title: Upstream regulator of genomic imprinting in rice is a small RNA-associated chromatin remodeler [PCR_product_deep_sequencing]

overall_design: We have performed deep sequencing of PCR products amplified from endosperm (WPxPB1) to check genomic imprinting status of CLSY3, ARF22 and SHH1 genes


title: Upstream regulator of genomic imprinting in rice is a small RNA-associated chromatin remodeler [ChIP_seq]

status: Public on Jul 29 2024

city: Bangalore

first: Shivaprasad

last: Padubidri V.

address: UAS-GKVK Campus

country: India

institute: National Centre for Biological Sciences - TIFR

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Knock down of CLSY3 in rice caused endosperm development defects. We investigated the molecular effects of loss of CLSY3 in rice  endosperm by profiling sRNA, targeted bisulfite PCR,whole genome bisulfite sequencing and mRNA transcriptomes .These analyses revealed the  roles of CLSY3 in Endosperm

overall_design: We have  performed ChIP-seq with C-terminal GFP tagged and C-terminal 2X myc tagged CLSY3 transgenic plants panicle.

first: Padubidri

middle: V

last: Shivaprasad

first: Avik

middle: K

last: Pal


title: Spatial analysis of microRNA regulation at defined tumor hypoxia levels reveals biological traits of aggressive prostate cancer

city: Oslo

first: Heidi

last: Lyng

address: Montebello

department: Department of Radiation Biology

country: Norway

institute: Oslo University Hospital

type: Non-coding RNA profiling by high throughput sequencing

summary: Mechanisms regulating the gene expression program at different hypoxia severity levels in patient tumors are not understood. We aimed to determine microRNA (miRNA) regulation of this program at defined hypoxia levels from moderate to severe in prostate cancer. Biopsies from 95 patients were used, where 83 patients received the hypoxia marker pimonidazole before prostatectomy. Forty hypoxia levels were extracted from pimonidazole-stained histological sections and correlated with miRNA and gene expression profiles determined by RNA-sequencing and Illumina bead arrays. This identified miRNAs associated with moderate (n=7) and severe (n=28) hypoxia and predicted their target genes. Scores of miRNAs or target genes showed prognostic significance, as validated in external cohort of 417 patients. The target genes showed enrichment of gene sets for cell proliferation and MYC activation at all hypoxia levels and PTEN inactivation at severe hypoxia. This was confirmed by RT-qPCR for MYC and PTEN, by Ki67-immunohistochemistry, and by gene set analysis in an external cohort. To assess whether miRNA regulation occurred within the predicted hypoxic regions, a method to quantify co-localization of multiple histopathology parameters at defined hypoxia levels was applied. A high Ki67-proliferation index co-localized significantly with hypoxia at all levels. The co-localization index was strongly associated with poor prognosis. Absence of PTEN-staining co-localized significantly with severe hypoxia. Scores for miRNAs correlated with the co-localization index for Ki67-staining and hypoxia, consistent with miRNA regulation within the overlapping regions. This was confirmed by showing miR-210-3p expression within severe hypoxia by in situ hybridization. Cell line experiments (22Rv1, PC3) were conducted to determine whether miRNAs and target genes were regulated directly by hypoxia. Most of them were hypoxia-unresponsive, and probably regulated by other mechanisms such as MYC activation. In conclusion, cancer cells residing within moderate and severe hypoxic regions in aggressive prostate tumors in patients exhibit different proliferative gene expression programs regulated by miRNAs.

overall_design: miRNA expression  was determined from small RNA-sequencing data of biopsies from the dominant (index) lesion of 91 patients treated with prostatectomy. Hypoxia levels were measured by pimonidazole immunostaining of biopsy sections.

first: Vilde

middle: E

last: Skingen

first: Unn

middle: B

last: Salberg

first: Tord

last: Hompland

first: Christina

middle: S

last: Fjeldbo

first: Hanna

last: Helgeland

first: Kari-Anne

middle: M

last: Frikstad

first: Harald

last: Ragnum

first: Ljiljana

last: Vlatkovic

first: Knut

middle: H

last: Hole

first: Therese

last: Seierstad


title: Constitutive BoHAK5 expression and early robust induction of salicylic acid biosynthesis genes as candidates to explain the tolerance of broccoli to limiting potassium

city: Valencia

first: Eduardo

last: Bueso

state: Valencia

address: Ingeniero Fausto Elio, s/n

department: IBMCP

country: Spain

institute: Universitat Politècnica de València

type: Expression profiling by high throughput sequencing

summary: Potassium is vital for optimum plant growth and crop yield and so is an important component of fertilizers. However, our knowledge of the physiological and molecular response in response to limiting potassium conditions is incomplete. Here we performed a detailed transcriptomic analysis of the model plant, Arabidopsis and the crop plant, broccoli to compare and contrast the response to this abiotic stress. Our results show that broccoli plants lose potassium at a much slower rate than Arabidopsis. This may be explained by the high levels of expression of the BoHAK5 gene. Both Arabidopsis and broccoli showed characteristic responses observed in other plant species, related to oxidative stress and hypoxia and jasmonic, salicylic and abscisic acid signaling. In broccoli, we also observed alterations in the gene expression patterns of enzymes and in the levels of intermediates involved in the biosynthesis of glucosinolates, which are important molecules contributing to the added nutritional value of broccoli. Lastly, we provide evidence for concomitant alterations in the transport of several other ions, such as iron, phosphate and nitrate. Our data help to characterize the response of broccoli to limiting potassium conditions and may provide insights for the development of varieties of broccoli with reduced fertilization requirements.

overall_design: Transcriptomic response in roots and leaf in broccoli after 1, 3 and 6 days

first: Yenush

last: Lynne

first: Bueso

last: Eduardo

first: Mulet

last: José Miguel

first: Núria

last: Andrés-Colás


overall_design: Gene expression profiling of PC-3 and 22Rv1 cells exposed to hypoxia at 0.2%, 0.5%, 1%, 2% and 5% O2 and normoxia (95% air, control) was carried out, two replicates of hypoxia and control for each O2 concentration.


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

city: London

first: Probir

last: Chakravarty

address: 1 Midland Road

country: United Kingdom

institute: Francis Crick Institute

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

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

first: Erik

last: Sahai

first: Eishu

last: Hirata


title: Spatial analysis of microRNA regulation at defined tumor hypoxia levels reveals biological traits of aggressive prostate cancer [cell lines]

first: Heidi

last: Lyng

overall_design: miRNA expression profiling of PC-3 and 22Rv1 cells exposed to hypoxia at 0.2%, 0.5%, 1%, 2% and 5% O2 and normoxia (95% air, control) was carried out, two replicates of hypoxia and control for each O2 concentration.

first: Vilde

last: Skingen

first: Unn

last: Salberg


title: RING1 missense variants reveal sensitivity of DNA damage repair to H2A monoubiquitination dosage during neurogenesis [scRNA-seq]

city: Ann Arbor

first: Charles

middle: William

last: Ryan

state: Michigan

address: 1241 Catherine St, 4909 Buhl Building

department: Human Genetics

country: USA

institute: University of Michigan

summary: Polycomb repressive complex 1 (PRC1) modifies chromatin through catalysis of histone 2A lysine 119 monoubiquitination (H2AK119ub1). RING1 and RNF2 interchangeably serve as the catalytic subunit within PRC1. Pathogenic missense variants in PRC1 core components reveal functions of these proteins that are obscured in knockout models. While Ring1a knockout models remain healthy, the microcephaly and neuropsychiatric phenotypes associated with a pathogenic RING1 missense variant implicate unappreciated functions. Using an in-vitro model of neurodevelopment, we observe that RING1 contributes to the broad placement of H2AK119ub1, and that its targets overlap with those of RNF2. PRC1 complexes harboring hypomorphic RING1 bind target loci but do not catalyze H2AK119ub1, reducing H2AK119ub1 by preventing catalytically active complexes from accessing the locus. This results in delayed DNA damage repair and cell cycle progression in neural progenitor cells (NPCs). Conversely, reduced H2AK119ub1 due to hypomorphic RING1 does not generate differential expression that impacts NPC differentiation. The greater reduction of H2AK119ub1 dosage observed in NPCs expressing hypomorphic RNF2, however, exhibited defects in both DNA repair and transcriptional regulation. These findings suggest that the DNA damage response is more sensitive to H2AK119ub1 dosage change than is regulation of gene expression.

overall_design: To determine the effect of H2AK119ub1 dosage on gene expression, we subjected cultured neural progenitor cells to bulk RNA sequencing for our RING1/RNF2 allelic series.

first: Samantha

last: Regan

first: Yao

last: Tsan

first: Stephanie

last: Bielas


title: RING1 missense variants reveal sensitivity of DNA damage repair to H2A monoubiquitination dosage during neurogenesis.


title: RING1 missense variants reveal sensitivity of DNA damage repair to H2A monoubiquitination dosage during neurogenesis [bulk RNA-seq]


title: The Characteristics of cell in Psoriatic Resolved Lesions Revealed and Onsite Recurrent by Single-cell Transcriptome

city: Guangzhou

first: Lu

last: peng

address: No.106, Zhongshan 2nd Road

department: Dermatology

country: China

institute: Guangdong Provincial People's Hospital

summary: Skin biopsy from the previously resolved region (PN), the autologous on-site recurrence with psoriatic plaque (PP)and the non-lesion control (CN) were obtained from the lower back of four male plaque psoriasis patients. Single-cell RNA sequencing (scRNA-seq) was performed after cell dissociation. In our article, sub-clustering and characterization of T cells were demonstrated, functional comparisons of T-cell subsets were conducted further among PN, PP and CN.

overall_design: The previously resolved region (PN), the autologous on-site recurrence with psoriatic plaque (PP)and the appearing-normal control (CN) were obtained from the lower back of four male plaque psoriasis patients. Single-cell RNA sequencing (scRNA-seq) were performed after cell dissociation. The specimen named “CN” has combinated the cells of appearing-normal control from Patient D1 and Patient D2.

last: Peng

first: Wenqi

last: Liu

first: Yufan

last: Cheng

first: Ling

last: Chen

first: Zhu

last: Shen


title: Local and systemic transcriptome and spliceome changes induced by Meloidigyne incognita in tomato roots.

city: Knoxville

first: Tarek

last: Hewezi

state: TN

address: 2431 Joe Johnson Drive

department: Plant Sciences

institute: University of Tennessee

summary: The aim of study is to investigate DEGs, long non-coding RNAs and alternative splicing events in the development of galls and neighboring region compared to the non-infected whole root induced by root-knot nematode (RKN, Meloidogyne incognita). Total RNA was extracted, ribosomal depleted libraries were prepared, and then high throughput RNA sequencing was performed using the Illumina NovaSeq 6000. High quality, paired-end reads were then aligned to the tomato reference genome (Heinz1706 assembly SL4.0) and uniquely mapped reads were counted using Htseq. Finally, differentially expressed genes (DEGs) between whole roots-galls and whole roots-neighboring region were identified using DESeq package and downstream analyses were performed.

overall_design: Susceptible Heinz1706 tomato plants were grown and 10-days-old plants were inoculated with 1000 second-stage juvenile(J2) M.incognita. 4 and 11 days post inoculation, galls, neighboring region which is two near side of galls and non-infected whole roots were collected as three biological replicates.

first: Selin

last: Ozdemir


title: Serine arginine-rich splicing factor (SRSF7) cooperates with the histone methyltransferase KMT5a to promote the type I interferon response via transcriptional activation of IRF7

city: Bryan

first: Kristin

last: Patrick

state: Texas

address: 8441 Riverside Parkway

department: MPIM

institute: Texas A&M

summary: Tight regulation of macrophage immune gene expression is required to fight infection without risking harmful inflammation. The contribution of RNA binding proteins (RBPs) to shaping the macrophage response to pathogens remains poorly understood. Transcriptomic analysis revealed that a member of the serine/arginine-rich (SR) family of mRNA processing factors, SRSF7, is required for optimal expression of a cohort of interferon stimulated genes (ISGs) in macrophages. Using genetic and biochemical assays, we discovered that in addition to its canonical role in regulating alternative splicing, SRSF7 drives transcription of interferon regulatory transcription factor 7 (IRF7) to promote antiviral immunity. At the Irf7 promoter, SRSF7 maximizes STAT1 transcription factor binding and RNA polymerase II elongation via cooperation with the H4K20me1 histone methyltransferase KMT5a (SET8). These studies define an unorthodox role for an SR protein in activating transcription and reveal an unappreciated RNA binding protein-chromatin network that orchestrates macrophage antiviral gene expression.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) of 3xFLAG-GFP or 3xFLAG-SRSF7 in RAW 264.7 cells 4h post stimulation with LPS (100 ng/ml).

first: Haley

last: Scott

first: Mackenzie

last: Smith

first: Aja

last: Coleman

first: Summer

last: Apostalo

first: Allison

last: Wagner

first: Robert

last: Watson


title: Upstream regulator of genomic imprinting in rice is a small RNA-associated chromatin remodeler [targeted bisulfite PCR]

first: Shivaprasad

last: Padubidri V.

type: Methylation profiling by high throughput sequencing

summary: Knock down of CLSY3 in rice caused endosperm development defects. We investigated the molecular effects of loss of CLSY3 in rice  endosperm by profiling sRNA, targeted bisulfite PCR and mRNA transcriptomes .These analyses revealed the  roles of CLSY3 in Endosperm

overall_design: We have performed targeted bisulfite PCR

first: Padubidri

last: Shivaprasad

first: Avik

last: Pal


title: Cadherin adhesion complexes direct cell aggregation in the epithelial transition of Wnt-induced nephron progenitor cells

city: Los Angeles

first: Helena

last: Bugacov

state: CA

address: 1425 San Pablo St.

department: Development, Stem Cell and Regenerative Medicine

institute: USC STEM CELL

summary: The aggregation of nephron progenitor cells (NPC) is required to form the nephron precursor renal vesicle (RV) as they undergo the mesenchymal-to-epithelial transition (MET). Canonical Wnt signaling regulates the MET of NPCs via the effector molecule β-catenin. β-catenin acts as a transcriptional co-activator during the differentiation of NPCs, however, if it has any role as an adhesion regulating molecule via the cadherin-catenin complex is currently unknown. Using in vitro NPC culture system with gene editing and modulated Wnt signaling input, we investigated the morphological transition of NPCs and its underlying mechanism modeling the initial step of the MET in vivo. Increasing Wnt signaling input with the small molecule agonist CHIR resulted in the aggregation of NPCs similar to the generation of the nephron anlagen. By removing β- and α-catenin from the NPCs, NPCs are sorted out from these aggregates, demonstrating the key role of the cadherin-catenin complex in the aggregation. β-catenin was essential for the induction, however, the removal of α-catenin did not affect the transcriptional profile of NPCs. Removal of extracellular Ca2+ resulted in the transient loss of cell-cell contacts suggesting the role of cadherins in the aggregation process. The analysis of in vitro bulk RNA-seq cadherin expression profile matched the in vivo cadherin expression profile determined by single-cell RNA-seq. The combined removal of pre-existing and inducible cadherins phenocopied the results of β- and α-catenin KO experiments highlighting the crucial role of the cadherin-catenin complex by multiple lines of evidence. Notably, the induction of NPCs is independent of their aggregation. The in vitro modeling of nephron development provides a mechanistic understanding how cell adhesion is regulated via the cadherin-catenin complex during nephrogenesis.

overall_design: Primary nephron progenitors (NPCs)were isolated using the methods published in Brown et al., 2015 and were cultured in NPEM media. NPCs were then transfected using Lipofectamine™ MessengerMAX™ Transfection Reagent (Thermofisher, Cat # LMRNA015) with gene editing components (sgRNAs) and mCherry mRNA to distinguish transfected cells. Cells were cultured and lifted for FAC sorting. RNA was extracted from FAC sorted samples using mCherry+ NPCs were sorted on  BD SORP FACS Aria IIu into RLT buffer with 1:100 beta-mercaptoethanol prior to RNA isolation. RNA was isolated using Qiagen RNeasy Micro Kit (74004).  Total RNA integrity was determined using Agilent Bioanalyzer or 4200 Tapestation. Library preparation was performed with 10ng of total RNA with a Bioanalyzer RIN score greater than 8.0. ds-cDNA was prepared using the SMARTer Ultra Low RNA kit for Illumina Sequencing (Takara-Clontech) per manufacturer's protocol. cDNA was fragmented using a Covaris E220 sonicator using peak incident power 18, duty factor 20%, cycles per burst 50 for 120 seconds. cDNA was blunt ended, had an A base added to the 3' ends, and then had Illumina sequencing adapters ligated to the ends. Ligated fragments were then amplified for 12-15 cycles using primers incorporating unique dual index tags. Fragments were sequenced on an Illumina NovaSeq-6000 using paired end reads extending 150 bases.

first: Balint

last: Der

first: Andrew

last: McMahon


title: Upstream regulator of genomic imprinting in rice is a small RNA-associated chromatin remodeler [RNA-seq]

summary: Knock down of CLSY3 in rice caused endosperm development defects. We investigated the molecular effects of loss of CLSY3 in rice  endosperm by profiling sRNA and mRNA transcriptomes .These analyses revealed the  roles of CLSY3 in Endosperm development.

overall_design: We performed RNA seq analyses to look at the expression of the genes in WT and clsy3 kd at 20 days after anthesis. We performed RNAseq analyses to look at the expression of the genes in embryo and endosperm 25days days after anthesis.


title: Upstream regulator of genomic imprinting in rice is a small RNA-associated chromatin remodeler [ncRNA-Seq]

overall_design: We profiled sRNAs in the WT and clsy3 kd tissues from embryo and endosperm tissues to estimate the effect of loss of clsy3 the the sRNA transcriptome.


title: Epigenetic reprogramming orchestrates transcriptional responses in shade avoidance memory of Arabidopsis

city: Shanghai

first: Qican

last: Cheng

state: Shanghai

address: 2005 Songhu Road, Yangpu District

department: school of life sciences

institute: Fudan university

summary: An RNA-seq was conducted to investigate how REF6 and PIF7 participate in shade avoidance memory.

overall_design: Total RNA was extracted from seedlings grown under N (4-day-old seedlings + 8 h white light + 40 h recovering white light + 1 h triggering shade) and P (4-day-old seedlings + 8 h priming shade + 40 h recovering white light + 1 h triggering shade) conditions.


title: Effect of Vortioxetine treatment on gene expression of LN-229 glioblastoma cells over time

city: Zurich

first: Berend

last: Snijder

address: Otto-Stern-Weg 3, HPM building

country: Switzerland

institute: ETH Zurich

summary:
The atypical antidepressant Vortioxetine represents the most effective neuroactive drug with anti-glioblastoma efficacy across glioblastoma patient samples and model systems. To closer examine the temporal dynamics of Vortioxetine, we performed in-depth transcriptomic profiling of LN-229 cells following Vortioxetine treatment.
We performed gene expression profiling analysis using data obtained from RNA-seq across 6 different time points during the 24 hours following Vortioxetine treatment.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for LN-229 glioblastoma cells at baseline then at each respective time point after Vortioxetine treatment

first: Sohyon

last: Lee


title: Dose-dependent responses to canonical Wnt transcriptional complexes in the regulation of mammalian nephron progenitors

summary: Wnt/β-catenin signaling is a highly conserved molecular pathway that plays a crucial role in stem/progenitor systems and cancer. β-catenin, the main Wnt pathway effector, has two pools within a cell: one for cell-cell adhesion at the membrane and the other for transcriptional functions in the nucleus. However, the mechanism by which β-catenin mediates both roles remain unclear. The tightly controlled, well characterized system of nephrogenesis is an ideal model to decouple the roles of β-catenin at the membrane and in the nucleus. In kidney development, a delicate balance of nephron progenitor cell  self-renewal and differentiation is required for the mesenchymal to epithelial transition (MET) in nephrogenesis and is driven by Wnt/β-catenin signaling. Given an ability to isolate and manipulate large numbers of NPCs in tissue culture, we can dissect the dual nature of β-catenin as a transcriptional activator and component of a cell membrane complex in adhesion. We pioneered a method using CRISPR/Cas9 gene editing to rapidly remove β-catenin, Tcf/Lef factors and simultaneous cadherin genes in primary NPCs. We have characterized the effects of modulating β-catenin and integrated RNA-seq results from β-catenin’s removal with mouse ChIP-seq and mouse single cell RNA -seq data. Functional analysis of β-catenin removal provides strong evidence for β-catenin regulation of NPC proliferation, independent of a direct Lef/Tcf associated transcriptional program. Together these data suggest β-catenin mediates aggregation, the first step in MET, through β-catenin mediated cell adhesion complexes while simultaneous transcriptional activation within these structures initiates the nephrogenic program. The studies provide new insight into the direct transcriptional role of Lef/Tcf/β-catenin complexes associated with the initiation of a nephron forming program. Overall, this study enhances an understanding of the molecular mechanisms underlying kidney development and the dual nature of β-catenin stem/progenitor systems at large.

overall_design: Primary nephron progenitors (NPCs)were isolated using the methods published in Brown et al., 2015 and were cultured in NPEM media. NPCs were then transfected using Lipofectamine™ MessengerMAX™ Transfection Reagent (Thermofisher, Cat # LMRNA015) with gene editing components (sgRNAs or Cre mRNA) and mCherry mRNA to distinguish transfected cells. Cells were cultured and lifted for FAC sorting. RNA was extracted from FAC sorted samples using mCherry+ NPCs were sorted on  BD SORP FACS Aria IIu into RLT buffer with 1:100 beta-mercaptoethanol prior to RNA isolation. RNA was isolated using Qiagen RNeasy Micro Kit (74004).  Total RNA integrity was determined using Agilent Bioanalyzer or 4200 Tapestation. Library preparation was performed with 10ng of total RNA with a Bioanalyzer RIN score greater than 8.0. ds-cDNA was prepared using the SMARTer Ultra Low RNA kit for Illumina Sequencing (Takara-Clontech) per manufacturer's protocol. cDNA was fragmented using a Covaris E220 sonicator using peak incident power 18, duty factor 20%, cycles per burst 50 for 120 seconds. cDNA was blunt ended, had an A base added to the 3' ends, and then had Illumina sequencing adapters ligated to the ends. Ligated fragments were then amplified for 12-15 cycles using primers incorporating unique dual index tags. Fragments were sequenced on an Illumina NovaSeq-6000 using paired end reads extending 150 bases.  Basecalls and demultiplexing were performed with Illumina’s bcl2fastq2 software.  RNA-seq reads were then aligned to the combined mouse GRCm38 and human GRCh38 Ensembl release 76 primary assemblies with STAR version 2.5.1a1.  Gene counts were derived from the number of uniquely aligned unambiguous reads by Subread:featureCount version 1.4.6-p52. Isoform expression of known Ensembl transcripts were estimated with Salmon version 0.8.23.  Sequencing performance was assessed for the total number of aligned reads, total number of uniquely aligned reads, and features detected.  The ribosomal fraction, known junction saturation, and read distribution over known gene models were quantified with RSeQC version 2.6.24.  Normalized counts tables were ran through DeSEQ2 (Love, 2014 #412) to create differential gene expression tables with Log2Fc cut offs no less than 1 and p-adjusted values no greater than 0.05. Significance of Cre and Cas9 intersected gene lists was calculated with hypergeometric function in R. Differential expression tables were passed through Gene Ontology package clusterProfiler (Yu, 2012). Data was visualized using ggplot and Complex heatmap functions in R (Gu, 2016). Benjamini-Hochberg correction (False Discovery Rate) applied as well as significance associated with intersections compared to all genes expressed with gene normalized counts greater than or equal to 10 using hypergeometric test.

first: Sunghyun

last: Kim

first: Nils

last: Lindstrom


title: scRNA-Seq analysis of a glioblastoma patient sample after Vortioxetine or DMSO treatment

summary: Vortioxetine is the neuroactive drug with the highest ex vivo anti-glioblastoma efficacy across patients measured in our study. We used single-cell RNA sequencing (scRNA-seq) to analyze the transcriptomic response to Vortioxetine treatment.

overall_design: Dissociated and cryopreserved single-cell suspensions of a patient sample P024 were processed by fluorescence-activated cell sorting (FACS) to isolate viable CD45+ immune cells and CD45- glioblastoma cells. Sorted cells were subsequently treated with Vortioxetine or DMSO for 3 hours before proceeding to generate scRNA-Seq libraries.

first: Tobias

last: Weiss


title: Digital RNA with pertUrbation of Genes (DRUG-Seq) of LN-229 glioblastoma cells treated with 20 different drugs at two time points

summary:
DRUG-Seq is a miniatruized and multiplexed RNA-Seq protocol described in Ye et al., 2018 (https://www.nature.com/articles/s41467-018-06500-x)
To determine transcriptional changes upon drug perturbation in LN-229 glioblastoma cells, we profiled 20 drug responses across two time-points (6 and 22 hours) by DRUG-Seq

overall_design: These 20 drugs were selected to include pharmacoscopy-hit neuroactive drugs spanning diverse drug classes (n=11), pharmacoscopy-hit oncology drugs (n=7), pharmacoscopy-negative neuroactive drugs (n=2), and a DMSO control.


title: Deletion of a state-specific PD-1 enhancer modulates exhausted T cell fate and function [ATAC-seq]

city: Boston

first: Amy

last: Huang

state: MA

address: 25 Shattuck St

department: Immunology

institute: Harvard Medical School

summary: T cell exhaustion is a state of CD8+ T cell dysfunction elicited by chronic exposure to antigen and inflammation, arises in both cancer and chronic viral infection. The co-inhibitory receptor PD-1 plays a key role in mediating exhaustion, but complete ablation of PD-1 by gene knock-out leads to deeper functional deficits and poor T cell survival. We hypothesized that an intermediate level of PD-1 expression may confer an improved balance of exhausted CD8+ T cell functionality, so we deleted an exhaustion-associated enhancer of PD-1 which indeed resulted in a reduced expression level. We compared EnhDel, WT and PD-1 KO T cells using single-cell RNA-Seq and found that PD-1 KO but not EnhDel cells are strongly biased towards the terminally exhausted subset.  EnhDel cells also uniquely enrich for effector-associated genes and gene signatures. However, all three genotypes (EnhDel, WT and PD-1 KO) exhibit a similar chromatin accessibility landscape by ATAC-Seq, controlling for exhausted subset. These data suggest that tuning of PD-1 expression may uniquely permit the maintenance of an “effector” transcriptional profile in exhausted CD8+ T cells.

overall_design: Enhancer-deleted and PD-1 KO P14+ cells, or enhancer-deleted and wildtype P14+ cells, were co-transferred into recipient animals (n = 15-30) at a 75:25 or 50:50 ratio, respectively. Recipients were infected with LCMV Clone 13. At day 30, spleens were pooled into two biological replicates and transferred CD8+ T cells were isolated with magnetic enrichment. Progenitor and terminally-exhausted subsets for each genotype were isolated with flow-assisted sorting, and samples were processed for ATAC-Seq.

first: Sarah

middle: A

middle: Y

first: Megan

last: Fung

first: Chang-Yu

last: Chen

first: Brian

middle: C

last: Miller

first: Jared

last: Rowe

first: Natalia

last: Sifnugel

first: Mei-An

last: Nolan

first: Dillon

middle: G

last: Patterson

first: Jossef

middle: F

last: Osborn

first: Sydney

last: Loh

first: Marc

last: Schwartz

first: Richard

last: Davidson

first: Martin

middle: W

last: LaFleur

first: John

last: Doench

first: W

middle: N

last: Haining

first: Arlene

last: Sharpe

first: Debattama

middle: R

last: Sen


title: Extracellular vesicles miR-574-5p and miR-181a-5p as prognostic markers in NSCLC patients treated with nivolumab

status: Public on Jul 28 2024

city: Genova

first: simona

last: coco

address: L.go R. Benzi, 10

country: Italy

institute: IRCCS Ospedale Policlinico San Martino

summary:
Plasma from 245 patients with advanced NSCLC who received nivolumab as second-line therapy was collected and analyzed. EV-miRnome was profiled on 174/245 patients by microarray platform and selected EV-miRs were validated by qPCR. A prognostic model combining EV-miR and clinical variables was built using stepwise Cox regression analysis and tested on an independent patient cohort (71/245). EV-PD-L1 gene copy number was assessed by digital PCR. For 54 patients with disease control, EV-miR changes at best response versus baseline were investigated by microarray and validated by qPCR.
EV-miRNome profiling at baseline identified two EV-miR (miR-181a-5p, miR-574-5p) that, combined with performance status, are capable of discriminating patients unlikely from those that are likely to benefit from immunotherapy (median overall survival of 4 months or higher than 9 months, respectively).
EV-PD-L1 digital evaluation reported higher baseline copy number in patients at increased mortality risk, without improving the prognostic score. Best response EV-miRNome profiling selected six deregulated EV-miRs (miR19a-3p, miR-20a-5p, miR-142-3p, miR-1260a, miR-1260b, miR-5100) in responding patients. Their longitudinal monitoring highlighted a significant downmodulation already in the first treatment cycles, which lasted more than six months.

Grantee:  Simona Coco
Grantor:  Italian Ministry of Health
Grant ID:  CO-2016-02361470
Grant Title:  Exosomal miRNA signature as prognostic marker in advanced non-small cell lung cancer (NSCLC) patients treated with Nivolumab

overall_design: 174 plasma Exososome miRNAs were profiled comparing short survivors (patients with an overall survival lower than 9 months) versus long survivors ( patients with an OS higher than 9 months). 108 plasma exosome miRNAs derived from 54 NSCLC patients reporting disease control (i.e., 31 stable disease and 23 Partial Response; based on RECIST criteria) after treatment with nivolumab. For each patient, plasma samples were collected at baseline and at the best response time. The entire exosome miRNome was profiled comparing the changes from baseline to the best response.

first: Simona

last: Coco

first: Silvia

last: Marconi

first: Giovanna

last: Chiorino

first: Francesca

last: Guana

first: Luca

last: Longo

first: Irene

last: Vanni

first: Paola

last: Ostano

first: Carlo

last: Genova


title: Deletion of a state-specific PD-1 enhancer modulates exhausted T cell fate and function [scRNA-seq]

first: Amy

last: Huang

overall_design: Enhancer-deleted, PD-1 KO and WT P14+ CD8+ T cells were isolated from donor mice, mixed at a 33:33:33 ratio, and transferred to GFP+ recipient mice (n = 5). Recipients were infected with LCMV Clone 13. Transferred CD8+ T cells were isolated from spleens at day 29 of infection, then pooled into two biological replicates. Genotypes (Enhancer-deleted, PD-1 KO, WT) were then isolated by sorting and processed for scRNA-Seq.

first: Sarah

last: Weiss

first: Megan

last: Fung

first: Chang-Yu

last: Chen

first: Brian

last: Miller

first: Jared

last: Rowe

first: Natalia

last: Sifnugel

first: Mei-An

last: Nolan

first: Dillon

last: Patterson

first: Jossef

last: Osborn

first: Sydney

last: Loh


title: Deletion of a state-specific PD-1 enhancer modulates exhausted T cell fate and function

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

overall_design: Refer to individual Series


title: Effect of dexmedetomidine on ncRNA and mRNA profiles of cerebral ischemia-reperfusion injury in transient middle cerebral artery occlusion rats model

status: Public on Jul 27 2024

city: Zheng Zhou

first: Nasir

last: ABDUL

address: Jingba Road

department: The Second Clinical Medical School of Zhengzhou University

institute: Zhengzhou University

summary: Ischemic stroke poses a significant global health burden, with rapid revascularization treatments being crucial but often insufficient to mitigate ischemia-reperfusion (I/R) injury. Dexmedetomidine (DEX) has shown promise in reducing cerebral I/R injury, but its potential molecular mechanism, particularly its interaction with non-coding RNAs (ncRNAs), remains unclear. This study investigates DEX's therapeutic effect and potential molecular mechanisms in reducing cerebral I/R injury. A transient middle cerebral artery obstruction (tMACO) model was established to simulate cerebral I/R injury in adult rats. DEX was administered pre-ischemia and post-reperfusion. RNA sequencing and bioinformatic analyses were performed on the ischemic cerebral cortex to identify differentially expressed non-coding RNAs (ncRNAs) and mRNAs. The sequencing results showed 6494 differentially expressed (DE) mRNA and 2698 DE circRNA between the sham (S) and tMCAO (I/R) groups. Additionally, 1809 DE lncRNA, 763 DE mRNA, and 2795 DE circRNA were identified between the I/R group and tMCAO+DEX (I/R+DEX) groups. Gene ontology (GO) analysis indicated significant enrichment in multicellular biogenesis, plasma membrane components, and protein binding. KEGG analysis further highlighted the potential mechanism of DEX action in reducing cerebral I/R injury, with hub genes involved in inflammatory pathways. This study demonstrates DEX's efficacy in reducing cerebral I/R injury and offers insights into its brain-protective effects, especially in ischemic stroke. Further research is warranted to fully understand DEX's neuroprotective mechanisms and its clinical applications.

overall_design: The gene expression difference between the model group and the control group was compared, and the gene expression difference between the drug administration group and the model group was compared


title: Bulk RNA sequencing in SARS-CoV-2 infected patients

city: CHEONAN

first: Hyunkyung

last: Cha

state: Cheonan-si

address: Suncheonhyang 6-gil

department: Department of Otorhinolaryngology-Head and Neck Surgery

country: South Korea

institute: Soonchunhyang University Cheonan Hospital

summary: We report the application of bulk RNA sequencing technology for high-throughput profiling of SARS-CoV-2 infection in the human nasopharynx

overall_design: Examination of SARS-CoV-2-induced RNA expression in the nasopharynx of human

first: Hyun

middle: J

last: Kim

first: Sujin

first: Chan

last: Lee


title: Light-gated integrator for highlighting kinase activity in living cells

city: La Jolla

first: Wei

last: Lin

address: 9500 Gilman Dr

department: Pharmacology

institute: UC San Diego

summary: Protein kinases are key signaling nodes that regulate fundamental biological and disease processes. Illuminating kinase signaling from multiple angles can provide deeper insights into disease mechanisms and improve therapeutic targeting. While fluorescent biosensors are powerful tools for visualizing live-cell kinase activity dynamics in real time, new molecular tools are needed that enable recording of transient signaling activities for post hoc analysis and targeted manipulation. Here, we develop a light-gated kinase activity coupled transcriptional integrator (KINACT) that converts dynamic kinase signals into “permanent” fluorescent marks. KINACT enables robust monitoring of kinase activity across scales, accurately recording subcellular PKA activity, highlighting PKA activity distribution in 3D cultures, and identifying PKA activators and inhibitors in high-throughput screens. We further leverage the ability of KINACT to drive signaling effector expression to allow feedback manipulation of the balance of GαsR201C-induced PKA and ERK activation and dissect the mechanisms of oncogenic G protein signaling.

overall_design: We generated two dual-stable cell lines: one A-KINACT effector cell line stably expressing A-KINACT and integrating TetO-driven EGFP-fused PKA inhibitory peptide (TetO-PKI-EGFP), and one A-KINACT control cell line stably expressing A-KINACT and integrating TetO-EGFP. A-KINACT-expressing HEK293T cells showing mTagBFP2-GαsR201C overexpression and A-KINACT-driven PKI-EGFP expression were identified and collected via FACS to assess differences in transcription versus control cells expressing A-KINACT-driven EGFP.

first: Abhishek

last: Phatarphekar

first: Yanghao

last: Zhong

first: Longwei

last: Liu

first: Hyung-Bae

last: Kwon

first: William

last: Gerwick

first: Yingxiao

last: Wang

first: Sohum

last: Mehta

first: Jin

last: Zhang


title: Human breastmilk memory T cells throughout lactation manifest an activated and tissue-oriented profile with prominent regulation

status: Public on Jul 26 2024

city: Utrecht

first: Elise

middle: S.

last: Saager

address: Lundlaan 6

department: Centre for Translational Immunology

country: Netherlands

institute: University Medical Centre Utrecht

summary: Breastfeeding provides important immunological benefits to the neonate, but how different immunoactive components in breastmilk contribute to immunity remains poorly understood. Here, we characterized human breastmilk T cells using single cell RNA sequencing and flow cytometry. Breastmilk contained predominantly memory T cells with immune response signaling, proliferation and an effector Th1/cytotoxic profile with high cytokine production capacities. High activation markers were balanced by an enriched Treg population and immune regulatory markers in conventional memory T cells. Gene and surface expression of tissue-residency markers indicated that breastmilk T cells represent tissue-adapted rather than circulatory T cells. In addition, breastmilk T cells had a broad homing profile and higher activation markers in these migratory subsets. The partly overlapping transcriptome profile between breastmilk and breast tissue T cells, particularly cytotoxic T cells, might support a role in local immune defense in the mammary gland. However, unique features of breastmilk, such as regulatory T cells, might imply an additional role in neonatal immune defense. We found some correlations between the breastmilk T-cell profile and clinical parameters, most notably with maternal and household factors. . Together, our data suggest that breastmilk contains an adapted T cell population that exerts their function in specific tissue-sites.

overall_design: scRNAseq data of plate-sorted T cells (sort-seq) from 1 month postpartum breastmilk (n=7) were clustered together to reveal T-cell populations and characteristic DEG present in human breastmilk.


title: The epithelial transcriptome of hidradenitis suppurativa tunnels is more similar to cutaneous squamous cell carcinoma than to benign infundibular cysts

city: Norfolk

first: Clinton

last: Enos

state: Virginia

address: 721 Fairfax Avenue, Suite 200

institute: Eastern Virginia Medical School

summary: The goal is to utilize the Nanostring Geomx platform to examine the spatial moecular signature of hidradenitis suppurativa tunnel epithelium compared to the epithelium of a malignant neoplasm, invasive squamous cell carcinoma, and a benign epidermal cyst.

overall_design: A total of 4 HS, 3 SCC, 3 cyst, and 3 healthy control (HC) samples were collected from surgical archival tissue. Utilizing Nanostring GeoMX Digital Spatial profiler, regions of interst were selected to include epithelium and adjacent stroma in HS tunnels, cyst walls, and SCC epithelium.

first: Waleed

last: Adawi

first: Chunghwan

last: Ro

first: David

last: Scoville

first: Yan

last: Luang

first: Stefan

last: Phelan

first: Alice

last: Roberts


title: Preventive Treatment with a CD73 Small Molecule Inhibitor Enhances Immune Surveillance in K-Ras Mutant Pancreatic Intraepithelial Neoplasia

city: Houston

first: Zhongming

last: Zhao

state: TX

address: 7000 Fannin St Suite 600

department: McWilliams School of Biomedical Informatics

institute: The University of Texas Health Science Center at Houston

summary: Immunoprevention is an emerging consideration for solid tumors, including pancreatic ductal adenocarcinoma (PDAC). We and others have shown that Kras mutations in genetic models of spontaneous pancreatic intraepithelial neoplasia (PanIN), which is a precursor to PDAC, results in CD73 expression in the neoplastic epithelium and some populations of infiltrating immune cells, including macrophages and CD8 T cells. CD73 is an ecto-enzyme that converts extracellular adenosine monophosphate (AMP) to adenosine, a critical immune inhibitory molecule in PDAC. We hypothesized inhibition of CD73 would reduce the incidence of PanIN formation and alter the immune microenvironment. To test our hypothesis, we used the KrasG12D; PdxCre1 (KC) genetically engineered mouse (GEM) model and tested the utility of AB-680, a small molecule inhibitor targeting CD73, to inhibit PanIN progression. AB-680, or vehicle control, was administered using oral gavage delivery three days/week at 10mg/kg, beginning when the mice were two months old and lasting three months. We euthanized the mice at five months old. In the KC model, we quantified significantly less pancreatitis, early and advanced PanIN, and quantified a significant increase in M1 macrophages in AB-680-treated mice. Single Cell RNA sequencing (scRNA-seq) of pancreata of AB-680 treated mice revealed increased infiltration of CD4+ T cells, CD8+ T cells, and mature B cells. The scRNA-seq analysis showed that CD73 inhibition reduced M2 macrophages, acinar, and PanIN cell populations. CD73 inhibition enhanced immune surveillance and expanded unique clonotypes of TCR and BCR, indicating that inhibition of CD73 augments adaptive immunity early in the neoplastic microenvironment.

overall_design: We used the genetically engineered KrasG12D/PdxCre1(KC) mouse model, which slowly developed pancreatic intraepithelial neoplasia (PanIN) and pancreatic ductal adenocarcinoma (PDAC) over 12-15 months. AB-680 (HY-125286) was dissolved in 20% SBE-β-CD (MedChem Express, HY-17031/CS-0731) and DMSO (Sigma-Aldrich, D8418) and administered oral gavage 10mg/kg three days a week. The Vehicle Control was DMSO in 20% SBE-β-CD and administered oral gavage three days a week. Pancreas tissue from two mice per group was collected. The two tissue samples from each respective group were combined and submitted as one sample per group for library preparation.

first: Lincoln

last: Strickland

first: Wendao

first: Jennifer

last: Bailey-Lundberg


title: Upstream regulator of genomic imprinting in rice is a small RNA-associated chromatin remodeler

city: Bangalore

first: Shivaprasad

last: Padubidri V.

address: UAS-GKVK Campus

institute: National Centre for Biological Sciences - TIFR


title: Basal-to-inflammatory transition enhances basal cell carcinoma (BCC) therapy resistance via crosstalk with a pro-inflammatory stromal niche [RNA-seq II]

city: Stanford

first: Anthony

last: Oro

address: 269 Campus Drvie

institute: Stanford University

summary: The goal of these RNA-seq experiments was to determine transcriptomic gene expression changes caused by 7 days of Il1a and Osm combination treatment of mouse primary BCC tumor-derived organoids compared to PBS control

overall_design: Mouse primary BCC tumor tissues were embedded as small pieces in basement membrane extract and grown in organoid medium for 7 days. Cells were treated with either PBS control or Il1 and Osm ligand combination for 7 days in organoid medium.

first: Nancy

last: Li

first: Sadhana

last: Gaddam


title: Effect of rasagiline on the serum exosome miRNA transcriptome of Parkinson's disease patients (2023)

city: Bunkyo

first: Zhiyang

last: Yu

state: Tokyo to

address: Hongo 2-1-1

department: Department of Neurology

country: Japan

institute: Juntendo University

summary: Background: Parkinson's disease (PD), a neurodegenerative disease characterised by bradykinesia, rest tremor and rigidit, affects approximately 6.1 million people worldwide. Although its aetiology was attributed to accumulation of misfolded alpha-synuclein species and subsequent loss of dopaminergic neurons in the substantia nigra, recently, systemic factors contributing to its initiation and progression have gained increasing recognition. Specifically, exosomes, a kind of extracellular vesicles in the size range of ∼30 to ∼200 nm, have been highlighted as crucial mediators in orchestrating the intricate intercellular communication in PD. Among its cargos, miRNAs, with its ability to promote target mRNA degradation and inihibit translation, have been identiifed as promising biomarkers and therpaeutic targets. Nonetheless, the effect of anti-parkinsonism medication on the serum exosome miRNA profiles of PD patients remain lagrely unexplored. Objective: To examine the effects of rasagiline, a potentially neuroprotective monoamine oxidase B inihibitor, on the serum exosome miRNA profile of PD patients.

overall_design: To investigate the effect of rasagilne on the serum exosome miRNA profiles of PD patients, we established four independent cohorts. Two cohorts included PD patients and healthy controls, while the others recorded data from PD patients before and after one year of rasagililne treatment.

first: Shinji

last: Saiki

first: Nobutaka

last: Hattori


title: Effect of rasagiline on the serum exosome miRNA transcriptome of Parkinson's disease patients (2020)


title: Effect of rasagiline on the serum exosome miRNA transcriptome of Parkinson's disease patients (2022)


title: Effect of rasagiline on the serum exosome miRNA transcriptome of Parkinson's disease patients (2021)


title: Comparative network-based analysis of toll-like receptor agonist, L-pampo signaling pathways in immune and cancer cells

city: Seoul

first: Sera

last: Park

state: Seocho-gu

address: 15-19 Gangnam-daero 41-gil

institute: MOGAM Institute for Biomedical Research

summary: Toll-like receptors (TLRs) are critical immune components to stimulate immune responses against various infections. Recently, TLR agonists have emerged as a promising way to activate anti-tumor immunity. L-pampo, a TLR1/2 and TLR3 agonist, induces robust humoral and cellular immune responses but also causes cancer cell death. In this study, we investigated the L-pampo-induced signal pathways in immune and cancer cells and how they interact with TLRs and cancer-related pathways. First, we constructed a template network with differentially expressed genes and influential genes using the weighted gene co-expression network analysis (WGCNA). Next, we obtained perturbed modules using the same method and extracted core subnetworks from the modules by running Walktrap. Finally, we attempted to link the core subnetworks for reconstructing major molecular signals regarding the response to L-pampo utilizing a shortest path finding algorithm, TOPAS. Our analysis revealed that the key molecular processes were PI3K-AKT, JAK-STAT, and oxidative phosphorylation (OXPHOS) with reactive oxygen species (ROS) in immune and cancer cells. Notably, the OXPHOS process may be the crucial regulator for the differential responses of L-pampo between cancer cells of different tissue types. Our computational approaches can be applied to similar research for inferring underlying molecular mechanisms using complex gene expression profiles.

overall_design: To determine whether L-pampo drives the differential responses of cellular networks in different cell types, we chose a differentiated human monocyte cell line, THP-1, that highly expresses TLR2/3 and induces various immune responses to L-pampo. Additionally, we utilized two cancer cell lines, PC-3 and SW620, that can express TLR2/3 but manifest differential responses by L-pampo treatment. Then, we investigated gene signatures associated with L-pampo in immune cells and cancer cells using RNA-seq and various computational methods.

first: Geuntae

last: Kim

first: Eunyoung

last: Chun

first: Jung

last: Yum


title: Implicating type 2 diabetes effector genes in relevant metabolic cellular models using promoter-focused Capture-C [Capture-C]

city: Philadelphia

first: Matthew

last: Pahl

state: PA

address: 3615 Civic Center Blvd

department: Center for Spatial and Functional Genomics

institute: Children's Hospital of Philadelphia

summary: We used ATAC-seq, RNA-seq, and promoter to create atlases of the regulatory elements to provide context to type 2 diabetes associated variants from publically available genome-wide association studies

overall_design: We performed ATAC-seq, RNA-seq, and Promoter Capture C/HiC on EndoC-BH1, HepG2, and SGBS cells

first: Pahl

last: Matthew

first: Pippin

last: Jim

first: Struan

last: Grant


title: Implicating type 2 diabetes effector genes in relevant metabolic cellular models using promoter-focused Capture-C [Hi-C]


title: Comprehensive data for studying serum exosome microRNA transcriptome in Parkinson's disease patients


title: Implicating type 2 diabetes effector genes in relevant metabolic cellular models using promoter-focused Capture-C [RNA-Seq]


title: Implicating type 2 diabetes effector genes in relevant metabolic cellular models using promoter-focused Capture-C


title: Helicases DDX5 and DDX17 promote Hepatitis B Virus transcription termination heterogeneity in infected human hepatocytes

city: Lyon

first: Xavier

last: Grand

address: 46 allée d'Italie

country: France

institute: INSERM U1052, UCBL1, ENS de Lyon

summary: Background & Aims: Transcription termination fine tunes gene expression and contributes to specify the function of RNAs in eukaryotic cells. Transcription termination of hepatitis B virus (HBV) is subjected to the recognition of the canonical polyadenylation signal (cPAS) common to all viral transcripts. The regulation of the usage of this cPAS and its impact on viral gene expression and replication is currently unknown. Approach & Results: To unravel the regulation of HBV transcript termination, we used a 3’ RACE-PCR assay together with single molecule sequencing both in in vitro infected hepatocytes and in chronically infected patients. The detection of a previously unidentified transcriptional readthrough indicated that the cPAS was not systematically recognized during HBV replication in vitro and in vivo. After gene expression downregulation, followed by RNA and chromatin immunoprecipitation experiments, we showed the role of the RNA helicases DDX5 and DDX17 in promoting viral transcriptional readthrough, which was, furthermore, associated to HBV RNA destabilization. Moreover, downregulation of DDX5 and DDX17 allowed the precise termination of HBV transcripts at cPAS, which was associated with increased viral replication. Conclusions: Our findings identify DDX5 and DDX17 as crucial determinants for HBV transcriptional fidelity and as host restriction factors for HBV replication.

overall_design: To investigate the role of DDX5 and DDX17 in HepG2-NTCP cells

first: Giraud

last: Guillaume

first: Grand

last: Xavier


title: PMEL is involved in snake color pattern transition from blotches to stripes [bulk RNA-seq]

city: Geneva

first: Athanasia

last: Tzika

address: Athanasia Tzika, LANE - Sciences III, 30 quai Ernest Ansermet, 1211 Genève 4, Switzerland

institute: University of Geneva

summary: Our genomic, bulk and single-cell transcriptomic, functional, and developmental characterization of the Terrazzo corn snake color morph and the extensive comparison with wild-type snakes puts forward the dual role of PMEL in snake skin coloration, both in the differentiation of chromatophores during embryogenesis and the melanogenesis in melanophores.

overall_design: To find which genes are impacted in the Terrazzo corn snake morph we performed bulk RNA-seq from skin samples of Terrazzo and WT individuals

first: Asier

last: Ullate-Agote


title: Basal-to-inflammatory transition enhances basal cell carcinoma (BCC) therapy resistance via crosstalk with a pro-inflammatory stromal niche [RNA-seq]

first: Anthony

last: Oro

summary: The goal of these RNA-seq experiments was to determine transcriptomic gene expression changes caused by Il1 or Osm (or both) treatments of mouse BCC cells at 24h and 48h timepoints versus PBS control

overall_design: 2 biological replicates each

first: Nancy

last: Li

first: Sadhana

last: Gaddam


title: Basal-to-inflammatory transition enhances basal cell carcinoma (BCC) therapy resistance via crosstalk with a pro-inflammatory stromal niche [ATAC-Seq]

summary: The goal of these ATAC-seq experiments was to determine changes in chromatin accesiblity caused by Il1a and Osm ligand treatment of mouse BCC cells at 48h timepoint versus PBS control

overall_design: There were two experimental conditions in this ATACseq experiment: (1) PBS control and (2) ligand-treated mouse BCC cells. All samples were performed in biological replicates.


title: Basal-to-inflammatory transition enhances basal cell carcinoma (BCC) therapy resistance via crosstalk with a pro-inflammatory stromal niche [CUT&Run]

summary: The objective of this experiment was to determine the DNA binding sites of transcription factor NFKB1 in mouse BCC cells upon Il1a and Osm ligand treatment at 48h timepoint versus PBS control

overall_design: There were two experimental conditions in this CUT&RUN experiment: (1) PBS control and (2) ligand-treated mouse BCC cells. All samples were performed in biological replicates. An input control was used for normalization.


title: Basal-to-inflammatory transition enhances basal cell carcinoma (BCC) therapy resistance via crosstalk with a pro-inflammatory stromal niche


title: Implicating type 2 diabetes effector genes in relevant metabolic cellular models using promoter-focused Capture-C [ATAC-Seq]


title: RAD21 promotes oncogenesis and lethal progression of prostate cancer

city: Cambridge

first: Sofia

last: Hu

email: sofiahu@mit.edu, xsu@cpdr.org

address: 500 Main Street

institute: MIT

summary: Higher levels of aneuploidy, characterized by imbalanced chromosome numbers, are associated with lethal progression in prostate cancer. However, how aneuploidy contributes to prostate cancer aggressiveness remains poorly understood. In this study, we assessed in patients which genes on chromosome 8q, one of the most frequently gained chromosome arms in prostate tumors, were most strongly associated with long-term risk of cancer progression to metastases and death from prostate cancer (lethal disease) in 403 patients and found the strongest candidate was cohesin subunit gene, RAD21, with an odds ratio of 3.7 (95% CI 1.8, 7.6) comparing the highest vs. lowest tertiles of mRNA expression and adjusting for overall aneuploidy burden and Gleason score, both strong prognostic factors in primary prostate cancer. Studying prostate cancer driven by the TMPRSS2-ERG oncogenic fusion, found in about half of all prostate tumors, we found that increased RAD21 alleviated toxic oncogenic stress and DNA damage caused by oncogene expression. Data from both organoids and patients indicate that increased RAD21 thereby enables aggressive tumors to sustain tumor proliferation, and more broadly suggests one path through which tumors benefit from aneuploidy.

overall_design: Ventral prostate tissue was separated from mice carrying the desired genotype and developed into organoids. Two organoid clones from each genotype (control Tmprss2-fl/fl-ERG and Tmprss2-ERG) were used.

first: Xiaofeng

last: Su

first: Duanduan

last: Ma

last: Vander Heiden


title: SMYD5 is a regulator of the mild hypothermia response [RNA-Seq]

city: Reykjavik

first: Hans

middle: Tomas

last: Bjornsson

state: Hofudborgarsvaedid

address: Sturlugata 8

department: Faculty of Medicine

country: Iceland

institute: University of Iceland

summary: Organisms have homeostatic mechanisms to respond to cold temperature to ensure survival including the activation of the mammalian neuroprotective mild hypothermia response (MHR) at 32°C. Utilizing a forward CRISPR-Cas9 mutagenesis screen, we identify the histone lysine methyltransferase SMYD5 as an epigenetic gatekeeper of the MHR. SMYD5 represses the key MHR gene SP1 at euthermia but not at 32°C. This repression is mirrored by temperature-dependent levels of H3K36me3 at the SP1-locus and globally indicating that the mammalian MHR is regulated at the level of histone modifications. We identified additional SMYD5-temperature dependent genes suggesting a broader MHR-related role for SMYD5. Our study provides an example of how the epigenetic machinery integrates environmental cues into the genetic circuitry of mammalian cells and suggests novel therapeutic avenues for neuroprotection after catastrophic events.

overall_design:
The purpose of doing an RNASeq on mouse Neural progenitor cells from cortex was to extend the list of gene transcripts known to react to the mild hypothermic response.
The purpose of doing an RNASeq on cortical and hippocampal cells from neonatal P10 mice exposed to hypoxic injury in vivo after euthermic and hypothermic (32°C) treatment was to extend the list of gene transcripts known to react to the mild hypothermic response.

first: Salvor

last: Rafnsdottir

first: Kijin

last: Jang

first: Sara

middle: T

last: Halldorsdottir

first: Meghna

last: Vinod

first: Arnhildur

last: Tomasdottir

first: Katrin

last: Möller

first: Tinna

last: Reynisdottir

first: Laufey

last: Atladottir

first: Kristin

last: Allison

first: Kevin

last: Ostacolo

first: Jin

last: He

first: Li

last: Zhang

first: Frances

last: Northington

first: Erna

last: Magnusdottir

first: Raul

last: Chavez-Valdez

first: Kimberley

last: Anderson


title: SMYD5 is a regulator of the mild hypothermia response [CRISPR-Cas9 screen]

first: Hans

last: Bjornsson

summary: Organisms have homeostatic mechanisms to respond to cold temperature to ensure survival including the activation of the mammalian neuroprotective mild hypothermia response (MHR) at 32°C. Utilizing a forward CRISPR-Cas9 mutagenesis screen, we identify the histone lysine methyltransferase SMYD5 as an epigenetic gatekeeper of the MHR. SMYD5 represses the key MHR gene SP1  at euthermia but not at 32°C. This repression is mirrored by temperature-dependent levels of H3K36me3 at the SP1-locus and globally indicating that the mammalian MHR is regulated at the level of histone modifications. We identified additional SMYD5-temperature dependent genes suggesting a broader MHR-related role for SMYD5. Our study provides an example of how the epigenetic machinery integrates environmental cues into the genetic circuitry of mammalian cells and suggests novel therapeutic avenues for neuroprotection after catastrophic events.

overall_design: To find additional regulators of the mild hypothermic response we performed a genome wide CRISPR-Cas9 screen using the SP1-MHI (HEK293+Cas9+SP1-MHI). We selected for stable sgRNAs with puromycin selection for 6 days and 8 hours and exposesd to 32°C the last 16h before FACS. We sorted our 5% most and least fluorescent cells. Next generation sequencing (NGS) was performed with single read, 80 cycles and 8 indexing cycles with PhiX spike in of 20% on NovaSeq S4 at deCODE genetics for HEK293WT+Cas9+SP1 and HEK293WT+Cas9(control). We did this experiment in 4 biological replicates.

middle: E


title: SMYD5 is a regulator of the mild hypothermia response

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: Metabolic activity affects RPE fate during cell reprogramming

city: Baltimore

first: Seth

last: Blackshaw

state: MD

address: 733 N Broadway

department: Neuroscience

institute: Johns Hopkins University

summary: To investigate transcriptomic changes that facilitate the reprogramming of retinal pigment epithelium (RPE) cells to neural retina, chicken RPE explants were cultured for 24 or 48 hours in the presence or absence of the reprogramming factor FGF2. To further interrogate the metabolic requirements of reprogramming, RPE explants were additionally cultured in the presence of the pyruvate dehydrogenase kinase inhibitor dichloroacetate in the presence or absence of FGF2.

overall_design: RPE explants were collected from specific pathogen free chickens at embryonic day 4. Biological replicates were collected in triplicate. Three explants were pooled per biological replicate. Explants were treated with A) maintenance media (control), B) 10 ng / ml FGF2, c) 20 mM dichloroacetate, or D) both 10 ng / ml FGF2 and 20 mM dichloroacetate. Each condition was assayed at both 24 and 48 hours in culture.

first: J

last: Perez-Estrada

last: Tangeman

first: M

last: Proto-Newton

first: H

last: Sanaka

first: B

last: Smucker

first: K

last: Del Rio-Tsonis


title: Single cell RNA sequencing of human PDAC-derived PDX

city: Shanghai

first: Christopher

last: Heeschen

address: South Chongqing Road 227

institute: Shanghai Jiao Tong University School of Medicine

overall_design: Three treatment samples of PDAC-003 ( CTRL, MCM-treatment, ETO-treatment) were labeled with HTO1 (-GTCAACTCTTTAGCG-), HTO2 (-TGATGGCCTATTGGG-), HTO3(-TTCCGCCTCTCTTTG-) separately, Subsequently pooled three treatments from  same PDAC samples together. As for PDAC-002 and PDAC-021 samples, two treatments (CTRL, MCM-treatment) were labeled with HTO1 (-GTCAACTCTTTAGCG-)  and HTO2 (-TGATGGCCTATTGGG-) . CTRL of PDAC-002 and PDAC-021, MCM-treatment of PDAC-002 and PDAC-021 were pooled together separately.

first: Minchun

last: Chen

first: Zhenyang

last: Guo


title: Novel Targets for Diabetic Foot Ulcer Healing: A Transcriptomic Analysis by Negative Pressure Wound Therapy

status: Public on Jul 25 2024

city: hefei

first: Ying

last: Tang

address: No.218 Jixi Road, Shushan District, Hefei City, Anhui Province, China.

institute: Department of Endocrinology, the First Affiliated Hospital of Anhui Medical University

summary: The results of the present study provide new insights into the mechanism through which NPWT promotes DFU wound healing

overall_design: Participants underwent wound debridement to remove blackened and necrotic soft and bone tissues, followed by NPWT. For NPWT, a vacuum assisted closure (VAC) negative pressure-assisted healing therapy system (KCI, USA) was used according to published protocols , at a dose of -125 mmHg for 1 week. Full-thickness skin tissue was taken within 0.5 cm of the wound edge before and after 1 week of NPWT, only after removing the negative pressure device. The collected granulation tissue was stored at -80ºC for examination.RNA-seq was performed on the granulation tissue of three patients with DFU before and after NPWT (before/after-patient 1, before/after-patient 2, before/after-patient 3)


title: RNA-seq of live cell 3-D models with engineered genetic sub-types of lung cancer

city: Atlanta

first: Adam

last: Marcus

state: GA

address: 1365C Clifton Rd., N.E., Room 4092

institute: Emory University

summary: An experimental system was designed to screen for targetable signaling pathways linked to early 3D invasion  in molecular subtypes, TP53 and LKB1, of KRAS-driven lung adenocarcinoma (LUAD). Live-cell imaging of human bronchial epithelial cells during 3D invasion was combined with RNA transcriptome profiling shown here.

overall_design: RNA-seq was performed in triplicate on for each sub-type. Total RNA was extracted from the spheroids at day 7 of 3D spheroid invasion assay using a RNeasy Kit (Qiagen, Valencia, CA, USA). The quantitation, integrity and purity of the extracted total RNA samples were assessed by the Emory Integrated Genomics Core (EIGC) using 2100 Bioanalyzer (Agilent) and RNA sequencing was performed by Novogene, Co., Ltd. Data processing, quality control, read alignment and statistical analyses were performed by the Emory Biostatistics and Bioinformatics Shared Resource, as previously described(21). Raw read data (fastq) QC was performed using FastQC v0.11.7. Post-filtered reads were mapped against Ensemble Human GRCh38.p12 release 95 reference genome using the STARaligner v2.7.0e. Expression quantification was obtained using featureCounts. Normalization and pairwise differential analysis were determined using median-ratios method in DESeq2 and log2 transformed. Differentially expressed genes (DEGs) were identified using a moderated t-test as implemented in the Limma R package. Heatmaps were created by unsupervised clustering of log2 transformed normalized expression data for the significant DEGs, and the resulting data were analyzed using a Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG).

first: Junghui

last: Koo

first: Chang-Soo

last: Seong

first: Rebecca

last: Parker

first: Bhakti

last: Dwivedi

first: Robert

last: Arthur

first: Ashok

last: Dinasarapu

last: Johnston

first: Henry

last: Claussen

first: Carol

last: Tucker-Burden

first: Suresh

middle: S

last: Ramalingam

first: Haian

last: Fu

first: Wei

last: Zhou

middle: I

first: Melissa

last: Gilbert-Ross


title: Disparate pathways for extrachromosomal DNA biogenesis and genomic DNA repair

first: Julia

last: Belk

state: California

address: 450 Serra Mall

summary: Oncogene amplification on extrachromosomal DNA (ecDNA) is a pervasive driver event in cancer, yet our understanding of how ecDNA forms is limited. Here, we couple a CRISPR-based method for ecDNA induction with extensive characterization of newly formed ecDNA to examine their biogenesis. We find that DNA circularization is efficient, irrespective of 3D genome context, with formation of 800kb, 1 Mb, and 1.8 Mb ecDNAs reaching or exceeding 15%. We show non-homologous end joining and microhomology-mediated end joining both contribute to ecDNA formation, while inhibition of DNA-PKcs and ATM have opposing impacts on ecDNA formation. EcDNA and the corresponding chromosomal excision scar can form at significantly different rates and respond differently to DNA-PKcs and ATM inhibition. Taken together, our results support a model of ecDNA formation in which double strand break ends dissociate from their legitimate ligation partners prior to joining of illegitimate ends to form the ecDNA and excision scar.

overall_design: Cells were electroporated with 2 Cas9•RNPs targeting the same chromosome arm to generate ecDNAs in the context of different drug treatments or genetic knockouts. 24 hours after electroporation samples were harvested and gDNA isolated. Targeted deep sequencing of the ecDNA (circle) or chromosomal (scar) junctions was performed.

first: John

last: Rose

first: Paul

last: Mischel

first: Howard

last: Chang


title: Genome-wide mapping of native G-loops in living cells

city: Chengdu

first: Zhihong

last: Xue

state: Sichuan

address: No. 20, 3rd section, South Renmin Road

institute: Sichuan University

summary: G-quadruplex (G4) are four‑stranded DNA secondary structures that form in guanine‑rich regions of the genome, which can enhance or repress gene expression. An R-loop is a special triple-stranded nucleic acid structure formed when nascent RNA invades double-stranded DNA (dsDNA) during transcription. G-loops are constituted by one or more DNA G4 on one strand and a stable RNA/DNA hybrid on the other. We developed the HepG4-seq for  mapping the native G4s and the HBD-seq for mapping native R-loops. We combined the HepG4-seq and HBD-seq to profile the genomic native G-loops, which are regions co-occupied by both native G4s and R-loops, in both HEK293 cells and mouse embryonic stem cells (mESCs).

overall_design: Cleavage Under Targets and Tagmentation (CUT&Tag) for mapping G4 and R-loop in HEK293 and mESC cells.


title: Effect of knockdown of GRP94 on gene expression of MKN-45 cells

first: Qimeng

last: Shi

address: Ruijin Second Road

institute: Shanghai Jiaotong University School of Medcine

summary: Anoikis resistance allows cancer cells to avoid death caused by detachment from the extracellular matrix's adhesion, enabling these cells to infiltrate and migrate to regions such as the peritoneum.Glucose-regulated protein 94 (GRP94, alternatively labeled as HSP90B1) is typically stress-induced and is part of the heat shock protein 90 (Hsp90) family.GRP94 is closely tied to cellular resistance to stress, apoptosis, proliferation, invasion, inflammation, and immunity.Our findings indicated that while attached culture didn't impact the apoptosis rate between shGRP94 and shCtrl groups, GRP94 knockdown weakened anoikis resistance, migration, and invasion abilities. Here, we aim to assess the impact of knocking down GRP94 on gene expression in the gastric cancer cell line MKN-45, to further elucidate potential molecular mechanisms by which GRP94 affects the biological characteristics of gastric cancer cells.

overall_design: To assess the impact of knocking down GRP94 on gene expression in the gastric cancer cell line MKN-45, GRP94 was  knocked down with shRNA.

first: Chen

last: Li


title: Single cell RNA-sequencing of esophageal epithelium from wild type and p63-KO mice

city: New York

first: Yinshan

last: Fang

state: New York

address: 650W 168th Street

institute: Columbia University Medical Center

summary: Cell fate determination and maintenance play critical roles in establishing and preserving tissue identity and function during organ development. Aberrant cell fate transitions can lead to cancer cells acquiring lineage plasticity, resulting in tumor heterogeneity and resistance to treatment. Trp63 (p63) is a transcription factor important for the formation of stratified squamous epithelium in the esophagus. Here, we report an unexpected role of p63 in protecting esophageal basal progenitor cells from committing to a neuroendocrine fate. Combination of single cell RNA sequencing analysis, lineage tracing and in vitro differentiation, we found that deletion of p63 in developing murine esophagus and in human embryonic stem cells results in extensive differentiation towards neuroendocrine lineage.

overall_design: We isolated esophageal epithelium from wild type and p63-KO mice at E18.5 and then applied for single cell RNA-sequencing

first: Yongchun

last: Zhang

first: Jian

last: Ge

first: Jianwen

last: Que


title: SMYD5 is a regulator of the mild hypothermia response [Cut & Run]

first: Hans

last: Bjornsson

overall_design: To look further at epigenetic modifications both in SMYD5 KD and WT cells and SMYD5 binding at 37°C and 32°C we decided to do CUT&RUN. We followed the Epicypher CUTANA protocol (v.1.7) on 300.000 cells per sample. We permeabilized the cells with 0.01% digitonin (Sigma, D141). For normalization we spiked in E. Coli DNA (Epicypher, 23618-1401) at the final concentration of 0.2 ng per sample. The library preparation was performed with TrueSeq-Chip Sample preparation Kit (Illumina, IP-202-9001). The following antibodies were used for the CUT&RUN: CTCF antibody (Cell Signaling, 2899S), H3K36me3 (Thermo Fisher Scientific, MA5-24687), H4K20me3 (EpiCypher, 13-0054), H3K9me3 (Abcam, ab176916), H3K4me3 (EpiCypher, 13-0041), H3K27me3 (Thermo Fisher Scientific, MA5-11198), SMYD5 (custom), and Rabbit IgG Antibody (Epicypher, 23613-0042). The sequencing was performed on a NovaSeq 6000 S4 at deCODE genetics, paired-end 8 cycles of index1, 8 cycles of index 2, 150 cycles read 1 and, 150 cycles read 2. Some samples were sequenced using a NextSeq 550 using the NextSeq 500/550 Mid Output Kit v2.5 (20024904) with 75 bp paired-end sequencing.  For all samples  we aimed for 10 million reads per sample.

first: Salvor

last: Rafnsdottir

first: Kijin

last: Jang

first: Sara

last: Halldorsdottir

first: Meghna

last: Vinod

first: Arnhildur

last: Tomasdottir

first: Katrin

last: Möller

first: Tinna

last: Reynisdottir

first: Laufey

middle: H

last: Atladottir

first: Kristin

last: Allison

first: Kevin

last: Ostacolo

first: Jin

last: He

first: Li

first: Frances

last: Northington

first: Erna

last: Magnusdottir

first: Raul

last: Chavez-Valdez

first: Kimberley

last: Anderson


title: Lipid engulfed macrophages interfere with protective T cell immunity and favor tumor progression in human colon carcinogenesis - Monocyte

city: Orbassano

first: Giulio

last: Ferrero

state: TO

address: Regione Gonzole, 10

department: Dept. Clinical and Biological Sciences

institute: University of Turin

summary: Obesity and altered lipid metabolism are associated with colorectal cancer (CRC). In this context, the presence of colonic lipid-engulfed macrophages (foam cells, FC) may hinder protective T cell immunity, thus favoring gut carcinogenesis.

overall_design: Triplicate RNASeq was performed on RNA extracted from In vitro-derived FC generated by culturing sorted CD14+ cells from PBMCs of healthy donors with M-CSF stimulating factor. Cells were grown in the absence or presence of oxLDL for 48h.

first: Elena

last: Daveri

first: Licia

last: Rivoltini


title: RNA-seq analysis of wild-type HEK293, HEK293 treated with DMSO, ML216 or NSC617145, wild-type mESCs and dhx9KO mESCs

first: Zhihong

last: Xue

summary: G4s are built by stacked guanine tetrads connected via Hoogsteen hydrogen bonds and can be formed by intra- or inter-molecular folding of the tetramers. R-loops are three-stranded structures which contain a DNA-RNA hybrid and a displaced single-stranded DNA. Both G4s and R-loops are involved in key biological processes, including transcriptional regulation, replication, genomic instability, class switch recombination in B cells, DNA damage and repair, and telomere maintenance. To understand the transcriptional regulation of G4s and R-loops, we performed the RNA-seq analysis on HEK293 cells treated with the inhibitors of G4 resolving helicase ML216 or NSC617145 and mESCs in which the resolving helicase DHX9 of G4 and R-loop was knockout.

overall_design: To investigate the effect of ML216 and NSC617145 in HEK293,we treated HEK293 with corresponding helicase inhibitors and performed RNA-seq. To invistegate the effect of Dhx9 in mESC,we established dhx9 knockout mESC using CRISPR/Cas9 and performed RNA-seq.


title: p65 signaling dynamics drive the developmental progression of hematopoietic stem and progenitor cells through cell cycle regulation

city: Ames

first: Raquel

last: Espin Palazon

state: Iowa

address: 2213 Pammel Drive, 3003 ATRB

institute: Iowa State University

summary: While hematopoietic stem and progenitor cells (HSPCs) and the signaling pathways that specify them have been well defined in vivo, deriving functional HSPCs from human pluripotent stem cells (hPSCs) is still an outstanding challenge in the stem cell field. This has impaired efforts to use patient-specific HSPCs and their derivatives to treat leukemia, lymphoma, anemia, and solid cancers. Identifying the missing element to derive functional human HSPCs will be critical to advance these therapies. Here, we found that in contrast to in vitro protocols where signaling pathways are typically manipulated statically, the natural developmental progression of HSPC fate in the embryo requires a precise and temporally controlled oscillatory signaling dynamics.  Particularly, our work reveals that NF-kB signaling works as an oscillatory clock that controls the developmental progression of HSPCs by driving phases of quiescence and proliferation precisely during embryonic development. This was revealed by live imaging of a custom NF-kB zebrafish reporter line, in conjunction with transcriptomic profiling and precise temporal manipulations of the NF-kB family member p65, the driver of these signaling events. We also show that in vitro protocols of human hematopoietic differentiation from iPSCs depend on initial pro-inflammatory signaling activation. In summary, our results uncover the functional role of pro-inflammatory signaling during HSPC specification, revealed their oscillatory dynamics, and define this signaling pathway as the main driver of HSPC fate progression. The integration of these dynamics in vitro will be fundamental to achieve the long-standing goal of generating and expanding patient derived functional HSPCs.

overall_design: To gain insights into how NF-kB in endothelium was contributing to HSPC specification, NFKB+ or NFKB- endothelial cells (kdrl+) were sorted and collected from 22hpf zebrafish embryos in triplicate and subject to RNA-seq.

first: Clyde

last: Campbell

first: Rodolfo

last: Calderon

first: Giulia

last: Pavani

first: Radwa

last: Barakat

first: Xiaoyi

last: Cheng

first: Fang

last: Liu

first: Xiyu

last: Peng

first: Karin

last: Dorman

first: Jeffrey

last: Essner

first: Maura

last: McGrail

first: Paul

last: Gadue

first: Deborah

middle: L

last: French

last: Espin-Palazon


title: FoxO transcription factors actuate the formative pluripotency specific gene expression programme

city: Vienna

first: Martin

last: Leeb

address: Dr.-Bohr-Gasse 9/3

department: Max Perutz Labs

country: Austria

institute: University of Vienna


title: FoxO transcription factors actuate the formative pluripotency specific gene expression programme [ChIP-Seq]

summary: Naïve pluripotency is sustained by a self-reinforcing gene regulatory network (GRN) comprising core and naïve pluripotency-specific transcription factors (TFs). Upon exiting naïve pluripotency, ES cells transition through a formative post-implantation-like pluripotent state. However, the mechanisms underlying disengagement from the naïve GRN and initiation of the formative GRN remain unclear. Here, we demonstrate that phosphorylated AKT acts as a gatekeeper that prevents nuclear localization of FoxO TFs in naïve ESCs. PTEN-mediated reduction of AKT activity allows nuclear entry by FoxO TFs, enforcing a cell fate transition by binding and activating formative pluripotency-specific enhancers. Indeed, FoxO TFs are necessary and sufficient for transition from the naïve to the formative pluripotent state. Our work uncovers a pivotal role for FoxO TFs and AKT signalling in mechanisms underlying the exit from naïve pluripotency, a critical early embryonic cell fate transition.

overall_design: Chromatine-immunoprecipitation DNA sequencing (ChIP-seq) for FoxO1 and FoxO3 in WT and Pten KO mESC, in pluripotency supporting conditions (2i) and 24hours after 2i withdrawal (N24).

first: Laura

last: Santini

first: Saskia

last: Kowald

first: Giovanni

last: Sestini

first: Nicolas

last: Rivron


title: FoxO transcription factors actuate the formative pluripotency specific gene expression programme [RNA-Seq]

overall_design: Quantseq experiments from ES cells frown in 2i and in differentiation conditions in N2B27 medium; WT, Pten KO, Tsc2 KOs treated with DMSO and with a variety of inhibitorrs are used


title: FoxO transcription factors actuate the formative pluripotency specific gene expression programme [ATAC-Seq]

overall_design: ATAC-seq of WT and Pten KO mESC, in pluripotency supporting conditions (2i) and 24hours after 2i withdrawal (N24).


title: Transcriptomics of PLXDC2 knockdown BMDM treated with PTEN

city: shanghai

first: jiaming

last: shen

address: SHANGHAI

institute: shanghai jiaotong university

summary: To evaluate whether PLXDC2 mediated the role of PTEN on macrophages, PLXDC2 was knocked down in M2-polarized BMDMs, and the resultant PLXDC2-knocked down cells together with control cells were treated with or without PTEN, before subjected to global transcriptome profiling.

overall_design: Bone-marrow (BM) cells were harvested from femurs and tibia of C57BL/6J mice. Differentiation was induced by recombinant mouse M-CSF in DMEM medium containing 1% penicillin/streptomycin and 10% FBS for 7 days to obtain the bone marrow-derived macrophages (BMDMs).To induce an M2-like phenotype, we treated BMDMs with IL-4 and IL-13 for 48 h.M2 BMDMs transfected with siNC or siPLXDC2#1,2 were treated with and without 100 ng/mL PTEN for 24 h, followed by RNA-seq analysis.


title: Transcriptomics of M0 macrophages treated with PTEN

summary: While naive BMDMs without further treatment are referred to as M0-like macrophages, which can be polarized to either M1-like or M2-like macrophages,To assess the consequence of PTEN binding on macrophages, we profiled the global transcriptome of M0-like BMDMs with and without PTEN treatment.

overall_design: Bone marrow-derived naive macrophages (M0 BMDMs) isolated from mice with and without treatment by 100 ng/mL PTEN for 24 h were subjected to RNA-seq.


title: Transcriptomics of BMDM treated with PTEN in the presence or absence of STAT1 inhibitors

summary: PTEN binds to and regulates polarization of TAMs to enhance antitumor immunity and suppress tumor growth，We hope to investigate whether STAT1 mediates the polarization effect of PTEN on TAMs as a downstream factor

overall_design: Bone-marrow (BM) cells were harvested from femurs and tibia of C57BL/6J mice. Differentiation was induced by recombinant mouse M-CSF in DMEM medium containing 1% penicillin/streptomycin and 10% FBS for 7 days to obtain the bone marrow-derived macrophages (BMDMs).To induce an M2-like phenotype, we treated BMDMs with IL-4 and IL-13 for 48 h.M2 BMDMs were treated with and without 100 ng/mL PTEN for 24 h in the presence or absence of STAT1 inhibitors (Fludarabine), followed by RNA-seq analysis.


title: Effect anti-TNF-alpha treatment on CD4+ T cell activation

city: Birmingham

first: Sarah

last: Dimeloe

address: Edgbaston

institute: University of Birmingham

summary: The experiment was designed to test how autocrine TNF-alpha impacts human CD4+ T cell activation. Naïve CD4+ T cells from healthy human blood donors were either left unactivated or activated with anti-CD3/anti-CD28 in presence of an isotype control antibody or anti-TNF-alpha antibody. RNA was extracter at 48 hours. Analysis revealed that anti-TNF-alpha treatment prevented upregulation of activation-induced T cell metabolic, signaling and effector molecules.

overall_design: Naïve CD4+ T cells from healthy human blood donors were either left unactivated or activated with anti-CD3/anti-CD28 in presence of an isotype control antibody or anti-TNF-alpha antibody. RNA was extracted at 48 hours.

first: Emma

last: Bishop


title: Research on the Mechanism of PTEN Mediated tumor-associated macrophages (TAMs) Polarization

overall_design: Refer to individual Series


title: Prohibitin 2 orchestrates transcription of long noncoding RNA and coding gene to accelerate tumorigenesis

first: He

last: Zhang

address: Tongji University, 1239 Siping Road, Yangpu District, Shanghai

institute: School of Life Science and Technology, Tongji University

summary: The spatial co-presence of aberrant long non-coding RNAs (lncRNAs) and abnormal coding genes contribute to the development of the malignancy in various tumors. However, precise coordinated mechanisms underlying this co-presence phenomenon in tumorigenesis remains incompletely understood. Here, we for the first time showed that PHB2 coordinately crosslinked transcription of a novel oncogenic CANT1-New-Isoform 2 (CANT2) lncRNA and coding tumor-suppressor gene CCBE1 for accelerating tumorigenesis of melanoma. In melanoma cells, PHB2 initially accessed open chromatin sites of the promoter of CANT2 lncRNA and recruited MLL2, resulting in an augmented H3K4 trimethylation and activating the transcription of CANT2. Intriguingly, PHB2 further bound with activated CANT2 transcript for targeting interaction with the promoter of tumor-suppressor CCBE1, recruiting histone deacetylase HDAC1 to decrease H3K27 acetylation of CCBE1 promoter and inhibiting CCBE1 transcription for significantly promoting cell growth and metastasis in vitro and in vivo. Our study elucidated a novel PHB2-mediating mechanism that crosslinked aberrant transcription of lncRNA and coding gene, thereby providing an interesting “one stone two birds” model in tumorigenesis.

overall_design: We first generated CANT2 lncRNA knockout stable A375 cell lines with CRISPR/Cas9. Then RNA-seq was conducted on RNAs extracted from these CANT2-KO and control A375 cells.

first: Tianyi

last: Ding


title: scRNA-Seq analysis of adult glioblastoma patient samples

city: Zurich

first: Berend

last: Snijder

address: Otto-Stern-Weg 3, HPM building

institute: ETH Zurich

summary: Glioblastoma is the most common type of malignant brain tumor among adults. We used single-cell RNA sequencing (scRNA-seq) to analyze the diversity of glioblastoma cells.

overall_design: Dissociated and cryopreserved single-cell suspensions of patient samples  (P007, P011, P012, P013) were processed by fluorescence-activated cell sorting (FACS) to isolate viable CD45+ immune cells and CD45- glioblastoma cells.

first: Sohyon

last: Lee

first: Tobias

last: Weiss


title: Transcriptional response to MMS in haploid and diploid Schizosaccharomyces pombe

status: Public on Jul 24 2024

city: Los Angeles

first: Joshua

last: Park

address: 1050 Childs Way Room 108

department: Molecular and Computational Biology

institute: University of Southern California

summary: Polyploidy has been implicated in genome instability and tumorigenesis. We use Schizosaccharomyces pombe diploids as a model for studying the consequences of whole genome duplications on genome integrity. In this study, our aim is to investigate the transcriptional profile between haploid and diploid S. pombe in unperturbed and MMS treated conditions (0.0075% MMS, 4 hours @ 32 degrees Celsius in YES media).

overall_design:
Investigate the transcriptional profile between haploid and diploid S. pombe in unperturbed and MMS treated conditions (0.0075% MMS, 4 hours @ 32 degrees Celsius in YES media).
Total RNA was extracted using Qiagen RNeasy kit. ERCC spike-ins were added in equal volume to all samples. Samples were sent to Novogene for mRNA purification (polyA selection), cDNA synthesis, library preparation, and paired-end sequencing.

first: Susan

last: Forsburg


title: Epigenetic regulation of p63 blocks squamous-to-neuroendocrine transdifferentiation in esophageal development and malignancy

first: Dimitris

last: Karagiannis

state: NY

institute: Columbia University

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


title: Ceramide metabolism alterations contribute to Tumor Necrosis Factor-induced melanoma dedifferentiation and predict resistance to immune checkpoint inhibitors in advanced melanoma patients [WM35]

city: Toulouse

first: Marie

last: Tosolini

address: 2 avenue Hubert Curien

department: CRCT

institute: INSERM

summary: Introduction: Advanced cutaneous melanoma is a skin cancer characterized by a poor prognosis and high metastatic potential. During metastatic spread, melanoma cells often undergo dedifferentiation toward an invasive phenotype, resulting in reduced expression of microphthalmia-associated transcription factor (MITF)-dependent melanoma antigens and facilitating immune escape. Tumor Necrosis Factor (TNF) is known to be a key factor in melanoma dedifferentiation. Interestingly, accumulating evidence suggests that TNF may play a role in melanoma progression and resistance to immunotherapies. Additionally, TNF has been identified as a potent regulator of sphingolipid (SL) metabolism, which could contribute to melanoma aggressiveness and the process of melanoma dedifferentiation. Methods: We conducted RNA sequencing and mass spectrometry analyses to investigate TNF-induced dedifferentiation in two melanoma cell lines. In vitro experiments were performed to manipulate sphingolipid metabolism using genetic or pharmacologic alterations in combination with TNF treatment, aiming to elucidate the potential involvement of this metabolism in TNF-induced dedifferentiation. Lastly, to evaluate the clinical significance of our findings, we performed unsupervised analysis of plasma sphingolipid levels in 48 patients receiving treatment with immune checkpoint inhibitors, either alone or in combination with anti-TNF therapy. Results: Herein, we demonstrate that TNF-induced melanoma cell dedifferentiation is associated with a global modulation of sphingolipid metabolism. Specifically, TNF decreases the expression and activity of acid ceramidase (AC), encoded by the ASAH1 gene, while increasing the expression of glucosylceramide synthase (GCS), encoded by the UGCG gene. Remarkably, knockdown of AC alone via RNA interference is enough to induce melanoma cell dedifferentiation. Furthermore, treatment with Eliglustat, a GCS inhibitor, inhibits TNF-induced melanoma cell dedifferentiation. Lastly, analysis of plasma samples from patients treated with immune checkpoint inhibitors, with or without anti-TNF therapy, revealed significant predictive sphingolipids. Notably, the top 8 predictive sphingolipids, including glycosphingolipids, were associated with a poor response to immunotherapy.  Discussion: Our study highlights that ceramide metabolism alterations are causally involved in TNF-induced melanoma cell dedifferentiation and suggests that the evolution of specific ceramide metabolites in plasma may be considered as predictive biomarkers of resistance to immunotherapy.

overall_design: WM35 were seeded at Day 0 and then treated in reverse kinetics for 6, 24, 48 and 72 hours with 50 ng/mL TNF or transfected with siCTRL or siASAH1-1 for 72 hours

first: Carine

last: Dufau

first: Bruno

last: Ségui


title: Ceramide metabolism alterations contribute to Tumor Necrosis Factor-induced melanoma dedifferentiation and predict resistance to immune checkpoint inhibitors in advanced melanoma patients [A375]

overall_design: A375 cells were seeded and then treated with or without 50 ng/mL TNF for 24, 48 or 72 hours


title: Effect of PTEN treatment on the subcutaneous tumor formation model of PTEN knockout B16F10 cells in C57BL/6J mice

summary: PTEN treatment can affect the tumor growth of PTEN knockout B16-F10 cells in C57BL/6J mice，We hope to analyze the changes that occur in various cells in tumors through single-cell RNA sequencing.

overall_design: PTEN-knockout B16F-10 cells were subcutaneously injected into C57BL/6J mice (1.0×10^5 cells per mouse). After tumors reached approximately 100 mm3 in volume, PTEN (10 μg/day, 7 days) or PBS were intratumorally injected and tumors were harvested for scRNAseq


title: Transcriptomics of PTEN knockout B16-F10 cells in C57BL/6J mice model treated with PTEN

summary: extracellular PTEN treatment did not affect the growth of PTEN knockout B16-F10 cells cultured in vitro. , To investigate whether extracellular PTEN act on the tumor microenvironment to exert a tumor-suppressive role in vivo，molecular changes caused by PTEN treatment inside the B16-F10-PTEN tumors were monitored by RNA sequencing.

overall_design: 1.0×105 PTEN knockout B16-F10 cells were subcutaneously injected into C57BL/6 mice.once tumors reached ~ 50-100 mm3 in volume, mice were randomly assigned to control or treatment groups and were intratumorally or intraperitoneally injected with PTEN proteins or PBS every day. tumors were harvested for scRNAseq


title: Expression changes in respiratory epithelial cells when treated with TGF-β1

city: Ilsandong-gu, Goyang-si

first: Dong-Hee

last: Han

state: Gyeonggi-do

address: 32, Dongguk-ro

department: Life science

institute: Dongguk University

summary: TGF-β1 is involved in various biological processes through downstream signals including SMAD, Akt, Erk, etc. TGF-β1 overexpressed by harmful stimuli can act as a causative factor in diseases such as accumulation of extracellular matrix and development of cancerous characteristics. Therefore, we sought to compare changes in expression of intracellular miRNAs to study diseases caused by TGF-β1. In our study, we aimed to discover therapeutic targets for TGF-β1-induced disease by investigating changes in miRNA expression and identifying the correlation between genes that can be regulated by miRNA.

overall_design: To investigate changes in the expression of two respiratory epithelial cells according to TGF-β1 exposure, A549 and BEAS-2B were treated with TGF-β1 at 1 and 5 ng/mL for 2 days.

first: Min

last: Kim


title: Effect of trained immune response in brain [RNA-Seq]

city: Munich

first: Vini

last: Tiwari

address: Feodor Lynen Straße 17

country: Germany

institute: Technical University Munich, (TUM), Deutsches Zentrum für Neurodegenerative Erkrankungen

summary: To investigate the transcriptomic profile of cells involved in trained immune response , we performed RNA seq on unlesioned tissue sections isolated from mice treated with BCG or Saline

overall_design: RNA-seq of lesion tissue isolated from mice treated with BCG or saline.

first: Bharat

last: Prajapati

first: Hao

last: Ji

first: Mikael

last: Simons


title: The E3 ubiquitin ligase function of the epigenetic reader PHF16 is crucial for injury-induced intestinal regeneration

city: Pohang

first: Somi

address: 47, Jigok-ro

department: Life Sciences

institute: POSTECH

summary: The intestine has a remarkable capacity to regenerate during homeostasis by the vigorous intestinal stem cells (ISCs) that reside in the base of the crypts. ISCs are easily impaired by DNA damage due to their active cell cycle. Intriguingly, certain cell populations possess the dedifferentiating ability to restore ISCs after ablation, although the molecular mechanisms by which these cells dedifferentiate into ISCs are largely unknown. Here, we performed single-cell RNA-seq using wild type and Phf16 knockout mice from normal or after irradiation to elucidate molecular mechanisms of Phf16 in intestinal regeneration. Our work characterizes that Phf16 is crucial for downregulating stemness in revival stem cells.

overall_design: To verify the function of PHF16 in intestinal regeneration, we generated PHF16 knockout mice using CRISPR-Cas9 system. We then performed single-cell RNA-seq analysis on crypt epithelial cells of WT and PHF16 knockout mice at three different time points after whole-body irradiation.

first: Jun-Yeong

last: Ahn

first: Sung Hee

last: Baek

first: Jong Kyoung


title: Comparison of the response of Col-0 and ate1 ate2 N-degron mutant to flg22

city: Maynooth

first: Emmanuelle

last: Graciet

state: Kildare

address: Biology Department

department: Biology

country: Ireland

institute: Maynooth University

summary: Arabidopsis mutants of the N-degron pathway have been shown to be either susceptible or resistant to a wide range of pathogens. This includes mutants for the arginine-transferases (ate1-2 ate2-1) and for the downstream E3 ubiquitin ligase PRT6 (prt6). In addition, ate1-2 ate2-1 mutants have been shown to have dampened gene expression changes upon inoculation with Pseudomonas syringae AvrRpm1. Here, we tested whether ate1-2 ate2-1 mutants were impaired for pattern-triggered immunity (PTI) and determined gene expression changes of wild-type Col-0 and of ate1-2 ate2-1 mutant seedlings in the absence/pressence of flg22.

overall_design: To investigate transcriptional differences between 10-day old seedlings of Col-0 (wild type) and the ate1-2 ate2-1 N-degron mutant upon 1 hour treatment with 1 uM flg22 (or mock control).  3 biological replicates per sample.

first: Brian

last: Mooney

first: Catherine

middle: M

last: Doorly

first: Melissa

last: Mantz

first: Pablo

last: García

first: Pitter

last: Huesgen


title: Analysis of the transcriptional effect of hypoxia on flg22 response

summary: We tested whether hypoxia affects plant transcriptional responses to flg22 and reveal that hypoxia represses flg22 responses and more broadly also pattern triggered immunity (PTI) in plants.

overall_design: To investigate the transcriptional differences upon combined hypoxia/flg22 treatment, compared to mock control, hypoxia alone or flg22 alone treatments.


title: RNAseq of periosteal stem/progenitor cells from patients with congenital pseudarthrosis of the tibia

city: Créteil

first: Céline

last: Colnot

address: 8 rue du Général Sarrail

institute: INSERM U955

summary: Congenital pseudarthrosis of the tibia (CPT) is a severe pediatric disorder affecting children. CPT is characterized by tibial bowing at birth leading to spontaneous fracture and fibrous non-union. Our study showed the presence of biallelic inactivation of the NF1 gene in the periosteum and in periosteal skeletal/stem progenitor cells (pSSPCs) at the pseudarthrosis site.

overall_design: To understand the impact of NF1 mutation on pSSPCs, we performed RNAseq analysis of primary SSPCs from the pathological periosteum of the pseudarthrosis site and from the non-pathological periosteum collected from the iliac crest of 2 CPT patients.

first: Simon

last: Perrin