve obesity-associated metabolic dysfunction with significant lower adverse effects than those observed in TZD. Our results not only provide an advanced molecular insight into the structural ligand binding details in PPARg, but also develop its linked selective and safe agonist for obesity treatment.

overall_design: To explore the machanism of NJT-2 on promotion of biege cell, we performed the unbiased RNA-Seq to detailed described the metabolic phenotype of NJT-2 treated adipocytes.

first: Qiang

last: Tian

first: Miaohua

last: Wang

first: Xueting

first: Owais

last: Ahmad

first: Dianhua

last: Chen

first: Wei

last: Zheng

first: Pingping

last: Shen

first: Nanfei

last: Yang


title: Impact of Tlr7-knockdown on gene expression of 4T1 spheroids.

city: Berlin

first: Benjamin

middle: N

last: Ostendorf

address: Augustenburger Platz 1

institute: Charité University Medicine

summary: We here profiled the transcriptional changes imparted by knockdown of Tlr7 in spheroids formed by the murine breast cancer cell line 4T1. Knockdown of Tlr7 in 4T1 spheroids led to downregulation of genes implicated in cancer cell invasion, proliferation, and metastasis.

overall_design: We established spheroids from 4T1 cells transduced either with non-targeting control shRNA or an shRNA leading to knockdown of Tlr7. Subsequently, transcriptional profiling was performed.

first: Veena

last: Padmanaban


title: RNA-guided RNA silencing by an 1 Asgard archaeal Argonaute

city: Wageningen

first: Daan

middle: C.

last: Swarts

address: Stippeneng 4

department: Laboratory of Biochemistry

institute: Wageningen University

type: Non-coding RNA profiling by high throughput sequencing

summary: The dataset contains small RNAs that associated with HrAgo1 during heterologous expression in E. coli. The goal of the study was to determine what type of small RNAs associate with HrAgo1 and from what RNA transcripts these small RNAs are derived

overall_design: His-MBP-tagged-HrAgo1 was heterologously expressed under the T7 promoter in E. coli strain BL21-Gold (DE3). His-MBP-tagged-HrAgo was purified using niNTA purification and size-exclusion chromatography. Nucleic acids were extracted from HrAgo1 and a small RNA library was generated and sequenced by GenomeScan (leiden, NL). Detailed methods are given in the associated publication.

first: Carolien

last: Bastiaanssen

first: Fabai

last: Wu

middle: C


title: MUC1-C DEPENDENCE FOR PROGRESSION OF PANCREATIC NEUROENDOCRINE TUMORS IDENTIFIES A DRUGGABLE TARGET FOR TREATMENT OF THIS RARE CANCER

city: Brookline

first: Hiroki

last: Ozawa

address: 450 Brookline

institute: Dana-Farber Cancer Institute

summary: Patients with rare pancreatic neuroendocrine tumors (pNETs) have limited access to effective targeted agents and invariably succumb to progressive disease. MUC1-C is a druggable oncogenic protein linked to driving pan-cancers. There is no known involvement of MUC1-C in pNET progression. The present work was performed to determine if MUC1-C represents a potential target for advancing pNET treatment. We demonstrate that the MUC1 gene is upregulated in primary pNETs that progress with metastatic disease. In pNET cells, MUC1-C drives E2F- and MYC-signaling pathways necessary for survival. Targeting MUC1-C genetically and pharmacologically also inhibits self-renewal capacity and tumorigenicity. Studies of primary pNET tissues further demonstrate that MUC1-C expression associates with (i) advanced NET grade and pathological stage, (ii) metastatic disease, and (iii) decreased disease-free survival. These findings demonstrate that MUC1-C is necessary for pNET progression and is a novel target for treating these rare cancers with anti-MUC1-C agents under clinical development.

overall_design: RNA Seq in pNET Cell line with MUC1-C shRNA silencing. We conpared DMSO and DOX treatment cells for differential gene expression analysis. MUC1shRNA (MISSION shRNA TRCN0000122938; Sigma, St. Louis, MO, USA)  was inserted into the pLKO-tet-puro vector (Plasmid #21915; Addgene, Cambridge, MA, USA) . Vector-transduced cells were selected for growth in 1–2 μg/ml puromycin. Cells were treated with 0.1% DMSO as the vehicle control or 500 ng/ml doxycycline (DOX; Millipore Sigma).


title: Selective depletion of HBV-infected hepatocytes by class A capsid assembly modulators require high levels of intrahepatic HBV core protein

city: Foster City

first: Cameron

last: Soulette

address: 333 Lakeside Dr

institute: Gilead Sciences

summary: Several classes of capsid assembly modulators (CAMs) are currently being developed for chronic hepatitis B (CHB) cure. Both class A (CAM-A) and class E (CAM-E) CAMs disrupt nucleocapsid assembly and reduce extracellular hepatitis B virus (HBV) DNA. However, only CAM-As have been shown to reduce the number of HBV-infected cells in the animal models. However, there has been limited efficacy to date of CAM-A molecules achieving this secondary mechanism of HBV-infected cell clearance in CHB clinical trials. To investigate this disconnect, we performed comparative experiments with tool compounds from each class to further explore these unique features and antiviral activity of CAM-A across HBV-infected primary human hepatocytes (PHH), as well as in two different HBV mouse models (immunodeficient mice repopulated with human hepatocytes and AAV-HBV). Mechanistic studies in HBV-infected PHH revealed that CAM-A, but not CAM-E, induced dose-dependent aggregation of HBV core protein (HBc) in the nucleus. Experiments with siRNA, resulted in identification of the ubiquitin-binding protein p62 as a factor negatively regulating the size of these aggregates. Furthermore, we found that only the CAM-A is able to induce HBc-positive cell death in vivo with the loss of HBV-infected cells positively correlated to the levels of intrahepatic HBc. Profiling of intrahepatic HBc levels across CHB patient liver biopsies demonstrated a significantly lower level of HBc per hepatocyte than either of the HBV mouse models. Taken together, these data demonstrate that CAMs of class A have a unique secondary mechanism that has potential to specifically affect viability of HBV-infected hepatocytes. At the same time, the clearance of infected hepatocytes may depend on the level of HBc expression thereby limiting the therapeutic potential for this class of molecules.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: Gene expression profile at single cell level of fixed CD55+ myeloid progenitors

city: Liege

first: Christophe

last: Desmet

address: B34, Avenue de l'Hopital 1

country: Belgium

institute: University of Liege

summary: Murine bone marrow CD55+ myeloid progenitors were fixed and submitted to Flex ScRNA-seq with CITE-seq in order to investigate eosinophil ontogeny and development.

overall_design: Murine bone marrow progenitors were fixed, CD55+ myeloid were sorted by FACS and submitted to the Flex 10x Genomics pipeline.

first: Joseph

last: Jorssen

first: Glenn

last: Van Hulst


title: Gene expression profile at single cell level of SiglecF positive bone marrow leukocytes.

summary: SiglecF positive bone marrow leukocytes were submitted to SCRNAseq in order to investigate eosinophil development and amplification during eosinophilia.

overall_design: SiglecF positive bone marrow leukocytes were sorted by FACS and submitted to the 10x Genomics pipeline.


title: Uranium-bearing dust induces differentiation and expansion of enteroendocrine cells In human colonoids

city: Albuquerque

first: Julie

middle: Goeun

last: In

state: NM

address: 915 Camino de Salud, IDTC 3270

institute: University of New Mexico

summary: Chronic exposure to environmental toxins and heavy metals has been tenuously linked to intestinal inflammation, increased susceptibility to pathogen-induced diseases, and higher incidences of colorectal cancer, all of which have been steadily increasing in prevalence for the past 40 years. The negative effects of heavy metals on barrier permeability and inhibition of intestinal epithelial healing have been described; however, transcriptomic changes within the intestinal epithelial cells and impacts on lineage differentiation are largely unknown. Uranium exposure remains an important legacy environmental and physiological health concern, with hundreds of abandoned uranium mines located in the Southwestern United States largely impacting underserved indigenous communities. Human colonoids were used to define the molecular and cellular changes that occur in response to uranium bearing dust (UBD) exposure. Here, we use single cell RNA sequencing to characterize the molecular changes that occur to proliferative and differentiated intestinal epithelial cells exposed to UBD. We demonstrate that this environmental toxicant disrupts proliferation and induces hyperplastic differentiation of secretory lineage cells, specifically enteroendocrine cells (EEC). These cell types show increased differentiation into de novo EEC sub-types not found in control colonoids. These findings highlight the significance of epithelial differentiation as major colonic responses to heavy metal-induced injury.

overall_design: Human colonoids were kept in high WNT3A/Rspo1/Noggin media and separated into control and uranium-bearing dust (UBD) conditions. UBD was added for 18h, then washed out. Colonoids were processed into single cells using enzymatic digestion then analyzed using the 10X chromium platform

first: Roger

last: Atanga


title: Study of the impact of Anti-IL5 neutralizing antibodies on bone marrow eosinophil maturation stages during IL-33 induced eosinophilia.

summary: RNA-seq of the 4 eosinophil maturation stages in the murine bone marrow during Interleukin-33 induced eosinophilia combined with Anti-IL5 or Control isotype injections.

overall_design: The 4 stages of eosinophil maturation (from I to IV) were sorted via FACS from the bone marrow of mice after IL-33 induced eosinophilia with additional treatment with Anti-IL5 or Control isotype.


title: RNA-seq of the 4 stages of maturation of eosinophils in the murine bone marrow in the steady state and in Interleukin-33- induced eosinophilia.

summary: Study of the transcriptional changes occurring during eosinophil maturation in the steady state and in IL-33 induced eosinophilia.

overall_design: Biological triplicates of the 4 stages of eosinophil maturation (from I to IV) were sorted via FACS from the bone marrow of mice in the steady state and in IL-33 induced eosinophilia.


title: Incremental increases in physiological fluid shear progressively alter pathogenic phenotypes and gene expression in multidrug resistant Salmonella

city: Tempe

first: Jiseon

state: AZ

address: 1001 S. McAllister Ave

institute: Arizona State University

summary: The ability of bacteria to sense and respond to mechanical forces has important implications for pathogens during the in vivo infection process, as they experience wide fluid shear fluctuations in the host.  However, relatively little is known about how mechanical forces encountered in the infected host drive microbial pathogenesis.  We previously demonstrated an inverse relationship between fluid shear-induced responses of classic gastrointestinal disease-causing Salmonella Typhimurium (x3339) and systemic multidrug resistant (MDR) S. Typhimurium (ST313 D23580) when the organisms were cultured under fluid shear forces like those in the intestinal tract and bloodstream.  To advance our understanding of how incremental increases in physiological fluid shear impact D23580 pathogenesis phenotypes and transcriptomic responses, we applied dynamic bioreactor technology to introduce and quantify incremental increases in fluid shear during culture. Our data indicate that D23580 responds dynamically to a range of physiological fluid shear levels by altering pathogenesis-related phenotypes (stress responses, host cell colonization) and transcriptomic responses (including genes important for adherence and invasion).  These phenotypic and molecular genetic changes directly correlated with incrementally increased fluid shear. This is the first demonstration that incremental changes in fluid shear alter stress responses and gene expression in any ST313 strain and offers new insight into how physiological fluid shear forces encountered by MDR bacteria in the infected host might impact their disease-causing ability in unexpected ways.

sample_organism: Salmonella enterica subsp. enterica serovar Typhimurium str. D23580

overall_design: RNA-Seq was performed on an Illumina HiSeq-2000 sequencer to profile the entire D23580 transcriptome. Three RNA sample types were processed: no bead (FS1), 1/8-PolyPropylene bead (FS3), and 1/8-Ceramic bead (FS4).  For all three RWV conditions (FS1, FS3 and FS4), the RNA samples were prepared from two sets of biological replicates of D23580 cultures grown in LB medium to stationary phase for 24 hours at 37 °C.

first: Jennifer

last: Barrila

first: Eric

middle: A

last: Nauman

first: Seth

middle: D

last: Nydam

first: Shanshan

first: Jin

last: Park

first: Ami

last: Gutierrez-Jensen

first: Christian

middle: L

last: Castro

first: C

last: Ott

first: Kristina

last: Buss

first: Jason

last: Steel

first: Cheryl

last: Nickerson


title: Bulk and single-cell RNA sequencing of human and murine eosinophil lineage cells

first: Christophe

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

overall_design: Refer to individual Series

first: Fabrice

last: Bureau

first: Jo

last: Caers

first: Frederic

last: Baron


title: Short-chain fatty acids abrogate Japanese encephalitis virus-induced inflammation in microglial cells via miR200a-3p/ZBTB20/Ikβα axis

status: Public on May 20 2024

city: Gurugram

first: Anirban

last: Basu

state: Haryana

address: Nainwal Mor, Manesar

department: Cellular and molecular neuroscience

country: India

institute: National Brain Research Centre

summary: Japanese encephalitis virus (JEV), a member of the Flaviviridae family, is a leading cause of viral encephalitis in humans. Survivors of this infection often develop life-long neurological sequelae. Short-chain fatty acids (SCFAs) produced in the gut are vital mediators of the gut-brain axis. We aimed to study microRNA-based mechanisms of SCFAs in an in vitro model of JEV infection. N9 microglial cells were pre-treated with SCFA cocktail before JEV infection. Cytokine bead analysis (CBA), immunoblotting and PCR were performed to analyse relevant inflammatory markers. microRNA sequencing was performed using Illumina Hiseq and Bioinformatical tools were used for differentially expressed (DE) miRNAs and Weighted gene co-expression network analysis (WGCNA). microRNA mimic/inhibitor experiments and luciferase assay were performed to study miRNA-target interaction. A significant reduction in MCP1 and TNFα along with reduced expression of phospho-NF-κB was observed in SCFA conditions. Significant attenuation of HDAC activity and protein expression was recorded. miRNA sequencing revealed 160 DE miRNAs in SCFA+JEV treated cells at 6 hours post infection (HPI). WGCNA revealed miR-200a-3p, a hub miRNA significantly upregulated in SCFA conditions. Transcription factor ZBTB20 was bioinformatically predicted and validated as a gene target for miR200a-3p. Further miRNA mimic/inhibitor assay demonstrated that miR-200-3p regulated ZBTB20 along with Iκβα that possibly dampened NF-κB signal activation downstream.

overall_design: N9 microglial cells were pre-treated with SCFA for 12 hours followed by JEV infection for 6hours and 24 hours. There were 4 groups in both the infection timeline namely, Control, SCFAControl, JEVcontrol, SCFAJEV. Total RNA extraction was performed using phenol chloroform method. Differential expression analysis in all 24 conditions was performed by small RNA sequencing (Illumina) according to the manufacturer's instructions.

first: Atreye

last: Majumdar

first: Indira

last: Priya Siva Venkatesh


title: Gene expression profile at single cell level of Bone Marrow Myeloid Progenitors

last: Desmet

summary: Murine bone marrow myeloid progenitors were submitted to SCRNAseq in order to investigate myelopoiesis

overall_design: Murine bone marrow myeloid progenitors were sorted by FACS and submitted to the 10x Genomics pipeline.

first: Joseph

last: Jorssen

first: Glenn

last: Van Hulst


title: Human CD127 negative ILC2s show immunological memory.

city: Solna

first: Itziar

last: Martinez Gonzalez

address: Biomedicum C7, Solnavägen 9

country: Sweden

institute: Karolinska Institute

summary: Group 2 Innate Lymphoid Cells (ILC2s) are key players in type 2 immunity and contribute to maintaining homeostasis and responding to inflammation. ILC2s are implicated in the development of type 2 inflammation-mediated chronic disorders like asthma. While memory ILC2s have been identified in mouse, it is unknown whether human ILC2s can acquire immunological memory. Here, we demonstrate the persistence of CD45RO, a marker previously linked to inflammatory ILC2s, in resting ILC2s that have undergone prior activation. A high proportion of these cells concurrently reduce the expression of the canonical ILC marker CD127 in a tissue specific manner. Through isolation and in vitro stimulation of CD127- CD45RO+ ILC2s, we observed an augmented ability to proliferate and produce cytokines. CD127- CD45RO+ ILC2s are found in both healthy and inflamed tissues and display a gene signature of cell activation. Our findings suggest that human ILC2s can acquire innate immune memory and warrants a revision of the current strategies to identify human ILC2s.

overall_design: Single cell sequencing of human dermal CD45+ Lin- cells

first: Sergio

last: Martinez-Høyer

first: Baltasar

last: Heesters

first: Hergen

last: Spits

last: Martinez-Gonzalez


title: Decoding Endothelial MPL and JAK2V617F Mutation: Insight into Cardiovascular Dysfunction in Myeloproliferative Neoplasms

city: STONY BROOK

first: Huichun

last: Zhan

state: NY

address: 15-089 HSC, 101 Nicolls Rd

institute: Stony Brook Medicine

summary: Background: Patients with JAK2V617F-positive myeloproliferative neoplasms (MPNs) and clonal hematopoiesis of indeterminate potential (CHIP) face a significantly elevated risk of cardiovascular diseases (CVDs). Endothelial cells (ECs) carrying the JAK2V617F mutation have been detected in many MPN patients. In this study, we investigated the molecular basis for the high incidence of cardiovascular complications in MPN patients. Methods: We investigated the impact of endothelial JAK2V617F mutation on CVD development using both transgenic murine models and MPN patient-derived induced pluripotent stem cell lines. Results and Conclusions: Our investigations revealed that JAK2V617F mutant ECs promote CVDs by impairing endothelial function and undergoing endothelial-to-mesenchymal transition (EndMT). Importantly, we discovered that inhibiting the endothelial thrombopoietin receptor MPL suppressed JAK2V617F-induced EndMT and prevented cardiovascular dysfunction caused by mutant ECs. Notably, the endothelial MPL receptor is not essential for the normal physiological regulation of blood cell counts and cardiac function, rendering it a promising therapeutic target for preventing or ameliorating cardiovascular complications in patients with MPNs.

overall_design: JAK2 wild-type and JAK2V617F mutant cardiac ECs (CD45-CD31+ ) were isolated from Tie2+FF1+ or Tie2-cre mice (n = 3 mice in each group) and their transcriptomic profiles were previously assessed using RNA sequencing1 . RNA sequencing was also conducted on JAK2 wild-type cardiac ECs (one pooled sample from 3 mice) and JAK2V617F mutant cardiac ECs (one pooled sample from 3 mice) isolated from CDH5+FF1+ mice and CDH5+FF1- control mice following high-fat diet treatment, and on human iPS-derived JAK2 wild-type and JAK2V617F mutant ECs treated with or without AMM2 between passage 3 and passage 5 (~15 days). Briefly, total RNA was extracted using the RNeasy mini kit (Qiagen, Hilden, Germany). The isolated RNA samples were sent to Novogene, Inc. for bulk RNA sequencing. Messenger RNA was purified from total RNA using poly-T oligo-attached magnetic beads and used to generate sequencing libraries using NEBNext Ultra™ RNA Library Prep Kit for Illumina® (New England BioLabs, MA, USA). The library preparations were sequenced on an Illumina platform. Index of the reference genome was built using Hisat2 v2.0.5 and paired-end clean reads were aligned to the reference genome. featureCounts v1.5.0-p3 was used to count the reads numbers mapped to each gene. Differential expression analysis between two groups was performed using the edgeR R package (3.22.5). The resulting P-values were adjusted using the Benjamini and Hochberg’s method for controlling the false discovery rate. Genes with an adjusted P-value < 0.05 found by edgeR were assigned as differentially expressed. Gene set enrichment analysis (GSEA) was performed using the complete list of genes according to their differential expression. Gene Ontology (GO) enrichment analysis of differentially expressed genes was implemented by the ClusterProfiler R package. GO terms with corrected P-value less than 0.05 were considered significantly enriched.

first: Haotian

last: Zhang

first: Nicholas

last: Kafeiti

first: Kyla

last: Masarik

first: Sandy

last: Lee

first: Xiaoxi

last: Yang

first: Haoyi

last: Zheng


title: Desert Hedgehog Downregulation Mediates Proliferative Inhibition by γ-Glutamylcyclotransferase Knockdown in Murine Glioblastoma Stem Cells

city: Kyoto

first: Hiromi

last: Ii

state: Kyoto

address: 5 Nakauchi-cho, Misasagi, Yamashina-ku,

institute: Kyoto Pharmaceutical University

summary: Glioblastoma is the most frequent type of adult-onset malignant brain tumors and has a very poor prognosis. A cell population called glioblastoma stem cells has been shown to be one of the mechanisms by which glioblastoma acquires therapy resistance. Therefore, there is a need to establish novel therapeutic strategies useful for inhibiting the cell population. γ-glutamylcyclotransferase (GGCT) is an enzyme involved in the synthesis and metabolism of glutathione, which is highly expressed in a wide range of cancer types including glioblastoma, and inhibition of its expression has been reported to have antitumor effects on various cancer types. In this study, we searched for pathways affected by the growth-promoting effect of GGCT overexpression in mouse embryonic fibroblasts NIH3T3 by comprehensive gene expression analysis and found the hedgehog pathway. Knockdown of GGCT inhibited proliferation of glioblastoma stem cells derived from a mouse glioblastoma model and caused decreased expression of desert hedgehog (Dhh), a representative ligand of the pathway, and the downstream target Gli1. To analyze the contribution of reduced Dhh, we activated the hedgehog pathway by forced expression of Dhh. We found that Dhh overexpression significantly restored the growth suppressive effect of GGCT knockdown. The results of this study indicate that high GGCT expression is important for Dhh expression and activation of the hedgehog pathway, which is required to maintain glioblastoma stem cell proliferation. Therefore, inhibition of GGCT function may be useful in suppressing stemness of glioblastoma stem cells accompanied with activation of the hedgehog pathway.

overall_design: To test the effect of overexpression of GGCT on the gene expression, mouse embryonic fibroblast cell line NIH3T3 was transfected with GGCT expression retroviral vector or control vector. Three independent experiments were performed and GGCT-overexpressing NIH3T3 cells were compared to control cells.

first: Masaya

last: Mori

first: Mitsugu

last: Fujita

first: Kozue

last: Nose

first: Ayako

last: Shimada

first: Risa

last: Shiraki

first: Yuhi

last: Sone

first: Chiami

last: Moyama

first: Keiko

last: Taniguchi

first: Susumu

last: Nakata


title: PanIN and CAF Transitions in Pancreatic Carcinogenesis Revealed with Spatial Data Integration [Xenium]

city: Baltimore

first: Luciane

middle: Tsukamoto

last: Kagohara

state: MD

address: 1650 Orleans St - Rm 485

department: Oncology

institute: Johns Hopkins University

summary: This study introduces a new imaging, spatial transcriptomics (ST), and single-cell RNA-sequencing (scRNA-seq) integration pipeline to characterize neoplastic cell state transitions during tumorigenesis. Our semi-supervised analysis pipeline was applied to examine pancreatic intraepithelial neoplasias (PanIN), the most frequent premalignant lesions that can develop into pancreatic adenocarcinoma (PDAC). Their strict diagnosis on FFPE samples has limited previous characterization of human PanINs within their microenvironment through single-cell approaches. We leverage unbiased whole transcriptome FFPE spatial profiling to enable this characterization in a rare cohort of matched low-grade and high-grade PanIN lesions to track progression and map cellular phenotypes relative to scRNA-seq data of advanced PDAC tumors. We demonstrate that cancer associated fibroblasts (CAF), including antigen-presenting CAFs (apCAF), are located close to PanINs. We further observed a transition from CAF-related inflammatory signaling to cellular proliferation during PanIN progression. We use these findings to guide panel design to perform single-cell validation with high-dimensional imaging proteomics and transcriptomics technologies. Altogether, our pipeline for spatial multi-omics characterization provides a resource for future PanIN studies. Moreover, our semi-supervised learning framework to spatial multi-omics has broad applicability across cancer types to decipher the spatiotemporal dynamics of carcinogenesis.

overall_design: 5 tissue sections from surgical sections of resected primary pancreatic ductal adenocarcinoma from 3 unique human subjects, which are Xenium data inputs to recreate results of validation experiments conducted as an extension of analyses conducted with GSE254829.

first: Alexander

middle: T

last: Bell

first: Jacob

last: Mitchell

first: Ashley

last: Kiemen

first: Elana

last: Fertig


title: The Hippo pathway transcription factors YAP and TAZ play HPV-type dependent roles in cervical cancer

city: Leeds

first: Andrew

last: Macdonald

state: West Yorkshire

address: Faculty of Biological Sciences

department: School of Molecular and Cellular Biology

country: United Kingdom

institute: University of Leeds

summary: Human papillomaviruses (HPVs) cause most cervical cancers and an increasing number of other anogenital and oral carcinomas, with most cases caused by HPV16 or HPV18. HPV hijacks host signalling pathways to promote proliferation, to drive carcinogenesis. Understanding these interactions could permit identification of much-needed therapeutics for HPV-driven malignancies. The Hippo signalling pathway is important in HPV+ cancers, with the downstream effector YAP playing a pro-oncogenic role. However, the significance of its paralogue TAZ remains largely uncharacterised in these cancers. We demonstrate that TAZ is dysregulated in a HPV-type dependent manner by a distinct mechanism to that of YAP and controls proliferation via alternative cellular targets. Analysis of cervical cancer cell lines and patient biopsies revealed that TAZ expression was only significantly increased in HPV18+ and HPV18-like cells and. TAZ knockdown reduced proliferation, migration, and invasion only in HPV18+ cells. RNA-sequencing of HPV18+ cervical cells revealed that YAP and TAZ have distinct targets, suggesting they promote different mechanisms. Thus, in HPV18+ cancers, YAP and TAZ play non-redundant roles. This analysis identified TOGARAM2 as a previously uncharacterised TAZ target and demonstrates its role as a key effector of TAZ-mediated proliferation, migration, and invasion in HPV18+ cancers.

overall_design: Stable monoclonal knockdown HeLa cells were generated using shRNA targeting YAP and WWTR1 individually. Two shRNA, targeting different sites, were utilised per gene to account for potential off-target effects. Knockdown stable cell lines were compared with a negative shRNA and analysed by RNA-Sequencing.

first: Molly

middle: R

last: Patterson

first: Joseph

last: Cogan

first: Rosa

last: Cassidy

first: Daisy

last: Theobald

first: Miao

last: Wang

first: James

last: Scarth

first: Chinedu

last: Anene

first: Adrian

last: Whitehouse

first: Ethan

last: Morgan


title: RNA-seq of the 4 stages of maturation of eosinophils in healthy human bone marrow.

first: Christophe

last: Desmet

summary: Study of the transcriptional changes occurring during eosinophil development in healthy human bone marrow.

overall_design: Biological triplicates of the 4 stages of eosinophil maturation (from I to IV) were sorted via FACS from human bone marrow and analyzed by RNA-seq to allow studying transcriptomic changes along human eosinophil development.

last: Jorssen

first: Glenn

last: Van Hulst


title: Epitranscriptomic cytidine methylation of the Hepatitis B viral RNA encapsidation signal ensure the reverse transcription of viral RNA [PA-m5C-seq 2]

city: Taipei

first: Kevin

last: Tsai

address: 128 Sec. 2, Academia Rd. Nankang

department: Institute of Biomedical Sciences

country: Taiwan

institute: Academia Sinica

summary: Epitranscriptomic RNA modifications have emerged as important regulators of the fate and function of both cellular and viral RNAs. One prominent modification, the cytidine methylation 5-methylcytidine (m5C), is found on the RNA of HIV-1, where m5C enhances the translation and splicing of HIV-1 RNA. However, whether m5C functionally enhances the RNA of other pathogenic viruses remain elusive. Here, we report that the RNA of hepatitis B virus (HBV) is enriched with a high level of m5C, mediated mainly through the cellular methyltransferase NSUN2. Intrigingly, the most prominent cluster of NSUN2-deposited m5C is found on the epsilon hairpin, an RNA element required for viral RNA encapsidation and reverse transcription. Loss of m5C from HBV RNA due to depletion of NSUN2 resulted in a modest decrease in viral capsid protein (HBc) translation, yet this is accompaneied by a near-complete loss of the reverse transcribed viral DNA. Similarly, mutations introduced to remove the methylated cytidines resulted in a translation decrease and block of reverse transcription. Furthermore, pharmacological disruption of m5C deposition with a nucleoside analogue led to a significant decrease in HBV replication. Thus, our data indicates m5C methylations is a critical enhancer of the epsilon element in HBV reverse transcription. Our study suggests the theraputic potential of targeting the m5C methyltransfer process on the HBV 5’epsilon as an alternative antiviral stratagy.

overall_design: PA-m5C-seq: Photoactivated UV crosslinking of m5C antibody to m5C+ RNA, followed by immunoprecipitation and sequencing. RNA from HuH-7 cells, transfected with HBV replicon plasmid, pulsed with 6-thioguanosine (6SG).

first: Pei-Yi

last: Su


title: Single Cell Analysis of Treatment–Resistant Prostate Cancer: Implications of Cell State Changes for Cell Surface Antigen Targeted Therapies

city: Ithaca

first: Jooyoung

last: Park

address: 237 Tower Rd

institute: Cornell University

summary: Targeting cell surface molecules using radioligand and antibody–based therapies has yielded considerable success across cancers.  However, it remains unclear how the expression of putative lineage markers, particularly cell surface molecules, varies in the process of lineage plasticity, wherein tumor cells alter their identity and acquire new oncogenic properties.  A notable example of lineage plasticity is the transformation of prostate adenocarcinoma (PRAD) to neuroendocrine prostate cancer (NEPC)––a growing resistance mechanism that results in the loss of responsiveness to androgen blockade and portends dismal patient survival.  To understand how lineage markers vary across the evolution of lineage plasticity in prostate cancer, we applied single cell analyses to 21 human prostate tumor biopsies and two genetically engineered mouse models, together with tissue microarray analysis (TMA) on 131 tumor samples.  Not only did we observe a higher degree of phenotypic heterogeneity in castrate–resistant PRAD and NEPC than previously anticipated, but also found that the expression of molecules targeted therapeutically, namely PSMA, STEAP1, STEAP2, TROP2, CEACAM5, and DLL3, varied within a subset of gene–regulatory networks (GRNs).  We also noted that NEPC and small cell lung cancer (SCLC) subtypes shared a set of GRNs, indicative of conserved biologic pathways that may be exploited therapeutically across tumor types.  While this extreme level of transcriptional heterogeneity, particularly in cell surface marker expression, may mitigate the durability of clinical responses to novel antigen–directed therapies, its delineation may yield signatures for patient selection in clinical trials, potentially across distinct cancer types.

overall_design:
Tumor biopsies were dissociated per previously published protocols (PMID: 35981096). Single cell data for HMP22, HMP23A, HMP23B, HMP24 has not previously been published and is deposited under this GEO ID. Data for HMP04, HMP05, HMP08, HMP11A, HMP11B, HMP13, HMP14, HMP16, HMP17, HMP19, HMP20, HMP25, and HMP26 were re–analyzed, but has previously been published in the context of JAK/STAT signaling in prostate cancer under GSE210358 (PMID 35981096). Data from HP95, HP96, HP97, HP99, HP100, and HP101 are also re–analyzed and has previously been published by our group in the context luminal regeneration under https://duos.broadinstitute.org/ (accession no. DUOS-000115). We have also included processed RDS files for combined samples (N=23) for all cells and tumor cells. Of note, HP and HMP IDs correspond to MSK–HP IDs as presented in this publication in Supplementary Table 3.

The list of the re-analyzed sample in GSE210358:
SRX16781303 GSM6428952 HMP04 SAMN30111353
SRX16781304 GSM6428953 HMP05 SAMN30111352
SRX16781305 GSM6428954 HMP08 SAMN30111351
SRX16781314 GSM6428955 HMP11_1       SAMN30111350
SRX16781315 GSM6428956 HMP11_2       SAMN30111349
SRX16781316 GSM6428957 HMP13 SAMN30111348
SRX16781317 GSM6428958 HMP14 SAMN30111347
SRX16781319 GSM6428960 HMP16 SAMN30111345
SRX16781318 GSM6428959 HMP15 SAMN30111346
SRX16781320 GSM6428961 HMP17 SAMN30111344
SRX16781321 GSM6428962 HMP19 SAMN30111343
SRX16781322 GSM6428963 HMP20 SAMN30111342
SRX16781323 GSM6428964 HMP25 SAMN30111341
SRX16781325 GSM6428965 HMP26 SAMN30111340

first: Samir

last: Zaidi

first: Michael

last: Haffner

first: Charles

last: Sawyers


title: Hair follicles modulate skin barrier function

city: Ann Arbor

first: Sunny

last: Wong

state: MI

address: 3217 Cancer Center

department: Dermatology

institute: University of Michigan

summary: We generated KA mutant mice (Krt14-CreERT + ABCA12-flox/flox) and controls (ABCA12-flox/flox) that enabled us to assess the effects of deleting a critical barrier gene, ABCA12, in the skin epidermis following tamoxifen treatment.  These studies identified numerous differentially expressed genes (DEGs) upon barrier disruption/ABCA12 deletion, including compensatory upregulation of genes involved with epidermal differentiation and barrier function.  Overall, this data-set provides a global view of molecular changes that occur in adult mouse skin following barrier perturbation.

overall_design: KA mice or controls were induced with tamoxifen at 8 weeks of age.  Two weeks later, disease features were apparent in mutants, and total RNA was isolated from separated telogen skin epidermis, and submitted for bulk RNA-sequencing.  Skin from 4 KA mutants (# 4854, 4855, 4876, 4877) and 3 controls (# 4857, 4853, 4875) were compared.

first: Noah

last: Ford

first: Rachel

middle: E

last: Benedeck

first: Matthew

last: Mattoon

first: Jamie

middle: K

last: Peterson

first: Arlee

last: Mesler

first: Natalia

last: Veniaminova

first: Danielle

last: Gardon

first: Shih-Ying

first: Yoshikazu

last: Uchida

middle: Y


title: Eomes expression identifies the early bone marrow precursor to classical NK cells

city: Chicago

first: Bana

last: Jabri

state: Illinois

address: 900 E 57th Street

department: Department of Medicine

institute: University of Chicago

summary: Classical NK cells are the prototypical type 1 innate lymphocytes mounting cytolytic and IFNg responses to intracellular pathogens and tumors. A similar but distinct population termed ILC1 was recently described, differing from NK cells in multiple though relatively subtle ways, including tissue residency vs recirculation, and levels of NK receptors. Whether the differences reflect distinct lineages or mere variations in program expression is not fully understood. Here, using transcription factor reporter mice and cell transfers of bone marrow precursors, we found that, upon transfer in vivo, Eomes-expressing, lineage-negative, NK receptor-negative cells acquired properties typically associated with classical NK cells including the capacity to prevent metatstatic disease. In contrast, as previously reported, PLZFhigh cells mostly generated ILC1, ILC2, or ILC3. These findings identify the Eomes-expressing NK precursor as the long-elusive bone marrow precursor to classical NK cells, and demonstrate that the NK and ILC1 lineages diverge early during development.

overall_design: Tcf7+ precursors, EomeshighNKneg cells, and mature NK/ILC1s were sorted from three pooled cohorts of EomesGFP+/- Tcf7mCherry+/- mice (n=11, n=12, n=12). In each pooled sample, cell populations were mixed at a ratio of 15:3:2 for Tcf7+ precursors, EomeshighNKneg cells, and mature NK/ILC1s, respectively, then profiled using scRNA-seq.

first: Zhitao

last: Liang

first: Hope

last: Anderson

first: Veronica

last: Locher

first: Crystal

last: O'Leary

first: Samantha

last: Riesenfeld

first: Benjamin

last: McDonald

first: Albert

last: Bendelac


title: High-density resolution of the Kaposi’s Sarcoma associated Herpesvirus transcriptome identifies novel transcript isoforms generated by long-range transcription and alternative splicing [iSLK-WTBAC-PCR-cDNA-Seq]

city: Gainesville

first: Rolf

last: Renne

state: FL

address: 1200 Newell Dr.

institute: University of Florida

summary: Kaposi’s sarcoma-associated herpesvirus is etiologic agent of Kaposi’s Sarcoma and two B-cell malignancies. Recent advancements in sequencing technologies have led to high resolution transcriptomes for several human herpesviruses that densely encode genes on both strands. However, for KSHV progress remained limited due to overall low percentage of KSHV transcripts even during lytic replication. To address this challenge, we have developed a target enrichment method to increase the KSHV-specific reads for both short- and long-read sequencing platforms. Furthermore, we combined this approach with the Transcriptome Resolution through Integration of Multi-platform Data (TRIMD) pipeline developed previously to annotate transcript structures. TRIMD first builds a scaffold based on long-read sequencings and validates each transcript feature with supporting evidence from Illumina RNA-Seq and deepCAGE sequencing data. Our stringent innovative approach identified 994 unique KSHV transcripts, thus providing the first high-density KSHV lytic transcriptome. We describe a plethora of novel coding and non-coding KSHV transcript isoforms with alternative untranslated regions, splice junctions and open-reading frames, thus providing deeper insights on gene expression regulation of KSHV. Interestingly, as described for Epstein-Barr Virus, we identified transcription start sites that augment long-range transcription and may increase the number of latency-associated genes potentially expressed in KS tumors.

overall_design: Multi-platform RNA sequencing of reactivated BCBL-1 cells to study the transcriptome of Kaposi's Sarcoma associated Herpesvirus on a high resolution.

first: Ritu

last: Shekhar

first: Tina

last: O'Grady

first: Netanya

last: Keil

first: April

last: Feswick

first: David

last: Moraga

first: Scott

last: Tibbets

first: Erik

last: Flemington


title: High-density resolution of the Kaposi’s Sarcoma associated Herpesvirus transcriptome identifies novel transcript isoforms generated by long-range transcription and alternative splicing [Iso-Seq]


title: Mechanosensitive Membrane Domains Regulate Calcium Entry in Arterial Endothelial Cells to Protect Against Inflammation

city: Los Angeles

first: Soon-Gook

last: Hong

state: California

address: 630 Charles E Young Dr S

department: Medicine

institute: UCLA

summary: Endothelial cells (ECs) in the descending aorta are exposed to high laminar shear stress, which supports an anti-inflammatory phenotype. High laminar shear stress also supports flow-aligned cell elongation and front-rear polarity, but whether these are required for the anti-inflammatory phenotype is unclear. Here, we show that Caveolin-1-rich microdomains polarize to the downstream end of ECs exposed to continuous high laminar flow. These microdomains are characterized by high membrane rigidity, filamentous actin (F-actin), and raft-associated lipids. Transient receptor potential vanilloid-type 4 (Trpv4) ion channels are ubiquitously expressed on the plasma membrane but mediate localized Ca2+ entry only at these microdomains where they physically interact with clustered Caveolin-1. The resultant focal Ca2+ bursts activate endothelial nitric oxide synthase (eNOS) within the confines of these domains. Importantly, we find that signaling at these domains requires both cell body elongation and sustained flow. Finally, Trpv4 signaling at these domains is necessary and sufficient to suppress inflammatory gene expression, and ectopic activation of Trpv4 channels ameliorates the inflammatory response to stimuli both in vitro and in vivo. Our work reveals a novel polarized mechanosensitive signaling hub that dampens inflammatory gene expression in arterial ECs.

overall_design:
To investigate the effect of Trpv4 channel inhibition on endothelial cell phenotype, primary human aortic endothelial cells (HAECs) were seeded on 6-well plates and subjected to uni-directional laminar flow. After 48 h, the flow-conditioned HAEC monolayers were treated with either DMSO or Trpv4-specific antagonist GSK205 (20 uM) for 2 h under flow.
We then performed gene expression profiling analysis using data obtained from RNA sequencing of three replicates per treatment condition.
Comparative gene expression profiling analysis of RNA-seq data for DMSO-control HAECs and Trpv4-antagonist GSK205-treated HAECs in the presence of laminar flow (20 dynes/cm2).

first: Julia

last: Mack


title: High-density resolution of the Kaposi’s Sarcoma associated Herpesvirus transcriptome identifies novel transcript isoforms generated by long-range transcription and alternative splicing


title: High-density resolution of the Kaposi’s Sarcoma associated Herpesvirus transcriptome identifies novel transcript isoforms generated by long-range transcription and alternative splicing [KSHV_TRIMD_Ritu_Illumina]


title: High-density resolution of the Kaposi’s Sarcoma associated Herpesvirus transcriptome identifies novel transcript isoforms generated by long-range transcription and alternative splicing [BCBL-1-direct-cDNA-Seq]


title: High-density resolution of the Kaposi’s Sarcoma associated Herpesvirus transcriptome identifies novel transcript isoforms generated by long-range transcription and alternative splicing [BCBL-1-directRNA-Seq]


title: Epitranscriptomic cytidine methylation of the Hepatitis B viral RNA encapsidation signal ensure the reverse transcription of viral RNA [PAR-CLIP]

first: Kevin

last: Tsai

overall_design: PAR-CLIP: HuH-7 cells, transfected with HBV replicon plasmid along with FLAG-GFP or FLAG-NSUN2 expression plasmids, were lysed and subject to FLAG-antibody pulldown of proteins, the co-immunoprecipitated RNA isolated and sequenced.

first: Pei-Yi

last: Su


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

city: Bethesda

first: Javed

last: Khan

address: 37 Convent Dr.

department: Genetics Branch

institute: NCI, NIH

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

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

first: Meijie

last: Tian

first: Xinyu

last: Wen

first: Jun

last: S Wei


title: Epitranscriptomic cytidine methylation of the Hepatitis B viral RNA encapsidation signal ensure the reverse transcription of viral RNA [PA-m6A-seq]

overall_design: PA-m6A-seq: Photoactivated UV crosslinking of m56Aantibody to m6A+ RNA, followed by immunoprecipitation and sequencing. RNA from HuH-7 cells, WT or NSUN2 knockout cells, transfected with or without HBV replicon plasmid.


title: Epitranscriptomic cytidine methylation of the Hepatitis B viral RNA encapsidation signal ensure the reverse transcription of viral RNA [PA-m5C-seq]

overall_design: PA-m5C-seq: Photoactivated UV crosslinking of m5C antibody to m5C+ RNA, followed by immunoprecipitation and sequencing. RNA from HuH-7 cells, WT or NSUN2 knockout cells, transfected with or without HBV replicon plasmid.


title: Epitranscriptomic cytidine methylation of the Hepatitis B viral RNA encapsidation signal ensure the reverse transcription of viral RNA [RNA-Seq]

overall_design: Total RNA-seq: Poly(A)-selected mRNAs were fragmented to ~100nt long and sequenced. RNA from HuH-7 cells, transfected with or without HBV replicon plasmid.


title: High-density resolution of the Kaposi’s Sarcoma associated Herpesvirus transcriptome identifies novel transcript isoforms generated by long-range transcription and alternative splicing [KSHV_TRIMD_Ritu_CAGE]


title: RNA-Seq data from iPSC-derived human airway cells and colon organoids infected with BANAL-20-236, WK-521, and WK-521 dFCS

city: Minato-ku

first: Kei

last: Sato

state: Tokyo

address: 4-6-1 Shiroganedai

department: Department of Microbiology and Immunology

institute: The Institute of Medical Science, The University of Tokyo

summary: iPSC-derived human airway cells and colon organoids were infected with BANAL-20-236, SARS-CoV-2 (WK-521), and FCS-deleted SARS-CoV-2 (WK-521 dFCS) to investigate virological properties and reaction of infected hosts.

overall_design: BANAL-20-236, WK-521, and WK-521 delFCS were inoculated to iPSC-derived airway cells and colon organoids. The samples were incubated for 72 hours and then were subjected to RNA extraction and RNA sequencing, respectively.

first: Shigeru

last: Fujita

first: Arnon

last: Plianchaisuk

first: Yukie

last: Kashima

first: Mai

last: Suganami

first: Jumpei

last: Ito

first: Yutaka

last: Suzuki


title: RNA-Seq data from hamster lung hila and peripheries infected with BANAL-20-236, WK-521, and WK-521 dFCS

summary: Experimental golden hamsters were infected with BANAL-20-236, WK-521 (a parental strain of SARS-CoV-2), and WK-521 dFCS (a WK-521 without furin cleavage site [FCS]) to investigate virological properties and host response in vivo.

overall_design: Experimental golden hamsters were inoculated with saline, BANAL-20-236, WK-521, and WK-521 dFCS intranasally. Lung hila and peripheries were collected from the hamsters at 2 and 5 days post-infection and then were subjected to RNA extraction and RNA sequencing, respectively.


title: Immediate myeloid depot for SARS-CoV-2 in the human lung

city: San Francisco

first: Bushra

last: Samad

state: CA

address: 505 Parnassus Ave

institute: USCF Immunoprofiler

summary: In the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic1, considerable focus has been placed on a model of viral entry into host epithelial populations, with a separate focus upon the responding immune system dysfunction that exacerbates or causes disease.  We developed a precision-cut lung slice model2,3 to investigate very early host-viral pathogenesis and found that SARS-CoV-2 had a rapid and specific tropism for myeloid populations in the human lung.  Infection of alveolar macrophages was partially dependent upon their expression of ACE2, and the infections were productive for amplifying virus, both findings which were in contrast with their neutralization of another pandemic virus, Influenza A virus (IAV). Compared to IAV, SARS-CoV-2 was extremely poor at inducing interferon-stimulated genes in infected myeloid cells, providing a window of opportunity for modest titers to amplify within these cells. Endotracheal aspirate samples from humans with the acute respiratory distress syndrome (ARDS) from COVID-19 confirmed the lung slice findings, revealing a persistent myeloid depot.  In the early phase of SARS-CoV-2 infection, myeloid cells may provide a safe harbor for the virus with minimal immune stimulatory cues being generated, resulting in effective viral colonization and quenching of the immune system.

overall_design: Human lung slices were infected with SARSCoV2 for either 24, 48 or 72h. Non-infected controls were included for each time point.Samples came from 2 different subjects and we included 3/4 slices for the experiment.

first: Mélia

last: Magnen

first: Ran

last: You

first: Arjun

last: Rao

first: Ryan

last: Davis

first: Lauren

last: Rodriguez

first: Olivier

last: Bernard

first: Camille

last: Simoneau

first: Lisiena

last: Hysenaj

first: Kenneth

middle: H

last: Hu

first: Mazharul

last: Maishan

first: Catharina

last: Conrad

first: Oghenekevwe

middle: M

last: Gbenedio

first: Christina

last: Love

first: Prescott

middle: G

last: Woodruff

first: David

last: Erle

first: Carolyn

last: Hendrickson

middle: S

last: Calfee

first: Michael

last: Matthay

first: Jeroen

middle: P

last: Roose

first: Anita

last: Sil

first: Melanie

last: Ott

first: Charles

last: Langelier

first: Matthew

middle: F

last: Krummel

first: Mark

last: Looney


title: Risk assessment of parabens in a transcriptomics-based in vitro system

city: Dortmund

first: Jan

middle: Georg

last: Hengstler

address: Ardeystraße 67

department: Toxikologie / Systemtoxikologie

institute: Leibniz-Institut für Arbeitsforschung an der TU Dortmund

summary: Parabens have been used for decades as preservatives in food, drugs and cosmetics. The majority however, were banned in 2009 and 2014 leaving only methyl-, ethyl-, propyl-, and butyl-derivates available for subsequent use. Methyl- and propylparaben have been extensively tested in vivo, with no resulting evidence for developmental and reproductive toxicity (DART). In contrast, ethylparaben has not yet been tested for DART in animal experiments, and it is currently debated if additional animal studies are warranted. In order to perform a comparison of the four currently-approved parabens, we used a previously established in vitro test based on human induced pluripotent stem cells (iPSC) that are exposed to test substances during their differentiation to neuroectodermal cells. EC50 values for cytotoxicity were 906 µM, 698 µM, 216 µM and 63 µM for methyl-, ethyl-, propyl- and butylparaben, respectively, demonstrating that cytotoxicity increases with increasing alkyl chain length. Genome-wide analysis demonstrated that FDR-adjusted significant gene expression changes occurred only at cytotoxic or close to cytotoxic concentrations, for example 1,720 differentially expressed genes (DEG) at 1,000 µM ethylparaben, 1 DEG at 316 µM, and no DEG at 100 µM or lower concentrations. The highest concentration of ethylparaben that did not induce any cytotoxicity nor DEG was 1670-fold above the highest published concentrations reported in biomonitoring studies (60 nM ethylparaben in cord blood). In conclusion, cytotoxicity and gene expression alterations of ethylparaben occurred at concentrations of approximately three orders of magnitude above human blood concentrations; moreover, the substance fitted well into a scenario where toxicity increases with the alkyl chain length, and gene expression changes only occur at cytotoxic or close to cytotoxic concentrations. Therefore, no evidence was obtained suggesting that in vivo DART with ethylparaben would lead to different results as the methyl- or propyl derivates.

overall_design: Human iPSCs were differentiated for 6 days according to a neuroepithelial protocol (UKN1) and incubated with parabens at the same time. Cytotoxicity was measured by a CellTiter-Blue® Cell Viability Assay. RNA was isolated from the cells and a whole transcritpome analysis was performed with Affymetrix HG-U133 Plus 2.0 arrays and Illumina RNA sequencing. Paraben-induced gene expression changes were determined and analyzed with statistical models.

first: Florian

last: Seidel

first: Franziska

last: Kappenberg

first: Susann

last: Fayyaz

first: Andreas

last: Scholtz-Illigens

first: Anna

last: Cherianidou

first: Katharina

last: Derksen

first: Patrick

last: Nell

first: Rosemarie

last: Marchan

first: Karolina

last: Edlund

first: Marcel

last: Leist

first: Agapios

last: Sachinidis

first: Jörg

last: Rahnenführer

first: Reinhard

last: Kreiling


title: Novel epigenetic based differentiation therapy for Acute Myeloid Leukemia

city: Pamplona

first: Xabier

last: Agirre

state: Navarra

address: Pio XII, 55

institute: CIMA

summary: Despite the development of novel therapies for acute myeloid leukemia (AML), outcomes remain poor for most patients, and therapeutic improvements are an urgent unmet need. Although treatment regimens promoting differentiation have succeeded in the treatment of acute promyelocytic leukemia (APL), their role in other AML subtypes needs to be explored. Here we identified and characterized two lysine (K) deacetylase inhibitors (DACi), CM-444 and CM-1758, exhibiting higher capacity to promote myeloid differentiation in all AML subtypes at low non-cytotoxic doses than commercial HDACi. Analyzing the acetylome after CM-444 and CM-1758 treatment revealed modulation of non-histone proteins involved in the enhancer–promoter chromatin regulatory complex, including bromodomain proteins (BRDs). This acetylation was essential for enhancing the expression of key transcription factors directly involved in the differentiation therapy provided by CM-444/CM-1758 in AML. In summary, these compounds may represent effective differentiation-based therapeutic agents across AML subtypes with a novel mechanism for treatment of AML.

overall_design: RNA-Seq study in AML cell lines treated with different HDAC inhibitors

first: Edurne

last: José-Enériz

first: Naroa

last: Gimenez-Camino

first: Obdulia

last: Rabal

first: Leire

last: Garate

first: Estibaliz

last: Miranda

first: Fernando

last: García

first: Elena

last: Sáez

first: Amaia

last: Vilas-Zornoza

first: Patxi

last: Martín-Uriz

first: Luis

middle: V

last: Valcárcel

first: Esteban

last: Tamariz-Amador

first: Nicolás

last: Martinez-Calle

first: María

middle: J

last: Larrayoz

last: Calasanz

first: Javier

last: Muñoz

first: José

middle: I

last: Martín-Subero

first: Antonio

last: Pineda-Lucena

first: Julen

last: Oyarzabal

first: Felipe

last: Prósper


title: High Throughput Plant Activation Domain Identification and Mapping

status: Public on May 19 2024

city: Durham

first: Lucia

middle: C.

last: Strader

state: NC

address: 124 Science Drive 3332 French Family Science Center

department: Biology

institute: Duke University

summary: Arabidopsis gene expression is regulated by more than 1,900 transcription factors (TFs), which have been identified genome-wide by the presence of well-conserved DNA binding domains. Activator TFs contain activation domains (ADs) that recruit coactivator complexes; however, for most Arabidopsis TFs, we lack knowledge about the presence, location, and transcriptional strength of their ADs. To address this gap, we experimentally identified Arabidopsis ADs on a proteome-wide scale, finding that over half of Arabidopsis TFs carry an AD. We annotated 1,553 ADs, the vast majority of which were previously unknown. We used the dataset generated to develop a neural network to accurately predict ADs and to identify sequence features necessary to recruit coactivator complexes. We uncovered six distinct sequence feature combinations that resulted in activation activity, providing a framework to interrogate activation domain sub-functionalization. Furthermore, we identified activation domains within the ancient AUXIN RESPONSE FACTOR (ARF) family of transcription factors, finding conservation of AD positioning in distinct clades. Our findings provide a deep resource for understanding transcriptional activation, a framework for examination of function within intrinsically disordered regions, and a predictive model of activation domains.

overall_design: Arabidopsis TFs (PADI), or ARF regions (ARF Evolution and Pilot) were fragmented into 40 amino acid tiles with a 10 amino acid step size were made as oligo pools and cloned into plasmid libraries. Yeast transformations ensured integration of a single synthetic TF, each expressing one tile, at the URA3 locus. TF activity was induced with beta-estradiol and TF activity was measured using cell sorting based on the ratio of reporter to synthetic TF (GFP:mcherry). The distribution of tiles across GFP:mCherry ratio bins was determined with Next-generation sequencing of the integrated TF. Median GFP:mcherry values were used to calculate activation domain scores.

first: Nicholas

last: Morffy

middle: C


title: Epitranscriptomic cytidine methylation of the Hepatitis B viral RNA encapsidation signal ensure the reverse transcription of viral RNA

first: Kevin

last: Tsai

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

overall_design: Refer to individual Series

first: Pei-Yi (Alma)

last: Su

first: Shin-Chwen Bruce

last: Yen


title: Histological Pattern of Tumor Inflammation and Stromal Density Correlate with Patient’s Demographics and Immuno-Oncologic Transcriptional Profile in Oral Squamous Cell Carcinoma

first: Vasileios Ionas

last: Theofilou

address: 650 W Baltimore St

department: Oncology and Diagnostic Sciences

institute: School of Dentistry, University of Maryland Baltimore

summary: The aim of this work is to demonstrate the correlation of Oral Squamous Cell Carcinoma (OSCC) tumor inflammation and stromal density with patient demographics and with the differential gene expression (DGE) analysis of 8 OSCC tumor tissue. Using the IO-360 nanoString platform, we showed that at a transcriptional level, decreased cytokine signaling and activation of oncogenic pathways were observed in immune excluded tumors and absence of peritumoral stromal inflammation. Smokers, and males correlated with decreased global levels of immune cells and predominance of myeloid populations.

overall_design:
Under an Institutional Review Board (IRB) approved protocol at the University of Maryland School of Medicine (UMSOM) (HP-00073603),  whole excision tumor tissues were collected prospectively at the day of surgery from 87 OSCC upon informed patient consent. Among the 87 tissue samples, 8 were used for RNA extraction and were processed and analyzed using  nCounter Human Pan Cancer IO-360 code set + panel standard cat # XT-CSPS-HIO360-12 and Master Kit-NAA-AKIT-012 platform that was purchased from nanoString Technology (Seattle, WA).   160 366480)

Please note that, while the samples are same as GSE163131 samples, a new record has been created due to additional characteristics which are not relevant to the GSE163131-associated publication.

first: Rania

last: Younis

first: Vasileios


title: Transcriptomics profile of adipose and adipocyte with Gls perturbation

status: Public on May 18 2024

city: Stockholm

first: Jiawei

last: Zhong

address: Blickagången 16, Flemingsberg

department: Department of Medicine, Huddinge

institute: Karolinska Institutet

summary: We generated single nucleus transcriptomic atlas for white and brown adipose tissue from adipocyte-specific Gls knockout mouse, and also generated bulk RNA-seq from in vitro differentiated human white adipocytes with/without GLS knock down.

overall_design: For snRNA-seq, brown and white adipose tissues were isolated from adipocyte-specific Gls knockout mice and Cre- neagtive Littermates and flash frozen in liquid nitrogen. The nuclei was extracted and performed snRNA-seq. For bulk RNA-seq, in vitro differentiated white adipocytes where transfected eight days post beginning of differentiation, with scrumble or GLS targeting siRNA, cells were harvested and total RNA extracted at D13 of differentiation.

first: Jutta

last: Jalkanen

first: Lucas

last: Massier

first: Gianluca

last: Renzi


title: Deficiency of lymphatic ERG and FLI1 in systemic sclerosis is associated with rarefaction of skin blood and lymphatic vasculature

city: Boston

first: Maria

last: Trojanowska

state: MA

address: 75 E. Newton St.

department: Arthritis and Autoimmune Diseases Research Center

institute: Boston University

summary:
ERG and FLI1 are important regulators of angiogenesis, but their role in lymphatic vasculature is less known. The goal of this study was to determine the role of ERG and FLI1 in postnatal lymphangiogenesis and their involvement in SSc lymphatic system defects. IF were used to detect ERG and FLI1 in SSc and healthy controls (HC) skin biopsies. Human lymphatic endothelial cells (LECs) were used in cell culture experiments. Transcriptional and chromatin analysis of ERG or FLI1 silenced LECs was performed using microarrays and ATAC-seq, respectively. Effects of ERG and FLI1 deficiency on in vitro tubulogenesis was examined using Matrigel assay. Erg and Fli1 endothelial specific knockouts (ErgCKO and Fli1CKO) and lymphatic specific knockouts (ProxErgCKO and ProxFli1CKO) were generated to examine vessel regeneration. ERG and FLI1 protein levels were reduced in blood and lymphatic vasculature in SSc skin biopsies. ERG and FLI1 were shown to regulate genes involved in lymphatic vessel specification, including VEGFR3/FLT4, LYVE-1, and PROX1, and this effect was, at least in part, due to chromatin remodeling. ERG/FLT4 pathway regulated in vitro tubulogenesis in LECs. Deficiency of Erg and Fli1 impaired function of blood and lymphatic vessel during wound healing. ERG and FLI1 are essential regulators of blood and lymphatic vessel regeneration. Deficiency of ERG and FLI1 in SSc endothelial cells, may contribute to rarefaction of blood and lymphatic vasculature in SSc patients.
Microarray analysis was performed at The Boston University Microarray Core Facility. Affymetrix CEL files were normalized to produce gene-level expression values using the implementation of the Robust Multiarray Average (RMA) in the affy package (version 1.36.1) included within in the Bioconductor software suite (version 2.12) and an Entrez Gene-specific probe set mapping from BrainArray (version 16.0.0). RLE and NUSE quality metrics were computed using the affy PLM Bioconductor package (version 1.34.0). All microarray analyses were performed using the R environment for statistical computing (version 2.15.1).

overall_design: Human Lymphatic Endothelial Cells (LECs) were transfected with 10 nM of scrambled siRNA (siSCR), or siRNAs against ERG (siERG), FLI1 (siFLI1), or both ERG and FLI1 (siERG and siFLI1) for 24 hours. After the transfection, total RNAs were extracted with TRI reagent (MRC, Inc., Cincinnati, OH). 3 samples for each siRNA treatment group were compared with Human Gene 2.0 ST arrays.

first: Yamashita

last: Takashi


title: Single-cell Total-RNA Profiling Unveils Regulatory Hubs of Transcription Factors

city: Houston

first: Chenghang

last: Zong

state: TX

address: 1 Baylor Plaza

institute: Baylor College of Medicine

summary: Recent development of RNA velocity uses master equations to establish the kinetics of the life cycle of RNAs from unspliced RNA to spliced RNA (i.e., mature RNA) to degradation. To feed this kinetic analysis, simultaneous measurement of unspliced RNA and spliced RNA in single cells is greatly desired. However, the majority of single-cell RNA-seq chemistry only captures mature RNA species to measure gene expressions. Here, we develop a one-step total-RNA chemistry-based scRNA-seq method: snapTotal-seq. We benchmarked this method with multiple single-cell RNA-seq assays in their performance in kinetic analysis of cell cycle by RNA velocity. Next, with LASSO regression between transcription factors, we identified the critical regulatory hubs mediating the cell cycle dynamics. We also applied snapTotal-seq to profile the oncogene-induced senescence and identified the key regulatory hubs governing the entry of senescence. Furthermore, from the comparative analysis of unspliced RNA and spliced RNA, we identified a significant portion of genes whose expression changes occurred in spliced RNA but not to the same degree in unspliced RNA, indicating these gene expression changes are mainly controlled by post-transcriptional regulation. Overall, we demonstrate that snapTotal-seq can provide enriched information about gene regulation, especially during the transition between cell states.

overall_design: We performed scRNA-seq with snapTotal-seq protocol on HEK293T cell line to dissect the gene regulatory network of cell cycle. We also applied snapTotal-seq to investigate the TF regulatory network driving oncogene-induced senescence. We utilized HPNE cells with inducible KRASG12D to profile the gene expression dynamics during oncogene-induced senescence. After the activation of KRASG12D with doxycycline, we collected the cells at day1, day2, day3 and day5, and performed scRNA-seq with snapTotal-seq protocol.

first: Yichi

last: Niu

first: Jiayi

last: Luo


title: Glutamine antagonist JHU083 reprograms immunosuppressive tumor-associated macrophages to drive tumor immunity in urologic cancers, single-cell RNA-seq

city: Baltimore

first: Alex

middle: Jea-Eun

last: Lee

address: 1650 Orleans Street

institute: Johns Hopkins University

summary: Glutamine metabolism in the tumor microenvironment is emerging as a critical regulator of immune-mediated anti-tumor responses. We report potent tumor growth inhibition by the glutamine antagonist prodrug JHU083 in urologic tumors by JHU083-reprogrammed tumor-associated macrophages (TAMs) and tumor-infiltrating monocytes (TIMs). Using orthogonal approaches, we show that JHU083-mediated glutamine antagonism in the tumor microenvironment induces TNF, inflammatory, and mTORC1 signaling in different intra-tumoral TAM clusters. Additionally, we report that JHU083 increases proliferation in tissue-resident macrophages intratumorally and in different TAM sub-clusters. Functionally, we report that JHU083-reprogrammed TAMs have increased tumor cell phagocytosis and diminished pro-angiogenic capacities. In vivo inhibition of glutamine consumption in TAMs results in increased glycolysis, broken TCA cycle, and disruption in purine metabolism. Although the effect of glutamine antagonism was less profound on tumor-infiltrating T cells for their anti-tumor activity, it promoted a stem cell-like phenotype in CD8+ T cells and decreased the CD4+ Treg abundance. Additionally, we report that JHU083 causes a global shutdown in glutamine utilizing metabolic pathways in tumor cells, leading to reduced HIF-1, c-MYC phosphorylation, and induction of tumor cell apoptosis, all key anti-tumoral features.

overall_design: Subcutaneous B6CaP tumors were harvested from mice treated with JHU083 or control. Tumors were dissociated and sorted based on CD45 expression with fluorescence-activated cell sorting (FACS).


title: Single cell RNA sequencing (scRNAseq) of urine-derived cells, peripheral blood mononuclear (PBMC), and tumor cells from bladder cancer patients.

status: Public on May 17 2024

city: New York

first: Michelle

middle: A

last: Tran

state: New York

address: 1470 Madison Ave

institute: Icahn School of Medicine

summary: As no one previously examined urine-derived cells from bladder cancer patients, we performed scRNAseq to profile the diversity of these cells and their transcriptional profiles. We used scRNAseq to compare the profiles of urine-derived cells to matched tumor cells and PBMC from bladder cancer patients.

overall_design:
We spun down urine-derived cells from urine, isolated PBMC using Ficoll-based separation, and dissociated cells from tumor. We analyzed each specimen individually using scRNAseq.

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Submitter states that missing raw files are due to file loss.
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first: Tran

last: Michelle

first: Bhardwaj

last: Nina

first: Sfakianos

last: John

first: Galsky

last: Matthew

first: Farkas

last: Adam


title: Glutamine antagonist JHU083 reprograms immunosuppressive tumor-associated macrophages to drive tumor immunity in urologic cancers, bulk RNA-seq

overall_design: Subcutaneous B6CaP tumors were harvested from mice treated with JHU083 or control. Tumors were dissociated and macrophages were fluorescence-activated cell sorted (FACS) by CD45+CD3-Ly6G-CD11b-F4/80+ and submitted for bulk RNA-sequencing.


title: Effect of PDD00017273 and GSK2606414 on MZ1-induced cell death

city: Shinagawa-ku

first: Fumiaki

last: Ohtake

institute: Hoshi University

summary: Targeted protein degradation is a groundbreaking modality in drug discovery ; however, further improvement of the degradation efficacy is demanded. Here, we identify small molecules that enhance the targeted degradation of the anticancer target BRD4 and related hard-to-degrade targets BRD2/3 induced by CRL2VHL- or CRL4CRBN -based PROTACs. The chemicals include inhibitors of cellular signaling pathways such as poly-ADP ribosylation (PARG inhibitor PDD00017273), unfolded protein response (PERK inhibitor GSK2606414), and protein stabilization (HSP90 inhibitor luminespib). Mechanistically, PARG inhibition  promotes TRIP12-mediated K29/K48-linked branched ubiquitylation of BRD4 by  facilitating chromatin dissociation of BRD4 and formation of the BRD4–PROTAC– CRL2VHL ternary complex; by contrast, HSP90 inhibition promotes BRD4 degradation  after the ubiquitylation step. Consequently, these signal inhibitors sensitize cells to the  PROTAC-induced apoptosis. Combining the treatment with these chemicals further  increases the efficacy of a highly potent PROTAC, SIM1. We propose that various cell intrinsic signaling pathways spontaneously counteract chemically induced target  degradation, which can be liberated by specific inhibitors.

overall_design: To further investigate the effect of PDD or GSK2606414 on the MZ1-induced apoptosis, we performed RNA-Seq analysis. HCT116 cells were treated with either PDD, GSK, or MZ1 for 24 h, and total RNA was isolated.

first: Yoshino

last: Akizuki

first: Yuki

last: Mori


title: Effect of depletion of PRMT2 on gene expression in the HL-60 acute myeloid leukemia cell line

city: Dijon

first: Laurent

last: DELVA

address: 7 boulevard Jeanne d’Arc

department: Inserm U1231 (CTM)

country: France

institute: Université de Bourgogne

summary: Arginine methylation is catalyzed by protein arginine methyltransferases (PRMTs) and is involved in various cellular processes, including cancer development. PRMT2 expression is increased in several cancer types although its role in acute myeloid leukemia (AML) remains unknown. Here, we investigate the role of PRMT2 in a cohort of patients with AML, PRMT2 knockout AML cell lines as well as a Prmt2 knockout mouse model. In patients, low PRMT2 expressors are enriched for inflammatory signatures, including the NF-κB pathway, and show inferior survival. In keeping with a role for PRMT2 in control of inflammatory signaling, bone marrow-derived macrophages from Prmt2 KO mice display increased pro-inflammatory cytokine signaling upon LPS treatment. In PRMT2-depleted AML cell lines, aberrant inflammatory signaling has been linked to overproduction of IL6, resulting from a deregulation of the NF-κB signaling pathway, therefore leading to hyperactivation of STAT3. Together, these findings identify PRMT2 as a key regulator of inflammation in AML.

overall_design: To investigate the functions of PRMT2 in AML, we used the CRISPR-Cas9 editing strategy on the HL-60 cells to generate stable PRMT2 knockout AML cell lines. We performed single-cell sorting on edited cells. Then, PRMT2 gene and protein expressions were screened on all the clones. The best KO clones (named 'AA13' and AA14') were selected and prepared for bulk RNA-sequencing, as well as the wild type (WT) HL-60 cell line as a control. All 3 samples were run on triplicates.

first: Sauter

last: Camille

first: Aucagne

last: Romain

first: Delva

last: Laurent

first: Fournier

last: Cyril


title: ChIP-seq of BRD4 of HCT116 cells treated with MZ1 or PDD00017273

city: Maebashi

first: Naoko

last: Hattori

state: Gunma

address: 3-39-15 Showa-mach

department: Institute for Molecular and Cellular Regulation

institute: Gunma University

summary: Targeted protein degradation is a groundbreaking modality in drug discovery ; however, the regulatory mechanisms are still not fully understood. Here, we identify cellular signaling pathways that modulate the targeted degradation of the anticancer target BRD4 and related hard-to-degrade targets BRD2/3 induced by CRL2VHL- or CRL4CRBN -based PROTACs. The chemicals identified as degradation enhancers include inhibitors of cellular signaling pathways such as poly-ADP ribosylation (PARG inhibitor PDD00017273), unfolded protein response (PERK inhibitor GSK2606414), and protein stabilization (HSP90 inhibitor luminespib). Mechanistically, PARG inhibition promotes TRIP12-mediated K29/K48-linked branched ubiquitylation of BRD4 by facilitating chromatin dissociation of BRD4 and formation of the BRD4–PROTAC–CRL2VHL ternary complex; by contrast, HSP90 inhibition promotes BRD4 degradation after the ubiquitylation step. Consequently, these signal inhibitors sensitize cells to the PROTAC-induced apoptosis. These results suggest that various cell-intrinsic signaling pathways spontaneously counteract chemically induced target degradation at multiple steps, which could be liberated by specific inhibitors.

overall_design: BRD4 binding profiles in HCT116 cells treated with MZ1 and PDD00017273 were generated by ChIP-sequencing using illumina NextSeq 2000


title: Functional Optimization of Designer Cardiac Organoids Enabled by Machine Learning Techniques

city: Syracuse

first: Yuanhui

last: Song

address: 318 Bowne Hall

institute: Syracuse University

summary: Stem cell organoids are powerful models for studying organ development, disease modeling, drug screening, and regenerative medicine applications. The convergence of organoid technology, tissue engineering, and artificial intelligence (AI) could potentially enhance our understanding of the design principle for organoid engineering. In this study, we utilized micropatterning techniques to create a designer library of 230 cardiac organoids with 7 geometric designs (Circle 200, Circle 600, Circle 1000, Rectangle 1:1, Rectangle 1:4, Star 1:1, and Star 1:4). We employed manifold learning techniques to analyze single organoid heterogeneity based on 10 physiological parameters. We successfully clustered and refined our cardiac organoids based on their functional similarity using unsupervised machine learning approaches, thus elucidating unique functionalities associated with geometric designs. We also highlighted the critical role of calcium rising time in distinguishing organoids based on geometric patterns and clustering results. This innovative integration of organoid engineering and machine learning enhances our understanding of structure-function relationships in cardiac organoids, paving the way for more controlled and optimized organoid design.

overall_design: To elucidate the transcriptomics difference between these distinct organoid population, we performed bulk RNA sequencing on rectangle 1:4 and circle 600 organoids .

first: Andrew

last: Kowalczewski

first: Shiyang

last: Sun

first: Nhu

last: Mai

first: Plansky

last: Hoang

first: Xiyuan

last: Liu

first: Huaxiao

last: Yang

first: Zhen

last: Ma