title: Single cell RNA-seq of immune cells in multiple mouse models of outer retinal degeneration

summary: The goal of this study focues on comparing the molecular signatures of immune cells, primarily microglia, in ectopically distinct mouse models of outer retinal degeneration. These models include the sodium iodate induced RPE injury model, rhodopsin-P23H knockin mice, an advanced aging model of 2-year-old mice and young (2-month old) wild-type control mice.

overall_design: Profiling of retinal immune cells in mouse models of outer retinal degeneration


title: Threonine fuels glioblastoma through YRDC-mediated codon-biased translational reprogramming (tRNA-Seq)

status: Public on Jan 06 2024

city: Pittsburgh

first: Xujia

last: Wu

state: Pennsylvania

address: 5117 Centre Avenue

department: Hillman Cancer Center

country: USA

institute: University of Pittsburgh

type: Non-coding RNA profiling by high throughput sequencing

summary: To understand the underlying mechanism of YRDC, an enzyme catalysing the formation of N6-threonylcarbamoyladenosine (t6A) on ANN-decoding tRNAs, and the interplay between threonine and YRDC in glioblastoma stem cells (GSCs), we carried out comparative transcriptomic analysis in patient-derived GSC456 with or without YRDC deprivation and with or without threonine restriction (control media, 800 μM threonine; threonine-restricted media, 4 μM). To further investigate the translational regulation of YRDC and threonine in GSCs, we performed ribosome profiling in GSC456 with or without YRDC deprivation and with or without threonine restriction (control media, 800 μM threonine; threonine-restricted media, 4 μM). To investigate the effects of YRDC and threonine purtubation on tRNA expression in GSCs, we performed tRNA sequencing in GSC456 with or without YRDC deprivation and with or without threonine restriction (control media, 800 μM threonine; threonine-restricted media, 4 μM).

overall_design: Comparative gene expression profiling analysis of RNA-seq data, tRNA-seq data and comparative ribosome footprint analysis of ribosome profiling data for GSC456 with or without YRDC deprivation and with or without threonine restriction.

first: Jeremy

middle: N

last: Rich


title: Threonine fuels glioblastoma through YRDC-mediated codon-biased translational reprogramming (Ribosome profiling)

type: Expression profiling by high throughput sequencing

summary: To understand the underlying mechanism of YRDC, an enzyme catalysing the formation of N6-threonylcarbamoyladenosine (t6A) on ANN-decoding tRNAs, and the interplay between threonine and YRDC in glioblastoma stem cells (GSCs), we carried out comparative transcriptomic analysis in patient-derived GSC456 with or without YRDC deprivation and with or without threonine restriction (control media, 800 μM threonine; threonine-restricted media, 4 μM). To further investigate the translational regulation of YRDC and threonine in GSCs, we performed ribosome profiling in GSC456 with or without YRDC deprivation and with or without threonine restriction (control media, 800 μM threonine; threonine-restricted media, 4 μM).

overall_design: Comparative gene expression profiling analysis of RNA-seq data and comparative ribosome footprint analysis of ribosome profiling data for GSC456 with or without YRDC deprivation  and with or without threonine restriction.


title: Threonine fuels glioblastoma through YRDC-mediated codon-biased translational reprogramming

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

overall_design: Refer to individual Series


title: Simultaneous nanopore profiling of mRNA m6A and pseudouridine reveals translation coordination

city: Chicago

first: Tao

last: Pan

state: Illinois

address: 929 E. 57th Street

department: Biochemistry and Molecular Biology

institute: University of Chicago

summary: N6-methyladenosine (m6A) and pseudouridine (Ψ) are the two most abundant modifications in mammalian mRNA, but the coordination of their biological functions remains poorly understood. We develop a machine learning-based nanopore direct RNA sequencing method (NanoSPA) that simultaneously analyzes m6A and Ψ in the human transcriptome. Applying NanoSPA to polysome profiling, we reveal opposing transcriptomic co-occurrence of m6A and Ψ and synergistic, hierarchical effects of m6A and Ψ on the polysome.

overall_design: Comparative gene expression, m6A and psU analysis on input and polysome fractions of siCtrl, siMETTL3 and siTRUB1 samples

first: Sihao

last: Huang

first: Adam

middle: C

last: Wylder


title: COPD basal cells are primed towards secretory to multi-ciliated cell imbalance driving increased resilience to environmental stressors

city: Neuherberg

first: Johannes

last: Beckers

address: Ingolstaedter Landstr. 1

department: Institute of Experimental Genetics

country: Germany

institute: Helmholtz Zentrum Muenchen

summary: Introduction: Environmental pollutants irritate and injure the bronchial elevator, thereby provoking disease progression in chronic obstructive pulmonary disease (COPD). Epithelial resilience mechanisms to environmental nanoparticles in health and disease are poorly characterized.   Methods: We delineated the impact of prevalent pollutants such as carbon and zinc oxide nanoparticles, on cellular function and progeny in primary human bronchial epithelial cells (pHBEC) from end-stage COPD, early disease and pulmonary healthy individuals. After nanoparticle exposure of pHBECs at the air liquid interface, cell cultures were characterized by  functional assays as well as transcriptome and protein analysis, complemented by single cell analysis in serial samples of  pHBEC culture focussing on basal cell differentiation.    Results: In end-stage COPD, environmentally abundant doses of zinc oxide nanoparticles (ZnO) aggravated a pro-secretory phenotype at the expense of the multi-ciliated epithelium alongside a reduction of barrier integrity and increased resilience towards cell damage. Similar effects on cellular composition and function were induced by co-treatment of early stage COPD pHEBC cultures with cigarette smoke extract. Time-resolved single cell transcriptomics revealed a unique end stage COPD associated basal cell state characterized by altered Wnt and Notch pathway activation. A conserved set of COPD specific genes persisted from this disease associated basal cell state into its differentiated progeny.  Conclusion: We identified COPD stage specific gene program alterations in basal stem cells that affect the cellular composition of the bronchial elevator and may control epithelial resilience mechanisms in response to environmental nanoparticles. The identified phenomena likely inform treatment or prevention strategies.

overall_design: We performed gene expression microarray analysis in primary human bronchial epithelial cells from patients with COPD-IV and non-CLD (chronic lung disease) controls. Cells were treated with Printex 90, zinc oxide nanoparticles, LPS, or sham.

first: Mircea

middle: G

last: Stoleriu

first: M

last: Ansari

last: Strunz

first: A

last: Schamberger

last: Heydarian

first: Y

last: Ding

first: C

last: Voss

first: J

last: J Schneider

last: Gerckens

first: Gerald

last: Burgstaller

last: Castelblanco

first: T

last: Kauke

last: Fertmann

last: Schneider

last: Behr

first: Martin

last: Irmler

first: Oliver

last: Eickelberg

first: B

last: Schubert

first: S

last: M Hauck

first: O

last: Schmid

first: R

last: Hatz

last: Stoeger

first: H

last: B Schiller

last: Hilgendorff


title: Inhibition of METTL3 Results in a Cell-Intrinsic Interferon Response That Enhances Antitumor Immunity [ONT direct RNA-seq]

city: Cambridge

first: Byron

last: Andrews

state: Choose County

address: Storm Therapeutics, Moneta Building, Babraham Research Campus

country: United Kingdom

institute: Storm Therapeutics

summary: Therapies that enhance antitumor immunity have altered the natural history of many cancers. Consequently, leveraging nonoverlapping mechanisms to increase immunogenicity of cancer cells remains a priority. Using a novel enzymatic inhibitor of the RNA methyltransferase METTL3, we demonstrate a global decrease in N6-methyladenosine (m6A) results in double-stranded RNA (dsRNA) formation and a profound cell-intrinsic interferon response. Through unbiased CRISPR screens, we establish dsRNA-sensing and interferon signaling are primary mediators that potentiate T-cell killing of cancer cells following METTL3 inhibition. We show in a range of immunocompetent mouse models that although METTL3 inhibition is equally efficacious to anti–PD-1 therapy, the combination has far greater preclinical activity. Using SPLINTR barcoding, we demonstrate that anti–PD-1 therapy and METTL3 inhibition target distinct malignant clones, and the combination of these therapies overcomes clones insensitive to the single agents. These data provide the molecular and preclinical rationale for employing METTL3 inhibitors to promote antitumor immunity in the clinic.

overall_design:
Nanopore sequencing was undertaken on AT3 cells treated with DMSO, STM2457 20µM or STM3006 2µM for 48 hours. All samples were processed in duplicate. Total RNA was isolated using TRIzol LS reagent (Invitrogen) according to manufacturer’s instructions and ethanol precipitation. RNA concentration was quantified with a Qubit Fluorometer (Thermo Fisher Scientific). Extracted total RNA was processed to obtain polyA-tailed transcripts with Dynabeads (Invitrogen) according to the manufacturer's instructions. A target amount of 200ng was used to create a readable transcript library for a single flowcell (type FLO-MIN106) using a protocol based on Oxford Nanopore Technology's DirectRNA SQK-RNA002 kit protocol. Each sample was sequenced on a single flowcell with version R9.4.1 chemistry.
A GridION instrument controlled by MINKNOW software (v20.10.6) was loaded with four flowcells and four channel libraries were run separately and simultaneously. Readfish (v3.0.0) selective sequencing32 was used during the GridION run to eject abundant and irrelevant transcripts. Briefly, the accessions of 81 mouse transcripts comprising mitochondrial transcripts and cytoplasmic ribsomal protein-encoding transcripts were used to generate an exclude list. Reads were basecalled in real time using Guppy (v4.2.3). FAST5 files were recorded for later processing by Nanopolish (v0.11.2)33. Basecalled reads and Fast5 signal files were processed to prepare for Nanocompore (v1.0.3) comparison analysis according to the author's instructions17. Briefly, fastq files were aligned to a mouse transcriptome version M20 with minimap2 (v2.2.0)34 to obtain bam alignment files, then Nanopolish's eventalign module was used to estimate kmer pore transition ('dwell') times from the alignment and the Fast5 signal files. The Nanopolish-compress tool was used to average nanopolish output by kmer. Nanocompore was used to discern significant differences in signal current intensity and dwell time between the control and treatment replicate pairs, using the GMM-logit option to use Gaussian Mixture Modelling to discern the presence of two different nucleotide modification states, and logit regression analysis to calculate the difference in site characteristics between the samples.

first: Andrew

middle: A

last: Guirguis

first: Yaara

last: Ofir-Rosenfeld

first: Kathy

last: Knezevic

first: Wesley

last: Blackaby

first: David

last: Hardick

first: Yih-Chih

last: Chan

first: Ali

last: Motazedian

first: Andrea

last: Gillespie

first: Dane

last: Vassiliadis

first: Enid

middle: Y

last: Lam

first: Kevin

last: Tran

first: Michael

middle: E

last: Harbour

first: Lina

last: Vasiliauskaite

first: Claire

middle: J

last: Saunders

first: Georgia

last: Tsagkogeorga

first: Aleksandra

last: Azevedo

first: Joanna

last: Obacz

first: Ewa

middle: S

last: Pilka

first: Marie

last: Carkill

first: Laura

last: MacPherson

first: Elanor

last: Wainwright

first: Brian

last: Liddicoat

first: Benjamin

last: Blyth

first: Mark

middle: R

last: Albertella

last: Rausch

last: Dawson


title: Basal epithelial subpopulations underlie and predict chemotherapy resistance in Triple-Negative Breast Cancer

status: Public on Jan 05 2024

city: Odense M

first: Vijay

last: Tiwari

state: Denmark

address: Campusvej 55

department: Institut for Molekylær Medicin (IMM)

country: Denmark

institute: Queen’s University Belfast

summary: Triple-Negative Breast Cancer (TNBC) is the most aggressive breast cancer subtype, characterized by extensive intratumoral heterogeneity, high metastasis and chemoresistance, leading to poor clinical outcomes. Despite progress, the mechanistic basis of these aggressive behaviours remains poorly understood. Using single-cell and spatial transcriptome analysis, here we discovered basal epithelial subpopulations located within the stroma that exhibit chemoresistance characteristics. The subpopulations are defined by distinct signature genes that show a frequent gain in copy number and exhibit an activated epithelial-to-mesenchymal transition program. A subset of these genes can accurately predict chemotherapy response and are associated with poor prognosis. Interestingly, among these genes, elevated ITGB1 participates in enhancing intercellular signaling while ACTN1 confers a survival advantage to foster chemoresistance. Furthermore, by subjecting the transcriptional signatures to drug repurposing analysis we find that chemoresistant tumors may benefit from distinct inhibitors in treatment naïve versus post-NAC patients. These findings shed light on the mechanistic basis of chemoresistance while providing the best-in-class biomarker to predict chemotherapy response and alternate therapeutic avenues for improved management of TNBC patients resistant to chemotherapy.

overall_design: To investigate transcriptional profiule dynamics during epithelial to mesenchymal transition (EMT). HMLE cells treated with TGFB at Untreated, day1, day4, day7, day10, day15, day20

first: Mohammed

last: Inayatullah


title: Threonine fuels glioblastoma through YRDC-mediated codon-biased translational reprogramming (RNA-Seq)

first: Xujia

last: Wu

first: Jeremy

last: Rich


title: Memory consolidation in honey bees is enhanced by down-regulation of Down Syndrome Cell Adhesion Molecule and changes its alternative splicing

city: Birmingham

first: David

middle: William John

last: McQuarrie

address: School of Biosciences

institute: University of Birmingham

summary: Down syndrome cell adhesion molecule (Dscam) gene encodes a cell adhesion molecule required for neuronal wiring. A remarkable feature of invertebrate Dscam is massive alternative splicing generating thousands of different isoforms from three variable clusters of alternative exons. Dscam expression and diversity arising from alternative splicing have been studied during development, but whether they exert functions in differentiated brains has not been determined. Here, using honey bees, we find that Dscam expression is critically linked to memory retention as reducing expression by RNAi enhances memory after reward learning in adult worker bees. Moreover, alternative splicing of Dscam is altered in all three variable clusters after learning. Since identical Dscam isoforms engage in homophilic interactions, these results suggest a mechanism to alter inclusion of variable exons during memory consolidation possibly to alter neuronal connections for memory retention.

overall_design: To investigate the role of Dscam alternative exon splicing in learning and memory in Apis mellifera central brains using RNA-seq.

first: Roland

last: Arnold

first: Matthias

last: Soller


title: Methylation analysis in monozygotic(MZ) twins with differences in TSH

status: Public on Jan 04 2024

city: Yamadaoka,Suitashi

first: Yuya

last: Arakawa

state: Osaka

department: Graduate school of Medicine

country: Japan

institute: Osaka university

summary: Genome wide DNA methylation profiling of MZ twins whose within-pair difference in TSH were discordant. the Illumina Infinium HumanMethylation450 Bead Chip Kit (approximately 480,000 CpG sites) and the Illumina Infinium Methylation EPIC Bead Chip (approximately 900,000 CpG sites) were used to obtain DNA methylation profiles in peripheral blood samples.

overall_design: Bisulphite converted DNA from the 44 samples (22 pairs) were hybridised to the Illumina Infinium HumanMethylation450 Bead Chip Kit or the Illumina Infinium Methylation EPIC Bead Chip.

first: Mikio

last: Watanabe


title: Targeting postsynaptic glutamate receptor scaffolding proteins PSD-95 and PICK1 for obesity treatment [2]

city: Copenhagen

first: Lars

middle: Roed

last: Ingerslev

address: Blegdamsvej 3B

department: NNF Center for Basic Metabolic Research

institute: Copenhagen University

summary: Human genome-wide association studies (GWAS) suggest a functional role for central glutamate receptor signaling and plasticity in body weight regulation. Here, we utilize UK Biobank GWAS summary  statistics of body mass index (BMI) and body fat percentage (BF%) to identify genes encoding for proteins known to interact with postsynaptic AMPA and NMDA receptors. Loci in/near Discs Large Homolog 4 (DLG4) and protein interacting with C kinase 1 (PICK1) reached genome-wide significance (P<5x10-8) for BF% and/or BMI. To further evaluate the functional role of PSD-95 (gene name: DLG4) and PICK1 in energy homeostasis, we used dimeric PDZ-domain-targeting peptides of PSD-95 and PICK1 to demonstrate that pharmacological inhibition of PSD-95 and PICK1 induces prolonged weight-lowering effects in obese mice. Collectively, these data demonstrate that the glutamate receptor scaffolding proteins, PICK1 and PSD-95, are genetically linked to obesity and that pharmacological targeting of their PDZ domains represents a promising new therapeutic avenue for sustained weight loss.

overall_design: Comparative gene expression profiling analysis of RNA-seq data from male mouse hypothalamus treated with saline or inhibitors of PSD-95 (UCCB01-147 ) or PICK1 (mPD5)

first: Nicole

last: Fadahunsi

first: Jonas

last: Petersen

first: Sophia

last: Metz

first: Alexander

last: Jakobsen

first: Cecilie

middle: V

last: Mathiesen

first: Alberte

last: Buch-Rasmussen

first: Nigel

middle: K

last: Kurgan

first: Jeppe

last: Larsen

first: Rita

last: Andersen

first: Thomas

last: Topilko

first: Charlotte

last: Svendsen

first: Mia

last: Apuschkin

first: Grethe

last: Skovbjerg

first: Jan

middle: H

last: Schmidt

first: Grace

last: Houser

first: Sara

last: Jager

first: Anders

last: Bach

first: Atul

last: Deshmukh

first: Tuomas

middle: O

last: Kilpeläinen

first: Kristian

last: Strømgaard

first: Kenneth

middle: L

last: Madsen

first: Christoffer

last: Clemmensen


title: Central Inhibition of HDAC6 Re-Sensitizes Leptin Signaling During Obesity to Induce Profound Weight Loss

status: Public on Jan 09 2024

city: Boston

first: dongxian

last: guan

state: MA

address: Children's Hospital 300 Longwood Ave

institute: Boston Children's Hospital

summary: Leptin resistance during excess weigh gain significantly contributes to the recidivism of obesity to leptin-based pharmacological therapies. The mechanisms underlying the inhibition of Leptin receptor b (LepR) signaling during obesity is still elusive. Here we report that histone deactylase 6 (HDAC6) interacts with LepR, reducing the latter’s activity, and that pharmacological inhibition of HDAC6 activity disrupts this interaction and augments leptin signaling. Treatment of obese mice with blood brain barrier (BBB)-permeable HDAC6 inhibitors profoundly reduces food intake and leads to a potent weight loss without affecting the muscle mass. Genetic depletion of Hdac6 in AgRP-expressing neurons or administration with BBB-impermeable HDAC6 inhibitors result in a lack of such anti-obesity effect. Together, these findings represent the first report describing a mechanistically validated and pharmaceutically tractable therapeutic approach to directly increase LepR activity  as well as identifying centrally-, but not peripherally-acting HDAC6 inhibitors as potent leptin-sensitizers and anti-obesity agents.

overall_design: To investigate the leptin sensitizer celastrol-regulated orexigenic or anorexigenic genes in the hypothalamus, we decided that a pair-fed control should be included, in which the food intake of pair-fed mice match those treated with celastrol, along with satiated control mice, when analyzing celastrol-treated mice to identify the genes involved in leptin resistance. DIO mice were acclimated with DMSO with daily i.p. injections for 1 week and then randomly divided into 4 groups for a subsequent 4-day treatment. 1) Vehicle (Veh), treated with DMSO and ad libitum access to food, 2) CS, treated with Celastrol (100μg/kg/day, i.p.) and ad libitum access to food, 3) PF, pair-fed to Celastrol-treated group, in which 0.5g high fat diet per mice per day was given (0.34g for dark cycle and 0.16g for light cycle) (a little bit less than 0.8g, the average amount of diet Celastrol-treated mice consumed), 4) PF/CS, treated with Celastrol and given the same amount of food and at the same intervals as the PF group. On the fourth day around 8:00 am, all mice received a final injection and 6 hours later blood glucose (mg/dl) levels were measured before tissue collection.

first: Dongxian

last: Guan

first: Umut

last: ozcan


title: Comparative genomics incorporating translocation renal cell carcinoma mouse model reveals molecular mechanisms of tumorigenesis

city: Dallas

first: James

last: Brugarolas

state: Texas

address: 5323 Harry Hines Blvd.

department: Department of Internal Medicine

institute: University of Texas Southwestern Medical Center

summary: Translocation renal cell carcinoma (tRCC) most commonly involves an ASPSCR1-TFE3 fusion, but molecular mechanisms remain elusive and animal models are lacking. Here, we show that human ASPSCR1-TFE3 driven by Pax8-Cre (a credentialed clear cell RCC driver) disrupted nephrogenesis and glomerular development, causing neonatal death, while the clear cell RCC failed driver, Sglt2-Cre, induced aggressive tRCC (as well as alveolar soft part sarcoma) with complete penetrance and short latency. However, in both contexts, ASPSCR1-TFE3 led to characteristic morphological cellular changes, loss of epithelial markers, and an epithelial-mesenchymal transition. Electron microscopy of tRCC tumors showed lysosome expansion, and functional studies revealed simultaneous activation of autophagy and mTORC1 pathways. Comparative genomic analyses encompassing an institutional human tRCC cohort (including a hitherto unreported SFPQ-TFEB fusion) and a variety of tumorgraft models (ASPSCR1-TFE3, PRCC-TFE3, SFPQ-TFE3, RBM10-TFE3, and MALAT1-TFEB) disclosed significant convergence in canonical pathways (cell cycle, lysosome, and mTORC1) and less established pathways such as Myc, E2F, and inflammation (IL-6/JAK/STAT3, interferon-γ, TLR signaling, systemic lupus, etc.). Therapeutic trials (adjusted for human drug exposures) showed antitumor activity of cabozantinib. Overall, this study provides insight into MiT/TFE-driven tumorigenesis, including the cell of origin, and characterizes diverse mouse models available for research

overall_design: Samples were submitted to the New York Genome Center (NYGC) for nucleic acid extraction as well as RNA-sequencing (RNA-Seq). 6 Sglt2-Cre; ASPSCR1-TFE3 LSL/+ mice were used for RNA-Seq studies encompassing 22 samples: 11 kidney tumors; 4 retro-orbital tumors; 3 brain tumors as well as 4 non-transformed kidney samples. In addition, RNA isolation and RNA-Seq were performed on 6 normal kidney samples from 3 Sglt2-Cre control mice. Kidney samples from 5 Pax8-Cre; ASPSCR1-TFE3 LSL/+ fetus and 3 Pax8-Cre or ASPSCR1-TFE3 LSL/+ age-matched controls were submitted for RNA-Seq. All samples were from formalin-fixed and paraffin-embedded (FFPE) tissues.

first: Alana

last: Christie

first: Gopinath

last: Prakasam

first: Akhilesh

last: Mishra


title: Organogenetic transcriptomes of the Drosophila embryo at single cell resolution

city: Baltimore

first: Da

last: Peng

state: Maryland

address: 733 North Broadway, MRB 653

institute: Johns Hopkins University

summary: To gain insight into the transcription programs activated during organogenesis of Drosophila larval structures, we carried out single cell RNA sequencing during two periods of Drosophila embryogenesis: stages 10 – 12, when most organs are first specified and initiate morphological and physiological specialization, and stages 13 – 16, when organs have achieved their final mature architectures and are beginning to function.

overall_design: Two sets of biological replicates for stage 10-12 wild type (Oregon R) Drosophila melanogaster embryos and two sets of biological replicates for stage 13-16 wild type (Oregon R) Drosophila melanogaster embryos were collected, dissociated into single cells, and ran through 10X scRNA-seq pipeline.

first: Dorian

last: Jackson

first: Bianca

last: Palicha

first: Eric

last: Kernfeld

first: Nathaniel

last: Laughner

first: Ashleigh

last: Shoemaker

first: Rajprasad

last: Loganathan

first: Patrick

last: Cahan

first: Deborah

last: Andrew


title: Maternal inherited siRNA initiate piRNA cluster formation

city: Ithaca

first: Yicheng

last: Luo

state: NY

address: 215 Tower Rd

department: Molecular Biology and Genetics

institute: Cornell University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: In order to control transposable element (TE) activity, PIWI-interacting RNAs (piRNAs) have been evolved to silence TE transcriptionally and post-transcriptionally, and produced from heterochromatic genomic loci, called piRNA cluster. Maternal inherited piRNAs transmission is considered as the key step of piRNA cluster maintenance and induction of de nove piRNA cluster formation, however, how the original piRNAs were produced without maternal piRNAs deposition remains unclear. In this paper, we created a repetitive GFP reporter and found a Rhino-dependent piRNA cluster conversion spontaneously occurs to the reporter over generations. Importantly, this conversion is reversible by removal of maternal piRNAs of GFP reporter. Taking advantage of this reversible GFP reporter piRNA cluster, we found that siRNA is able to initiate piRNA cluster formation epigenetically, even when only maternal siRNA presence is capable to do so. Our results suggest the importance of siRNA on initiation of piRNA biogenesis and describe a whole picture of initiation and maintenance of piRNA cluster.

overall_design:
Genome occupancy profiling and piRNA production by high throughput sequencing

The submitter noted that
[1] the following processed data files are identical as they contain very few values:

dsGPP_66A6_oxidized_piRNA_1st_S19_R1_001_dsGPP_66A6_oxidized_piRNA_1st_S19_R1_001.dm6.21_22mer.unique.dup.bam.10.mir304_12.Minus.bg4.chopped.bg4
dsGPP_66A6_oxidized_piRNA_2nd_S19_R1_001_dsGPP_66A6_oxidized_piRNA_2nd_S19_R1_001.dm6.21_22mer.unique.dup.bam.10.mir304_12.Minus.bg4.chopped.bg4

dsGPP_66A6_oxidized_piRNA_1st_S19_R1_001_dsGPP_66A6_oxidized_piRNA_1st_S19_R1_001.dm6.21_22mer.unique.dup.bam.10.mir304_12.Plus.bg4.chopped.bg4
dsGPP_66A6_oxidized_piRNA_2nd_S19_R1_001_dsGPP_66A6_oxidized_piRNA_2nd_S19_R1_001.dm6.21_22mer.unique.dup.bam.10.mir304_12.Plus.bg4.chopped.bg4

[2] some samples do not have processed data as only BAM/FASTA files were generated.

first: Peng

last: He

first: Nivedita

last: Kanrar

first: Alexei

last: Aravin


title: Methylation analysis in monozygotic(MZ) twins with differences in FT4

first: Yuya

last: Arakawa

summary: Genome wide DNA methylation profiling of MZ twins whose within-pair difference in FT4 were discordant. the Illumina Infinium HumanMethylation450 Bead Chip Kit (approximately 480,000 CpG sites) and the Illumina Infinium Methylation EPIC Bead Chip (approximately 900,000 CpG sites) were used to obtain DNA methylation profiles in peripheral blood samples.

first: Mikio

last: Watanabe


title: Locus folding mechanisms determine modes of antigen receptor gene assembly (RNA-seq)

city: Philadelphia

first: Craig

last: Bassing

address: 3501 Civic Center Blvd

institute: Children's Hospital of Philadelphia

summary: The dynamic folding of genomes regulates numerous biological processes, including antigen receptor (AgR) gene assembly. We show that, unlike other AgR loci, homotypic chromatin interactions and bi-directional chromosome looping both contribute to structuring Tcrb for efficient long-range V(D)J recombination. Inactivation of the CTCF binding element (CBE) or promoter at the most 5’Vβ segment (Trbv1) impaired loop extrusion originating locally and extending to DβJβ CBEs at the opposite end of Tcrb. Promoter or CBE mutation nearly eliminated Trbv1 contacts and decreased RAG endonuclease-mediated Trbv1 recombination. Importantly, Trbv1 rearrangement can proceed independent of substrate orientation, ruling out scanning by DβJβ-bound RAG as the sole mechanism of Vβ recombination, distinguishing it from Igh. Our data indicate that CBE-dependent generation of loops cooperates with promoter-mediated activation of chromatin to juxtapose Vβ and DβJβ segments for recombination through diffusion-based synapsis. Thus, the mechanisms that fold a genomic region can influence molecular processes occurring in that space, which may include recombination, repair, and transcriptional programming.

overall_design: RNA-Seq was performed on sorted (B220-, CD19-, CD11b-, CD11c-, NK1.1-, TER119-, TCRβ-, TCRγ/δ-) DN thymocytes from two Rag1-/- mice per experiment with either a WT Tcrb locus or with the indicated mutations.

first: Brittney

last: Allyn


title: Characterization of CD4+ and CD8+ T cells responses in the Mixed Lymphocyte Reaction by flow cytometry and single cell RNA sequencing

city: Gif-sur-Yvette

first: Adèle

last: Mangelinck

state: Essonne

address: 22 route 128 - rue Francis Perrin

country: France

institute: Servier

summary: The Mixed Lymphocyte Reaction (MLR) consists in the allogenic co-culture of monocytes derived dendritic cells (MoDCs) with T cells from another donor. This in vitro assay is largely used for the assessment of immunotherapy compounds. Nevertheless, the phenotypic changes associated with lymphocyte responsiveness under MLR have never been thoroughly evaluated. Here, we used multiplex cytokine and chemokine assays, multiparametric flow cytometry and single cell RNA-sequencing to deeply characterize T cells activation and function in the context of CD4+- and CD8+-specific MLR kinetics. We showed that CD4+ and CD8+ T cells in MLR share common classical markers of response such as polyfunctionality, increased proliferation and CD25 expression but differ in their kinetics and amplitude of activation as well as their patterns of cytokines secretion and immune checkpoints expression. The analysis of immunoreactive KI67+CD25+ T cells identified PBK, LRR1 and MYO1G as new potential biomarkers of MLR response. Using cell-cell communication network inference and pathway analysis on single cell RNA-sequencing data, we also highlighted key components of the immunological synapse occurring between T cells and the stimulatory MoDCs together with downstream signaling pathways involved in CD4+ and CD8+ T cells activation. These results provide a deep understanding of the kinetics of the MLR assay for CD4+ or CD8+ T cells and may allow to identify new strategies for immunotherapy in cancer.

overall_design:
Gene expression profiling analysis of single cell RNA-seq data from MLR and paired T cells samples at different timepoints (day 1, 2, 5 and 6).
NOTE FROM SUBMITTER: the data consists in patient data so we cannot share sequencing reads due to privacy issues.


title: Locus folding mechanisms determine modes of antigen receptor gene assembly


title: Locus folding mechanisms determine modes of antigen receptor gene assembly (ChIP-seq)

overall_design: ChIP Seq was performed in DN thymocytes from ~5 Rag1-/- mice per experiment with either a WT Tcrb locus or with the indicated mutations with the indicated antibodies.


title: Locus folding mechanisms determine modes of antigen receptor gene assembly (Hi-C)

overall_design: Conventional HiC or HiC using the Arima Kit was performed in DN thymocytes from ~5 Rag1-/- mice per experiment with either a WT Tcrb locus or with the indicated mutations.


title: Transcriptomic analysis of neuroinflammation and occult infection in sudden infant death syndrome

first: Benjamin

last: Okaty

address: 77 Avenue Louis Pasteur

institute: Harvard Medical School

summary: Antemortem infection is a risk factor for sudden infant death syndrome (SIDS) – the leading postneonatal cause of infant mortality in the developed world. Manifestations of infection and attendant inflammation, however, are not always apparent in clinical settings or by standard autopsy, thus enhanced resolution approaches are needed. Here we screened postmortem SIDS tissues and fluids for inflammatory markers and applied metagenomics and transcriptomics to a subset of cases to look for evidence of occult infection and inflammation.

overall_design: Postmortem fluids from a cohort of SIDS and controls collected between 2012 and 2018 were screened for elevated inflammatory markers, specifically cerebrospinal fluid (CSF) neopterin and CSF and serum cytokines. Postmortem brain and liver tissue and CSF from 6 SIDS cases with high levels of neopterin ("inflammatory SIDS"), as well as 3 non-inflammatory SIDS cases were subjected to metagenomic next generation sequencing (mNGS; see BioProject PRJNA1021781 for data). Human parechovirus 3 (HPeV3) was detected in all tissues profiled of a single case (HPeV3+ SIDS), but no pathogens were detected in the other cases or controls. Brainstem tissue (at the level of obex+2mm) from the HPeV3+ SIDS case and 3 non-inflammatory SIDS cases was analyzed by single nucleus RNA sequencing (snRNAseq) to measure cell type-specific gene expression associated with neuroinflammation and infection (the data presented in this submission)


title: CHCHD4-TRIAP1 regulation of innate immune signaling mediates skeletal muscle adaptation to exercise

city: Bethesda

first: Paul

middle: M.

last: Hwang

address: 9000 Rockville Pike

institute: NHLBI

summary: Exercise training can stimulate the formation of fatty acid oxidizing slow-twitch skeletal muscle fibers which are inversely correlated with obesity, but the molecular mechanism underlying this transformation requires further elucidation.  Here, we report that the downregulation of the mitochondrial disulfide relay carrier CHCHD4 in skeletal muscle by exercise training decreases the import of TP53-regulated inhibitor of apoptosis 1 (TRIAP1) into mitochondria, which reduces cardiolipin levels and promotes VDAC oligomerization.  The subsequent release of mtDNA into the cytosol activates innate immune signaling through cGAS and NFKB, downregulating MyoD and promoting the formation of oxidative slow-twitch fibers.  In mice, CHCHD4 haploinsufficiency was sufficient to activate this pathway, leading to increased oxidative muscle fibers and decreased fat accumulation with aging.  The identification of a specific mediator regulating muscle fiber transformation provides an opportunity to understand further the molecular underpinnings of complex metabolic conditions such as obesity and could have therapeutic implications.

overall_design: To obtain a comprehensive view of the molecular changes associated with CHCHD4 and exercise training, skeletal muscles derived from both CHCHD4 Tg and wild-type mice, with or without exercise training, were subjected to RNA seq anyalsis

first: Jin

last: Ma

first: Ping-yuan

last: Wang

first: Zhichao

last: Wu

middle: M


title: Engineered mischarged transfer RNAs for correcting pathogenic missense mutations [MSR-seq]

first: Tao

last: Pan

summary: Missense mutations account for nearly 50% of pathogenic mutations in human genetic diseases, most lack effective treatments. Gene therapies, CRISPR-based gene editing, and RNA therapies including transfer RNA (tRNA) modalities are common strategies for potential treatments of genetic diseases. However, reported tRNA therapies are for nonsense mutations, how tRNAs can be engineered to correct missense mutations have not been explored. Here, we describe missense correcting tRNAs (mc-tRNAs) as a potential therapeutic modality for correcting pathogenic missense mutations. Mc-tRNAs are engineered tRNAs that are charged with one amino acid and read codons of another amino acid in translation in human cells. We first developed a series of fluorescence protein (FP)-based reporters that indicate successful correction of missense mutations via restoration of fluorescence signals. We engineered mc-tRNAs that effectively corrected Serine and Arginine missense mutations in the reporters and confirmed the amino acid substitution by protein mass spectrometry and mc-tRNA expression by tRNA sequencing. We examined the transcriptome response to the expression of mc-tRNAs and found some mc-tRNAs induced minimum transcriptomic changes. Furthermore, we applied an Arg-tRNAGln(CUG) mc-tRNA to rescue the autolytic activity of a pathogenic CAPN3 Arg-to-Gln mutant involved in limb-girdle muscular dystrophy type 2A. These results establish a versatile pipeline for mc-tRNA engineering and demonstrate the potential of mc-tRNA as an alternative therapeutic platform for the treatment of genetic disorders.

overall_design: Five different mistranslation reporters (EGFP(mutant)-mCherry fusion proteins) are co-expressed with or without their corresponding missense correcting tRNAs (mc-tRNAs)  in HEK293 cells. We studied the expression and modification levels of the mc-tRNAs, as well as their effects on global tRNA and mRNA expression levels using total RNA from cells expressing mistranslation reporters alone or together with their corresponding mc-tRNAs.

first: Yichen

last: Hou

first: Wen

last: Zhang

first: Philip

last: McGilvray

first: Marek

last: Sobczyk

first: Sihao

last: Huang

first: Noah

last: Pena

first: Chris

last: Katanski


title: Engineered mischarged transfer RNAs for correcting pathogenic missense mutations [mRNA-seq]


title: Tnf signaling deficiency promotes UVB-induced melanocyte stem cell activation and migration

city: ithaca

first: Luye

last: An

address: 618 tower road

department: biomedical sciences

institute: cornell university

summary: This study focuses on understanding the role of Tnf signaling in regulating UVB induced melanocyte stem cell activation and epidermal migration

overall_design: Comparative gene expression profiling analysis of RNA-seq data for whole skin with and without UVB irradiation in WT and TNFR1/2 double KO mice

first: Dahihm

last: Kim

first: William

last: Zhuang

first: Andrew

last: White


title: Cleavage of the pseudoprotease iRhom2 by the signal peptidase complex reveals an ER-to-nucleus signalling pathway

city: Leicester

first: Michael

last: Tellier

state: Select State

address: Lancaster Road

department: Department of Molecular and Cell Biology

institute: University of Leicester

summary: iRhoms are pseudoprotease members of the rhomboid-like superfamily and are cardinal regulators of inflammatory and growth factor signaling; they function primarily by recognizing transmembrane domains of their clients. Here, we report a mechanistically distinct nuclear function of iRhoms, showing that both human and mouse iRhom2 are non-canonical substrates of signal peptidase complex (SPC), the protease that removes signal peptides from secreted proteins. Cleavage of iRhom2 generates an N-terminal fragment that enters the nucleus and modifies the transcriptome, in part by binding C-terminal binding proteins (CtBPs). The biological significance of nuclear iRhom2 is indicated by elevated levels in skin biopsies of patients with psoriasis, tylosis with oesophageal cancer (TOC), and non-epidermolytic palmoplantar keratoderma (NEPPK); increased iRhom2 cleavage in a keratinocyte model of psoriasis; and nuclear iRhom2 promoting proliferation of keratinocytes. Overall, this work identifies an unexpected SPC-dependent ER-to-nucleus signaling pathway and demonstrates that iRhoms can mediate nuclear signaling.

overall_design: Poly(A)+ RNA-seq performed in biological triplicates in HEK293 cells inserted with a doxycycline inducible iRhom2 fragment. RNA-seq were performed at 0h, 3h, and 6h after doxycycline induction.

first: Iqbal

last: Dulloo

first: Clémence

last: Levet

first: Anissa

last: Chikh

first: Boyan

first: Diana

middle: C

last: Blaydon

first: Catherine

last: Webb

first: David

middle: P

last: Kelsell

first: Matthew

last: Freeman


title: The hagfish genome and the evolution of vertebrates

city: London

first: Ferdinand

last: Marlétaz

address: Gower Street

department: Genetics, Evolution & Environment

institute: University College London

summary: As the only surviving lineages of jawless fishes, hagfishes and lampreys provide a critical window into early vertebrate evolution. Here, we investigate the complex history, timing, and functional role of genome-wide duplications in vertebrates in the light of a chromosome-scale genome of the brown hagfish Eptatretus atami. Using robust chromosome-scale (paralogon-based) phylogenetic methods, we confirm the monophyly of cyclostomes, document an auto-tetraploidization (1RV) that predated the origin of crown group vertebrates ~517 Mya, and establish the timing of subsequent independent duplications in the gnathostome and cyclostome lineages. Some 1RV gene duplications can be linked to key vertebrate innovations, suggesting that this early genomewide event contributed to the emergence of pan-vertebrate features such as neural crest. The hagfish karyotype is derived by numerous fusions relative to the ancestral cyclostome arrangement preserved by lampreys. These genomic changes were accompanied by the loss of genes essential for organ systems (eyes, osteoclast) that are absent in hagfish, accounting in part for the simplification of the hagfish body plan; other gene family expansions account for hagfishes’ capacity to produce slime. Finally, we characterise programmed DNA elimination in somatic cells of hagfish, identifying protein-coding and repetitive elements that are deleted during development. As in lampreys, the elimination of these genes provides a mechanism for resolving genetic conflict between soma and germline by repressing germline/pluripotency functions. Reconstruction of the early genomic history of vertebrates provides a framework for further exploration of vertebrate novelties.

overall_design: RNA-seq from total RNA extracted from organs

last: Marletaz

first: Oleg

last: Simakov

first: Dan

last: Rokhsar


title: scGR-seq: simulatenous profiling of glycomic and transcriptomic in single cell [scRNA-Seq]

city: Tsukuba

first: Hiroaki

last: Tateno

state: Ibaraki

address: Tsukuba Central 2, 1-1-1 Umezono

institute: National Institute of Advanced Industrial Science and Technology (AIST)

summary: Previously we have developed integrated analysis of single cell glycome and the transcriptome called scGR-seq using oligonucleotide-labeled lectins. Due to its plate based method, the amount of cell that can be analyzed was limited. Here, adapting 10x Genomic platform we analyzed the integrated model in thousands of cell revealing heterogenicity of cellular glyco-state in cell population.

overall_design: Integrated single cell glycome and transcriptome profiling of Panc04.03, hTERT-HPNE and PBMC

first: Sunanda

last: Keisham

first: Sayoko

last: Saito

first: Satori

last: Kowashi


title: RNA-seq profiling of paralysed CD8 negative dendritic cells after systemic inflammatory syndrome

city: Parkville

first: Gordon

last: Smyth

state: Vic

address: 1G Royal Pde

department: Bioinformatics

country: Australia

institute: Walter and Eliza Hall Institute of Medical Research

summary: CD8 negative dendritic cells were harvested from the spleens of untreated and treated mice. Treated mice were inoculated intravenously with 20 nmol of CpG1680 to induce systematic inflammation five days before harvesting cells. RNA was extracted and profiled by RNA-seq. Three independent experiments were performed for each of treated and untreated groups, with 8 mice per group.

overall_design: RNA-seq profiles were generated of untreated CD8 negative dendritic cells (3 biological replicates) and CpG-pretreated CD8 negative dendritic cell (3 biological replicates).


title: Ultra-low error synthetic long-read single-cell sequencing reveals expressions of hypermutation clusters of isoforms in human liver cancer cells

city: Pittsburgh

first: Shuchang

last: Liu

state: PA

address: 203 Lothrop Street

institute: University of Pittsburgh

summary: The protein diversity of mammalian cells is determined by arrays of isoforms from genes. Protein mutation is essential in species evolution and cancer development. Accurate Long-read transcriptome sequencing at single-cell level is required to decipher the spectrum of protein expressions in mammalian organisms. In this report, we developed a synthetic long-read single-cell sequencing technology based on LOOPseq technique. We applied this technology to analyze 447 transcriptomes of hepatocellular carcinoma (HCC) and benign liver from an individual. Through Uniform Manifold Approximation and Projection (UMAP) analysis, we identified a panel of mutation mRNA isoforms highly specific to HCC cells. The evolution pathways that led to the hyper-mutation clusters in single human leukocyte antigen (HLA) molecules were identified. Novel fusion transcripts were detected. The combination of gene expressions, fusion gene transcripts, and mutation gene expressions significantly improved the classification of liver cancer cells versus benign hepatocytes. In conclusion, LOOPseq single-cell technology may hold promise to provide a new level of precision analysis on the mammalian transcriptome.

overall_design: HCC samples and benign liver samples were freshly dissected from a patient who underwent liver transplantation. Long-read single-cell transcriptome sequencing was performed by LoopSeq, Element Biosciences. Each library (tumor or benign) was sequenced by six runs and eventually pooled together.

first: Silvia

first: Yan-Ping

last: Yu

first: Bao-Guo

last: Ren

first: Tuval

middle: B

last: Yehezkel

first: Carolyn

last: Colbert

first: Wenjia

last: Wang

first: Alina

last: Ostrowska

first: Alejandro

last: Soto-Gutierrez

first: Jian-Hua

last: Luo


title: Transcriptome analysis of cultured human pulmonary artery smooth muscle cells (hPASMCs) to explore functional roles for CD26/DPP4 in stimulation with TGF-β1

status: Public on Jan 03 2024

city: Chiba

first: Takeshi

last: Kawasaki

address: 1-8-1, Inohana, Chuoku

department: Respirology

institute: Chiba University

summary: The purpose of this study was to evaluate the role of CD26/DPP4 in hPASMCs in response to TGF-β1. In experimental methods, hPASMCs were treated as follows. The hPASMCs were cultured in Smooth Muscle Cell Growth Medium 2 with 10% fetal bovine serum and were incubated at 37 ℃ in a 5% CO2 incubator. The cells at passage 6 were used for all experiments. The PASMCs were treated with CD26/DPP4 siRNA or non-specific control siRNA for 48 hours, then stimulated with TGF-β1 or PBS for 24 hours. Transcriptome analysis revealed that TGFβ treatment in cultured hPASMCs upregu-lated genes related to pulmonary vascular SMC proliferation, involving the Notch, PI3K-Akt, and NFκB signaling pathways.Conversely, DPP4-siRNA treatment in cultured hPASMCs downregulated these pathways.

overall_design: Examination of cultured hPASMC treated with  DPP4 knockdown (KD) +/- and  TGF-β1 stimulation +/-

first: Tadasu

last: Okaya

first: Yoshinori

last: Hasegawa

first: Osamu

last: Ohara

first: Koichiro

last: Tatsumi

first: Takuji

last: Suzuki


title: NOTCH2 Promotes Osteoclast Maturation and Metabolism and Modulates the Transcriptome Profile During Osteoclastogenesis

city: Farmington

first: Ernesto

last: Canalis

state: CT

address: 263 Farmington Avenue

department: Orthopaedic Surgery

institute: UConn Health

summary: Bone marrow derived macrophages (BMM) from Notch2tm1.1Ecan, harboring a NOTCH2 gain-of-function mutation, and control mice were cultured with macrophage colony stimulating factor (M-CSF) and receptor activator of NF-kB ligand (RANKL). Bulk RNA-Seq revealed enhanced cell metabolism, aerobic respiration, and mitochondrial function, all associated with osteoclastogenesis, in Notch2tm1.1Ecan cells. Single cell RNA-Seq data of BMMs treated with M-CSF or M-CSF and RANKL for 3 days identified 11 well-defined cellular clusters. There was an increased number of cells expressing gene markers associated with the osteoclast and with related clusters in Notch2tm1.1Ecan than in control BMMs.

overall_design: To obtain BMMs, the bone marrow from 7 to 8-week-old experimental and control sex-matched littermate mice was removed by flushing. Cells were cultured in a-minimum essential medium (a-MEM) in the presence of 10% fetal bovine serum and M-CSF at 30 ng/ml on uncoated plastic petri dishes, grown, collected and seeded on tissue culture plates in the presence of M-CSF and RANKL. For bulk RNA-Seq, cultures were carried out until multinucleated tartrate resistant acid phosphotase (TRAP)-positive cells were formed. For scRNA-Seq, BMMS were seeded on culture plates coated with Cellmatrix Type I-A Collagen, grown in a-MEM in the presence of either M-CSF at 30 ng/ml alone or M-CSF and RANK-L at 10 ng/ml for 3 days. The collagen gel was digested and cells collected and processed for scRNA-Seq.

first: Lauren

last: Schilling

first: Jungeun

first: Emily

last: Denker


title: Transcriptome analysis of paralysed CD8 dendritic cells after systemic inflammatory syndrome

first: Gordon

last: Smyth

summary: CD8 dendritic cells were harvested from the spleens of treated and untreated mice. Treated mice were innoculated intravenously with 40nmol of CpG1680 to induce systematic inflammation five days before harvesting cells. Cells from the treated mice show high CD103 expression. Cells from the untreated mice were FACS sorted based on CD103 expression into CD103 positive (low) and CD103 negative populations. RNA was extracted and profiled by RNA-seq. Two or three independent experiments were performed for each population, with 8 mice per group.

overall_design: RNA-seq profiles were generated of CD103 negative (3 biological replicates), CD103 low (2 biological replicates) and CD103 high dendritic cells (2 biological replicates).

first: Javier

last: Vega-Ramos

first: Jose

last: Villadangos

first: Aaron

middle: T

last: Lun


title: FLT3 tyrosine kinase inhibition modulates PRC2 and promotes differentiation in acute myeloid leukemia

first: Kathrin

last: Bernt

address: 3501 Civic Center Boulevard,

institute: Children's Hospital of philadelphia

summary: FLT3 is a receptor tyrosine kinase that is frequently mutated in AML. Inhibition of FLT3 induces myeloid differentiation in AML patients. We identified a role of FLT3 inhibition in decreasing EZH2 expression. To assess the impact of this regulation on PRC2 function, we performed H3K27me3 ChIP-Seq in a FLT3 mutated human AML cell line.

overall_design: Chromatin immunoprecipitation sequencing (ChIP-Seq) for H3K27me3 in MOLM14 cells treated with DMSO or quizartinib (AC220) for 24 hours. Drosophila S2 cells were spiked in prior to cell lysis for normalization of peaks.

first: Pamela

last: Sung

first: Simone

last: Riedel

first: Hongbo

last: Xie


title: Single cell RNA sequencing reveals endothelial cell killing and resolution pathways in experimental malaria-associated acute respiratory distress syndrome

city: Leuven

first: Emilie

last: Pollenus

address: Herestraat 49 bus 1044

country: Belgium

institute: KU Leuven

summary: Plasmodium parasites cause malaria, a global health disease that is responsible for more than 200 million clinical cases and 600 000 deaths each year. Most deaths are caused by various complications, including malaria-associated acute respiratory distress syndrome (MA-ARDS). Despite the very rapid and efficient killing of parasites with antimalarial drugs, 15% of patients with complicated malaria succumb. This stresses the importance of investigating resolution mechanisms that are involved in the recovery from these complications once the parasite is killed. To study the resolution of MA-ARDS, P. berghei NK65-infected C57BL/6 mice were treated with antimalarial drugs after onset of symptoms, resulting in 80% survival. Micro-Computed Tomography revealed an altered morphology of the lungs upon infection, with an increase in total and non-aerated lung volume due to edema. Whole body plethysmography confirmed a drastically altered lung ventilation, which was restored during resolution. Single-cell RNA sequencing indicated an increased inflammatory state in the lungs upon infection, which was accompanied by a drastic decrease in endothelial cells, consistent with CD8+ T cell mediated killing. During resolution, anti-inflammatory pathways were upregulated and proliferation of endothelial cells was observed. MultiNicheNet interactome analysis provided a wealth of possible pathways that could be involved in the disease resolution, which might be interesting to functionally validate. In conclusion, promotion of pro-resolving pathways that limit inflammation and promotion of endothelial cell proliferation may be an interesting approach for the design of adjunctive treatments to promote resolution after Plasmodium parasite killing by antimalarial drugs.

overall_design: Cells isolated from the lungs of uninfected control mice (CON), Untreated, PbNK65-infected C57BL/6 mice dissected at 8 days post infection (d8) and ART+CQ-treated, PbNK65-infected C57BL/6 mice dissected at 12 days post infection (d12). One sample per condition (CON, d8, d12) with each sample consisting of cells pooled from 4 mice. Cells from each mouse were labelled with TotalSeq B hashtag oligo's in order to be able to distinguish the cells from the 4 mice pooled in one sample.

first: Philippe

middle: E

last: Van den Steen


title: Engineered mischarged transfer RNAs for correcting pathogenic missense mutations

city: Chicago

first: Tao

last: Pan

address: 929 E. 57th Street

institute: University of Chicago

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

overall_design: Refer to individual Series


title: Directional endothelial communication by polarized extracellular vesicle release

status: Public on Jan 02 2024

city: Toronto

first: Sneha

last: Raju

state: ON

address: 101 College Street, PMCRT 3-309

country: Canada

institute: University of Toronto

summary: EVs were isolated from primary human aortic endothelial cells (ECs) (+/- IL-1b activation), quantified, and analysed by miRNA transcriptomics and proteomics.

overall_design: We performed miRNAHAEC EV-microRNA profiles were determined using HTG Molecular Diagnostic EdgeSeq technology (HTG Molecular Diagnostics, Inc, Tucson, AZ, USA).

first: Kathryn

last: Howe


title: Unveiling the Impact of ApoF Deficiency on Liver and Lipid Metabolism: Insights from Transcriptome-Wide m6A Methylome Analysis in Mice

city: Fuzhou

first: Mengting

last: Chen

address: Fujian Medical University, 1 Xuefubei Road, Minhou, Fuzhou, Fujian province, China

country: China

institute: Fujian Medical University

summary: The transcriptome-wide MeRIP sequencing in the ApoF KO mice has demonstrated the role of this crucial apolipoprotein in the liver health and the lipid metabolism.

overall_design: The aim of the present study was to elucidate the change of m6A methylome in the lipid metabolism disorder mice with ApoF gene knockout, based on the bioinformatics methods of transcriptome-wide m6A methylome epitranscriptomics.

first: Xuebin

last: Shen

first: Jian

last: Zhang

first: Yifan

last: Lin

first: Xinyue

last: Gao

first: Jionghong

last: Tu

first: Kunqi

first: An

last: Zhu

first: Shanghua

last: Xu


title: Fast and flexible profiling of chromatin accessibility and total RNA expression in single nucleus using Microwell-seq3

city: Hangzhou

first: Fang

last: Ye

state: Zhejiang

address: 866 Yuhangtang Rd

institute: Zhejiang University


title: The inositol phosphtase INPP4B sustains ILC1s and intratumoral NK cells through an AKT-driven survival pathway

status: Public on Jan 01 2024

city: St. Louis

first: Vincent

last: Peng

state: MO

address: 660 S. Euclid Ave.

institute: Washington University School of Medicine in St. Louis

summary: Innate lymphoid cells (ILCs) are a heterogeneous population of lymphocytes that coordinate early immune responses and maintain tissue homeostasis. Type 1 immune responses are mediated by natural killer (NK) cells and group 1 ILCs (ILC1s). Despite their shared features, NK cells and ILC1s display profound differences among various tissue microenvironments. Here, we identify the inositol polyphosphatase INPP4B as a hallmark feature of tissue-resident ILC1s and intratumoral NK cells using a scRNA-seq atlas of tissue-associated and circulating NK/ILC1s. Conditional deletion of Inpp4b in ILC1s and NK cells reveals that it is necessary for the homeostasis of tissue-resident ILC1s but not circulating NK cells at steady-state. Inpp4b-deficient cells display increased rates of apoptosis and reduced activation of the pro-survival molecule AKT. Furthermore, expression of Inpp4b by NK/ILC1s is necessary for their presence in the intratumoral environment and lack of Inpp4b impairs antitumor immunity. These findings highlight INPP4B as a novel regulator of tissue residency and antitumor function in ILC1s and NK cells.

overall_design: murine CD49a+ ILC1s from Inpp4b-flox and NCR1-iCre, Inpp4b-flox mice

first: Marco

last: Colonna


title: Increased Enhancer—Promoter Interactions during Developmental Enhancer Activation in Mammals

city: Irvine

first: Evgeny

last: Kvon

address: McGaugh Hall

institute: University of California, Irvine

summary: Remote enhancers are thought to interact with their target promoters via physical proximity, yet the importance of this proximity for enhancer function remains unclear. Here, we investigate the 3D conformation of enhancers during mammalian development by generating high-resolution tissue-resolved contact maps for nearly a thousand enhancers with characterized in vivo activities in ten murine embryonic tissues. 61% of developmental enhancers bypass their neighboring genes, which are often marked by promoter CpG methylation. The majority of enhancers display tissue-specific 3D conformations, and both enhancer–promoter and enhancer–enhancer interactions are moderately but consistently increased upon enhancer activation in vivo. Less than 14% of enhancer–promoter interactions form stably across tissues; however, these invariant interactions form in the absence of the enhancer and are likely mediated by adjacent CTCF binding. Our results highlight the general significance of enhancer–promoter physical proximity for developmental gene activation in mammals.

overall_design:
(capture hic) We generated high-resolution enhancer interactome maps across 10 mouse embryonic tissues for 935 bona fide developmental enhancers with characterized in vivo activity at mid-gestation, using enhancer-capture HiC.
(RNA-seq) Tissues were collected from E11.5 wildtype or enhancer-knockout embryos to perform differential expression analysis.

first: Zhuoxin


title: Expression of Atoh1, Gfi1, and Pou4f3 in the mature cochlea reprograms non-sensory cells into hair cells

city: Houston

first: Chenghang

last: Zong

state: TX

address: 1 Baylor Plaza

institute: Baylor College of Medicine

summary: Mechanosensory hair cells of the mature mammalian organ of Corti do not regenerate, and loss of hair cells leads to permanent hearing loss. Although non-mammalian vertebrates are able to regenerate hair cells by the division and transdifferentiation of adjacent supporting cells, many humans with severe hearing loss completely lack hair cells and supporting cells, with the organ of Corti being replaced by a flat epithelium of non-sensory cells To determine if mature non-sensory cells can produce hair cells in vivo, we reprogrammed non-sensory cells of the mature cochlea with three hair cell transcription factors, Gfi1, Atoh1, and Pou4f3.  We were able to generate significant numbers of hair cell-like cells in the non-sensory inner and outer sulci of the cochlea after 2 weeks of reprogramming. New hair cells continued to be added to the non-sensory regions of the cochlea over the next seven weeks, and expressed hair cell proteins such as Myosin VIIa, parvalbumin, and prestin and formed stereociliary hair bundles. We confirmed the identity of these cells by high depth single cell RNA-seq. Significantly, cells adjacent to converted hair cells expressed markers of supporting cells, suggesting that transcription factor reprogramming of non-sensory tissue in the adult animal can generate mosaics of sensory cells similar to those seen in the organ of Corti. Generating such sensory mosaics by reprogramming may represent a potential strategy for hearing restoration in humans.

overall_design: FACS sorted adult mouse cochlear cells from control (Fbxo2CreERT2::Rosa26-tdTomato) animals and experimental (Fbxo2CreERT2::Rosa26-tdTomato::Rosa26-GAP) animals based on tdTomato and EpCAM flourescnece.

first: Andrew

last: Groves

first: Melissa

last: McGovern

first: Yichi

last: Niu


title: RFX6 maintains gene expression and function of adult human islet ⍺ cells

city: Stanford

first: Vy

middle: N

last: Coykendall

state: California

address: 279 Campus Drive W

department: Developmental Biology

institute: Stanford University


title: Keratin 5 Basal Cells are Temporally Regulated Developmental and Tissue Repair Progenitors in Bladder Urothelium.

city: Columbus

first: Xin

last: Wang

state: OH

address: 700 Children's Dr,

department: The Kidney and Urinary Tract Center

institute: Nationwide Children's hosptial

summary: Urothelium forms a distensible yet impermeable barrier, senses and transduces stimuli, and defends the urinary tract from mechanical, chemical and bacterial injuries. Biochemical and genetic labeling studies support the existence of one or more progenitor populations with the capacity to rapidly regenerate the urothelium following injury, but slow turnover, a low mitotic index, and inconsistent methodologies obscure progenitor identity. The progenitor properties of basal Keratin 5 urothelial cells (K5-UC) have been previously investigated, but those studies focused on embryonic or adult bladder urothelium. Urothelium undergoes desquamation and apoptosis after birth, which requires postnatal proliferation and restoration. Therefore, we mapped the fate of bladder K5-UCs across postnatal development/maturation and following administration of cyclophosphamide to measure homeostatic and reparative progenitor capacities, respectively. In vivo studies demonstrate that basal K5-UCs are age-restricted progenitors in neonates and juveniles, but not in adult mice. Neonatal K5-UCs retain a superior progenitor capacity in vitro, forming larger and more differentiated urothelial organoids than adult K5-UCs. Accordingly, K5-UC transcriptomes are temporally distinct, with enrichment of transcripts associated with cell proliferation and differentiation in neonates. Induction of urothelial proliferation is sufficient to restore adult K5-UC progenitor capacity. Our findings advance the understanding of urothelial progenitors and support a linear model of urothelial formation and regeneration, which may have significant impact on therapeutic development or tissue engineering strategies.

overall_design: Given the distinct progenitor capacities observed by neonatal and adult K5-UCs, we sorted K5-UCs from P7 and P35 bladders and used RNAseq to determine whether functional differences could be explained by gene expression.

first: Brian

last: Becknell

first: Mohammad

last: El-Harakeh

first: Felipe

last: Rodriguez-Tirado

first: Kelly

last: Grounds

first: Birong

last: Li

first: Ashley

last: Jackson


title: RFX6 maintains gene expression and function of adult human islet ⍺ cells [RNAseq]

summary: Transcriptomic profiling by high-throughput sequencing of primary human alpha and beta cells following RFX6 loss of function

overall_design: Profiling of FACS-purified human pancreatic RFX6 shRNA-mediated knockdown GFP+ alpha or beta cells by RNA-Seq, compared to non-targeting shRNA control for each donor

first: Coykendall

last: VMN

first: Kim

last: SK


title: RFX6 maintains gene expression and function of adult human islet ⍺ cells [CUT&RUN]

summary: RFX6 genome-binding profiling by CUT&RUN and high-throughput sequencing

overall_design: We analyzed the genomic targets of RFX6 by RFX6-FLAG overexpression in primary human pseudoislets and CUT&RUN, comparing anti-Flag antibody to anti-IgG control for each donor


title: Lineage-specific intolerance to oncogenic drivers restricts histological transformation

city: NEW YORK

last: Laughney

state: New York

address: 413 East 69th Street

department: Physiology, Biophysics, and Systems Biology

institute: Weill Cornell Medicine


title: Therapeutic application of human type 2 innate lymphoid cells via induction of granzyme B-mediated tumor cell death

city: Duarte

first: Zhenlong

address: 1500 East Duarte Road

institute: City of hope

summary: The therapeutic potential for human type 2 innate lymphoid cells (ILC2s) has been underexplored.  While not observed in mouse ILC2s, we have found that human ILC2s secrete granzyme B (GZMB) and directly lyse tumor cells by inducing pyroptosis and/or apoptosis, which is governed by a DNAM-1CD112/CD155 interaction that inactivates FOXO1. Over time, the high surface density expression of CD155 in AML impairs expression of DNAM-1 and GZMB, thus allowing for immune evasion. We describe a reliable platform capable of up to 2,000-fold expansion of human ILC2s within 4 weeks, whose molecular and cellular ILC2 profile has been validated by single-cell RNA sequencing. In both AML and solid tumor models, exogenously administered expanded human ILC2s show significant antitumor effects in vivo. Collectively, we demonstrate previously unreported properties of human ILC2s and identify this innate immune cell subset as a new member of the cytolytic immune effector cell family.

overall_design: Identification of ILC2 regulatory pathways treated by IL-15

first: Jianhua

last: Yu


title: Behavioral and transcriptomic analyses of mecp2 function in zebrafish

city: Madison

first: Nicholas

middle: Joel

last: Santistevan

state: WI

address: 145 Noland Hall, 250 N Mills St

department: Department of Integrative Biology

institute: University of Wisconsin – Madison

summary: Rett Syndrome (RTT), a human neurodevelopmental disorder characterized by severe cognitive and motor impairments, is caused by dysfunction of the conserved transcriptional regulator Methyl-CpG-binding protein 2 (MECP2). Genetic analyses in mouse Mecp2 mutants, which exhibit key features of human RTT, have been essential for deciphering the mechanisms of MeCP2 function; nonetheless, our understanding of these complex mechanisms is incomplete. Zebrafish mecp2 mutants exhibit mild behavioral deficits but have not been analyzed in depth. Here we combine transcriptomic and behavioral assays to assess baseline and stimulus-evoked motor responses and sensory filtering in zebrafish mecp2 mutants from 5-7 days post-fertilization (dpf). We show that zebrafish mecp2 function is dispensable for gross movement, acoustic startle response, and sensory filtering (habituation and sensorimotor gating), and reveal a previously unknown role for mecp2 in behavioral responses to visual stimuli. RNA-seq analysis identified a large gene set that requires mecp2 function for correct transcription at 4 dpf, and pathway analysis revealed several pathways that require MeCP2 function in both zebrafish and mammals. These findings show that MeCP2’s function as a transcriptional regulator is conserved across vertebrates and supports using zebrafish to complement mouse modeling in elucidating these conserved mechanisms.

overall_design: To investigate the role of MeCP2 as a transcriptional regulator in zebrafish, we performed differential gene expression analyses on 10 mutant and 10 wild type samples and highlighting the effects of the Q63X mutation in MECP2 in various molecular pathways.

middle: J

first: Colby

last: Ford

first: Cole

middle: S

last: Gilsdorf

first: Yevgenya

last: Grinblat


title: Lineage-specific intolerance to oncogenic drivers restricts histological transformation [scRNA-Seq]

summary: Lung adenocarcinoma (LUAD) and small cell lung cancer (SCLC) are thought to originate from different epithelial cell types in the lung. Intriguingly, LUAD can histologically transform into SCLC following treatment with targeted therapies. Here we designed models to follow the conversion of LUAD to SCLC and found the barrier to histological transformation converges on tolerance to Myc, which we implicate as a lineage-specific driver of the pulmonary neuroendocrine cell. Histological transformations are frequently accompanied by activation of the Akt pathway. Manipulating this pathway permitted tolerance to Myc as an oncogenic driver, producing rare, stem-like cells, transcriptionally resembling the pulmonary basal lineage. These findings suggest histological transformation may require the plasticity inherent to the basal stem cell, enabling tolerance to previously incompatible oncogenic driver programs.

overall_design: Single cell RNA sequencing was performed for 27 samples from genetically engineered mouse models. All samples were sorted from fresh, enzymatically dissociated lung tissues and, in most cases, triplicates were pooled before sequencing.

first: Eric

last: Gardner

first: Ethan

last: Earlie

first: Kate

first: Jerin

last: Thomas

first: Melissa

last: Hubisz

first: Benjamin

middle: D

last: Stein

first: Chen

last: Zhang

first: Lewis

last: Cantley

first: Harold

last: Varmus


title: Yeast eIF2A plays a minimal role in translation initiation in vivo

city: Bethesda

first: Swati

last: Gaikwad

state: MD

address: Building 6, Room 233

department: Section on Nutrient Control Of Gene Expression

institute: National Institutes of Health

summary: Earlier investigations have associated mammalian eIF2A with Met-tRNAi binding to the 40S subunit and its recruitment to specialized mRNAs in a GTP-independent manner. Additionally, eIF2A has been implicated in non-AUG start codon initiation, particularly under conditions where eIF2 function is attenuated by phosphorylation of its α-subunit during stress or starvation. However, the precise role of eIF2A in vivo translation remains unclear. Moreover, it's uncertain if the conserved ortholog in budding yeast can functionally substitute for eIF2 during stress. To address these questions, we conducted ribosome profiling on a yeast deletion mutant lacking eIF2A, alongside isogenic wild-type (WT) cells, both in the presence or absence of eIF2α phosphorylation induced by amino acid starvation.

overall_design: This study includes a total of 16 libraries comprised of 8 RNA-Seq libraries (mRNA) and 8 ribosome footprint profiling libraries (ribo). The samples used for library preparation were sourced from two biological replicates of both the BY4742 (WT) and yeast eIF2A∆ deletion mutant F2247 (eIF2A∆). The strains were cultured at 30ºC in synthetic complete (SC) medium to log-phase (A600 = 0.5–0.6). Additionally, the WT and eIF2A∆ strains were grown in SC media lacking isoleucine and valine (SC-Ile/Val) to log-phase (A600 = 0.5–0.6). Subsequently, they were treated with the antimetabolite Sulfometuron methyl (SM) at 1 µg/mL for 25 min to induce eIF2α phosphorylation.

first: Fardin

last: Ghobakhlou

first: Hongen

first: Alan

middle: G

last: Hinnebusch


first: Zhenlong

last: Li


title: Examination and comparison of the RNA extraction methods using mouse serum

city: Hirosaki

first: Mitsuru

last: Chiba

state: Aomori

address: 66-1 Hon-cho

department: Graduate School of Health Sciences

institute: Hirosaki University

summary: Serum miRNAs are considered useful as non-invasive biomarkers for various diseases, but the optimal method for extracting RNA from serum is currently unknown. In this study, several RNA extraction kits were used to determine which kit is the optimal method. RNA was extracted from the serum of 8-week-old C57BL/6NJcl male mice according to the protocol of each RNA extraction kit. The yield of extracted RNA samples was calculated and electrophoretic patterns were evaluated by Agilent bioanalyzer. Expression patterns of the extracted RNA samples were confirmed by Agilent mouse miRNA microarray. The results showed significant differences in RNA yields in the miRNeasy serum/plasma advanced kit, and mirVana™ PARIS™ RNA and Native Protein Purification Kit compared to almost all other samples. Furthermore, two peaks were identified in the miRNeasy serum/plasma advanced kit using small RNA kit of Agilent bioanalyzer, one at 20-40 nucleotides (nt) and the other around 40-100 nt whereas the other reagents had a single peak. In addition, a high correlation was observed between the two RNA extraction kits in microarray. These results suggest that the above two kits are suitable for miRNA extraction from mouse serum.

overall_design: RNAs were extracted using 200 μL of serum from 8-week-old C57BL/6NJcl male mice and 2 reagents: miRNeasy serum/plasma advanced kit (QIAGEN), and mirVana™ PARIS™ RNA and Native Protein Purification Kit (Thermo Fisher Scientific)  according to each manufacturer’s protocol. In addition, RNAs were extracted using exoRNeasy midi kit (QIAGEN) to extract RNA from the aqueous layer of 200 μL of serum to which 700 μL of QIAzol Lysis Reagent (QIAGEN) was added and separated in two layers to determine the amount of total RNA in the serum.

first: Keisuke

last: Yamamoto


title: Lineage-specific intolerance to oncogenic drivers restricts histological transformation [ATAC-Seq]

first: Ashley

last: Laughney

overall_design: Bulk ATAC-sequencing was performed for 15 high quality libraries across 8 biological conditions.


title: Effect of knockdown RUNX1-IT1 on gene expression in ES2 cells

city: Beijing

first: Pengfei

last: Zhao

address: No.17, Panjiayuan Nanli, South Huawei Road

institute: National Cancer Center/Cancer Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College

summary:
To investigate the function of RUNX1-IT1 in ovarian cancer progression, we constructed ES2 cell lines with knockdown of RUNX1-IT1 and the control group.
We then performed gene expression profiling analysis using data obtained from RNA-seq of 2 different cells.

overall_design: Comparative gene expression profiling of RNA-seq data from RUNX1-IT1 knockdown group and control group.

first: Xiao

last: Yu


summary: The therapeutic potential for human type 2 innate lymphoid cells (ILC2s) has been underexplored.  While not observed in mouse ILC2s, we have found that human ILC2s secrete granzyme B (GZMB) and directly lyse tumor cells by inducing pyroptosis and/or apoptosis, which is governed by a DNAM-1-CD112/CD155 interaction that inactivates FOXO1. Over time, the high surface density expression of CD155 in AML impairs expression of DNAM-1 and GZMB, thus allowing for immune evasion. We describe a reliable platform capable of up to 2,000-fold expansion of human ILC2s within 4 weeks, whose molecular and cellular ILC2 profile has been validated by single-cell RNA sequencing. In both AML and solid tumor models, exogenously administered expanded human ILC2s show significant antitumor effects in vivo. Collectively, we demonstrate previously unreported properties of human ILC2s and identify this innate immune cell subset as a new member of the cytolytic immune effector cell family.

overall_design: Identification of the phenotypes of expanded ILC2s and freshly isolated ILC2s from PBMCs

first: Jianhua


title: Clinical and molecular features of acquired resistance to immunotherapy in non-small cell lung cancer

city: Philadelphia

first: Jingya

last: Qiu

address: 421 Curie Blvd, 532

department: Radiation Oncology

institute: University of Pennsylvania

summary: Although cancer immunotherapy with PD-(L)1 blockade is now routine treatment for patients with lung cancer, remarkably little is known about acquired resistance. We examined 1,201 patients with NSCLC treated with PD-(L)1 blockade to clinically characterize acquired resistance, finding it to be common (occurring in more than 60% of initial responders), with persistent but diminishing risk over time, and with distinct metastatic and survival patterns compared to primary resistance. To examine the molecular phenotype and potential mechanisms of acquired resistance, we performed whole transcriptome and exome tumor profiling in a subset of NSCLC patients (n=29) with acquired resistance. Systematic immunogenomic analysis revealed that tumors with acquired resistance differentially expressed features of inflammation and interferon (IFN) signalling. In particular, relapsed tumors could be separated into those with either upregulated or with stable expression of IFNγ response genes. Tumors with upregulated IFNγ response genes were associated with putative routes of resistance characterized by signatures of persistent IFN signaling, immune dysfunction, and mutations in antigen presentation genes. Transcriptomic profiling of cancer cells from multiple murine models of acquired resistance to PD-(L)1 blockade also showed evidence of high IFN-stimulated genes coupled to an altered or attenuated induced response after in vitro IFNγ treatment. In summary, we characterized clinical and molecular features of acquired resistance to PD-(L)1 blockade in NSCLC and found evidence of ongoing but altered IFN response. The persistently inflamed, rather than excluded or deserted, tumor microenvironment of acquired resistance informs therapeutic strategies to effectively reprogram and reverse acquired resistance.

overall_design: Bulk RNA-seq of parental, exposed to low-level chronic IFNG treatment, and ICB-relapsed LLC1 and KP mouse lung cancer tumors (CD45-); 3 biological replicates for each condition

first: Danish

last: Memon

first: Adam

last: Schoenfeld

first: Darwin

last: Ye

first: George

last: Fromm

first: Hira

last: Rizvi

first: Xiang

first: Mohamed

middle: R

last: Keddar

first: Divij

last: Mathew

first: Kyung

last: Yoo

first: Jayon

last: Lihm

first: Jayalaksmi

last: Miriyala

first: Jennifer

last: Sauter

first: Jia

last: Luo

first: Andrew

last: Chow

first: Umesh

middle: K

last: Bhanot

first: Caroline

last: McCarthy

first: Chad

last: Vanderbilt

first: Cailian

last: Liu

first: Mohsen

last: Abu-Akeel

last: Plodkowski

first: Nicholas

last: McGranahan

first: Marta

last: Łuksza

last: Greenbaum

first: Taha

last: Merghoub

first: Ikbel

last: Achour

first: J

last: Barrett

first: Ross

last: Stewart

first: Pedro

last: Beltrao

first: Taylor

last: Schreiber

first: Andy

last: Minn

first: Martin

last: Miller

first: Matthew

last: Hellmann


title: Novel pan-ERR agonists ameliorate heart failure through enhancing cardiac fatty acid metabolism and mitochondrial function

first: Weiyi

last: Xu

address: One Baylor Plaza, 441E

department: Department of Molecular and Human Genetics

summary: Cardiac metabolic dysfunction is a hallmark of heart failure. Estrogen related receptors ERRalpha and ERRgamma are essential regulators of cardiac metabolism. Therefore, activation of ERR could be a potential theraputic intervention for heart failure. However, in vivo studies demonstrating the potential utility of ERR agonists for heart failure treatment are lacking, as compounds with pharmackinetics appropriate for in vivo use have not been available. Using a structure-based design approach, we designed and synthesized two structurally distinct pan-ERR agonists, SLU-PP-332 (332) and SLU-PP-915 (915), which significantly improved ejection fraction, ameliorated fibrosis and increased survival associated with pressure overload-induced heart failure without affecting cardiac hypertrophy. Mechanistically, a broad spectrum of metabolic genes was transcriptionally activated by ERR agonists, particulary genes involved in fatty acid metabolism and mitochondrial function. Using both in vitro and in vivo genetic dependency experiments, we show that ERRgamma is the main mediatorof ERR agonism-induced transcriptional regulation and cardioprotection and definitively demonstrated target specificity. Additionally, ERR agonism also led to downregulation of cell cycle and development pathways, which was partially mediated by E2F1 in cardiomyocyte. In summary, ERR agonists maintain oxidative metabolism, which confers cardiac protection against pressure overload-induced heart failure in vivo. Our results provide direct pharmacological evidence supporting the further in vivo development of ERR agonists as novel heart failure theraputics.

overall_design: To investigate the effect of ERR agonists 332 and 915 on the transcriptome of neonatal rat ventricular myocytes (NRVM). NRVMs were treated with vehicle (DMSO) or 10uM 915 for 72h.

first: Andrea

last: Wilderman

first: Lilei

last: Zhang

first: Thomas

last: Burris

first: Ronald

last: Evans


title: B3GALT6 Promotes Dormant Breast Cancer Cell Survival and Recurrence by Enabling Heparan Sulfate-Mediated FGF Signaling

first: Lewis

last: Chodosh

state: Pennsylvania

address: 421 Curie Blvd

department: Cancer Biology

summary: Breast cancer mortality results from incurable recurrent tumors, putatively seeded by dormant, therapy-refractory residual tumor cells (RTCs). Understanding the mechanisms enabling RTC survival is therefore essential for improving patient outcomes. We derived a dormancy-associated RTC signature that mirrors the transcriptional response to neoadjuvant chemotherapy in patients and is enriched for extracellular matrix-related pathways. In vivo CRISPR-Cas9 screening of dormancy-associated candidate genes identified the galactosyltransferase B3GALT6 as a functional regulator of RTC fitness. B3GALT6 is required for the linkage of glycosaminoglycans (GAGs) to proteins to generate proteoglycans and its germline loss-of-function causes skeletal dysplasias. We determined that B3GALT6-mediated biosynthesis of heparan sulfate GAGs predicts poor patient outcomes, promotes tumor recurrence by enhancing dormant RTC survival in multiple contexts, and does so via a B3GALT6-heparan sulfate/HS6ST1-heparan 6-O-sulfation/FGF1-FGFR2 signaling axis. These findings implicate B3GALT6 in cancer and suggest targeting of FGFR2 signaling as a novel approach to eradicate dormant RTCs, thereby preventing recurrence.

overall_design: To investigate the genomic and molecular changes through dormancy and recurrence , we designed an in-vitro time series experiment on an in-house cell line derived from a Her2/neu mouse model

first: Amulya

last: Sreekumar

first: Tien-chi

last: Pan

first: Dhruv

last: Pant


title: Mechanisms Driving Fasting-Induced Protection from Genotoxic Injury in the Small Intestine

first: Jason

last: Mills

department: Gastroenterology

summary: To elucidate the mechanisms required for short-term fasting to protect healthy intestinal cells from chemotherapy injury, we conducted an investigation comparing the effects of altered mTORC1 modulation in DDIT4 knockout male mice to wild-type male mice at various timepoints following doxorubicin treatment.

overall_design: Wild type and DDIT4 knockout male mice were ad libitum fed or fasted for 24 hours then treated with 20 mg/kg of doxorubicin by intraperitoneal injection. Small intestines were harvested 2, 6, 12, and 24 hours following doxoxrubicn admistration, in addition to untreated controls at 0, 2, and 6 hours. RNA was isolated from whole tissue proximal small intestine. Normalized and transformed gene expression values were used to generate heatmaps and line graphs for pathways of interest.

first: Kali

last: Deans-Fielder

first: Timothy

last: Wu

first: Steven

last: Bark

first: Yang-Zhe

last: Huang

first: Noah

middle: F

last: Shroyer


title: SHORT CHAIN FATTY ACIDS MITIGATE OSTEOCLAST-MEDIATED ARTHRITIC BONE REMODELLING

city: New York

first: Sergei

last: Koralov

state: NY

address: 550 1st Avenue

department: Pathology

institute: NYU School of Medicine

summary: Objective: To study the effects of Short Chain Fatty Acids (SCFAs) on arthritic bone remodeling.  Methods:  We treated a recently described preclinical murine model of psoriatic arthritis (PsA), R26STAT3Cstopfl/fl CD4Cre mice, with SCFA supplemented water. We also performed in vitro osteoclast differentiation assays in the presence of serum-level SCFAs to evaluate the direct impact of these microbial metabolites on maturation and function of osteoclasts. We further characterized the molecular mechanism of SCFAs by bulk transcriptomics analysis.  Results: The osteoporosis condition in R26STAT3Cstopfl/fl CD4Cre animals is attributed primarily to an expansion of osteoclast progenitor cells (OCPs), leading to robust osteoclast differentiation. We show that SCFA supplementation can rescue the osteoporosis phenotype in this model of PsA. Our in vitro experiments revealed an inhibitory effect of the SCFAs on osteoclast differentiation, even at very low serum concentrations. This suppression of osteoclast differentiation enabled SCFAs to impede osteoporosis development in R26STAT3Cstopfl/fl CD4Cre mice. Further interrogation revealed that bone marrow derived OCPs from diseased mice expressed a higher level of SCFA receptors than that of control mice and that the progenitor cells in the bone marrow of SCFA-treated mice presented a modified transcriptomic landscape, suggesting a direct impact by SCFAs on osteoclast progenitors.   Conclusion: We demonstrated how gut microbiota-derived SCFAs can regulate distal pathology, i.e., osteoporosis, and identified a potential therapeutic option for restoring bone density in rheumatic disease, further highlighting the critical role of the gut-bone axis in these disorders.

overall_design: Control mice and R26STAT3Cstopfl/fl CD4Cre mice were either untreated or treated with SCFAs. At ~ 8 weeks their bone marrow was extracted and cultured for 24 hours. Non-adherent progenitor cells were collected for RNA extraction and bulk sequencing. We then compared differential gene expression across genotype and treatment.

first: Katharine

middle: L

last: Yang

first: Briana

last: Mullins

first: Alannah

last: Lejeune

first: Ellie

last: Ivanova

first: Jong

last: Shin

first: Sofia

last: Bajwa

first: Richard

last: Possemato

first: Ken

last: Cadwell

first: Jose

middle: U

last: Scher


title: Microbiota modulation by dietary oat beta-glucan prevents Non-alcoholic fatty liver disease progression

city: Aachen

first: Kai Markus

last: Schneider

address: Pauwelsstraße 30

department: Medicine III

institute: University Hospital RWTH Aachen

summary: Background & Aims: Non-alcoholic fatty liver disease (NALFLD)-associated changes in gut microbiota are important drivers of disease progression toward fibrosis. Therefore, reversing microbiota alterations could ameliorate NAFLD progression. Oat beta-glucan, a non-digestible polysaccharides, has shown promising therapeutic effects on hyperlipidemia associated with NAFLD, but its impact on gut microbiota and most importantly NAFLD fibrosis remains unknown. Methods: We performed detailed metabolic phenotyping including body composition, glucose tolerance, and lipid metabolism as well as comprehensive characterization of the gut-liver axis in a western-style diet (WSD)-induced model of NAFLD and assessed the effect of a beta-glucan intervention on early and advanced liver disease. Gut microbiota was modulated using broad-spectrum antibiotic (Abx) treatment. Results: Oat beta-glucan supplementation did not affect WSD-induced body weight gain, glucose intolerance, and the metabolic phenotype remained largely unaffected. Interestingly, oat beta-glucan dampened NAFLD inflammation, associated with significantly reduced monocyte-derived macrophages (MoMFs) infiltration, fibroinflammatory gene expression, and strongly reduced fibrosis development. Mechanistically, this protective effect was not mediated by changes in bile acid composition or signaling, but was dependent on gut microbiota and was lost upon Abx treatment. Specifically, oat beta-glucan partially reversed unfavorable changes in gut microbiota, resulting in an expansion of protective taxa, including Ruminococcus, and Lactobacillus followed by reduced translocation of TLR ligands. Conclusions: Our findings identify oat beta-glucan as a highly efficacious food supplement that dampens inflammation and fibrosis development in diet-induced NAFLD. These results, along with its favorable dietary profile, suggest that it may be a cost-effective and well-tolerated approach to preventing NAFLD progression and should be assessed in clinical studies.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for liver tissue of male C57BL/6J mice.

first: Julius

last: Jaeger

first: Annette

last: Brandt

first: Wenfang

last: Gui

first: Timur

last: Yergaliyev

first: Angélica

last: Hernández-Arriaga

first: Mukil

middle: M

last: Muthu

first: Ahmed

last: Elashy

first: Karolina

last: Edlund

first: Antonio

last: Molinaro

first: Diana

last: Möckel

first: Jan

last: Sarges

last: Hengstler

first: Emina

last: Halibasic

first: Michael

last: Trauner

first: Florian

last: Kahles

first: Carolin

middle: V

first: Twan

last: Lammers

first: Hans-Ulrich

last: Marshall

first: Amélia

last: Camarinha Silva

first: Ina

last: Bergheim

first: Christian

last: Trautwein

first: Kai


title: RNA-Sequencing of EndoC-BH1 cells Overexpressing OAS1, OAS2, or OAS3

city: Boston

first: Jonathan

last: Dreyfuss

state: MA

address: 1 Joslin Pl

department: Bioinformatics & Biostatistics Core

institute: Joslin Diabetes Center

summary: EndoC-BH1 cells were plated in normal media and forward transfected with lipofectamine 3000 (2ug cDNA) of empty, OAS1, OAS2, or OAS3 plasmids from Vector builder

overall_design: There were 4 replicates of each plasmid for a total of 16 samples.

first: Dario

last: De Jesus


title: Molecular refinement of pilocytic astrocytoma in adult patients

city: Hamburg

first: Helena

last: Bode

address: Martinistrasse 52

institute: Research Institute Children‘s Cancer Center Hamburg

type: Methylation profiling by genome tiling array

summary: Genome wide DNA methylation profiling of 79 histologically diagnosed pilocytic astrocytoma (PA) in adults, of which we were able to upload 71 samples. The methylation profiling indicates that many of the histologically diagnosed PA do not show a concordant methylation classification.

overall_design: 71/79 samples of histologically diagnosed pilocytic astrocytoma that were analyzed. All samples passed the detection p-value cut-off of <0.01.

first: Ulrich

last: Schüller


title: A gene-expression signature in fibroblasts of Amyotrophic Lateral Sclerosis

city: Catania

first: Sebastiano

last: Cavallaro

address: Via Paolo Gaifami, 18

country: Italy

institute: IRIB-CNR

summary: In this study, we characterized transcriptome changes in skin fibroblasts of sporadic ALS patients (SALS) and controls and evaluated their utility as a molecular classifier for ALS diagnosis.

overall_design: Whole gene expression profiles of skin fibroblasts from SALS patients and healthy controls were analyzed. Fibroblast samples were obtained from three healthy individuals and nine patients diagnosed with SALS.

first: Giovanna

last: Morello

first: Valentina

last: La Cognata

first: Maria

last: Guarnaccia


title: Dietary tryptophan catabolite released by intratumoral Lactobacillus reuteri facilitates αPD-L1 therapy

city: Pittsburgh

first: Alok

last: Joglekar

address: Assembly 3045, 5051 Center Ave

department: Immunology

institute: University of Pittsburgh

summary: Here, we elucidate a critical microbial-host crosstalk between probiotic-released aryl hydrocarbon receptor (AhR) agonist indole-3-aldehyde (I3A) and CD8 T cells within the tumor microenvironment that potently enhances spontaneous antitumor immunity and facilitates ICI therapy in preclinical melanoma. We used single cell RNA sequencing to phenotype CD8+ T cells within tumors of L. reuteri -treated or untreated mice.

overall_design: CD3+ T cells from tumors of mice treated with L. reuteri or untreated mice were isolated by magnetic selection and FACS and subjected to single cell RNA sequencing

first: Mackenzie

last: Bender

first: Alex

middle: C

last: McPherson

first: Marlies

last: Meisel


title: Differential signaling effects of Escherichia coli and Staphylococcus aureus in human whole blood indicate distinct regulation of the NRF2 Pathway

city: Nashville

first: Ryan

last: Stark

state: Tennessee

address: 2200 Children's Way, 5121 Doctors' Office Tower

institute: Vanderbilt University Medical Center

summary: Escherichia coli and Staphylococcus aureus are two of the most common bacterial species responsible for severe sepsis. While it is observed that they have disparate clinical phenotypes, the signaling differences elicited by each bacteria that drive this variance remain unclear. Therefore, we utilized human whole blood exposed to heat-killed E. coli or S. aureus and measured the transcriptomic signatures. Relative to unstimulated control blood, heat-killed bacteria exposure led to significant dysregulation (up and down-regulated) of  >5000 genes for each experimental condition, with a slight increase in gene dysregulation by S. aureus. While there was significant overlap regarding pro-inflammatory gene ontology pathways, heat-killed E. coli had a more considerable impact on purine metabolism than heat-killed S. aureus, which had a more significant effect on nitric oxide-mediated processes. Utilizing Ingenuity Pathway Analysis, it was predicted that nuclear factor erythroid 2-related factor 2 (Nrf2) signaling, a main transcription factor in anti-oxidant responses, was predominately up-regulated in S. aureus relative to E. coli. Further, the use of pharmacologics that preferentially targeted the Nrf2 pathway led to differential cytokine profiles depending on the type of bacterial exposure. These findings reveal a significant inflammatory dysregulation between E. coli and S. aureus and provide insight into the potential of targeting unique pathways to curb the bacteria species-specific response.

overall_design: Total of 12 samples were analyzed, 4 were human whole blood exposure to heat-killed E. coli (HKEC),  4 were human whole blood exposure to heat-killed S. aureus (HKSA). RNA was extracted from each of these samples for bulk RNA-Sequencing analysis.

first: Allison

last: Pourquoi

first: Michael

last: Miller

first: Stephen

last: Koch

first: Katherine

last: Boyle

first: Victoria

last: Surratt

first: Hong

last: Nguyen

first: Sourav

last: Panja

first: Jean-Philippe

last: Cartailler

first: Shristi

last: Shrestha


title: The ganglioside GM3 protects against abdominal aortic aneurysm by suppressing ferroptosis in vascular smooth muscle cells

city: tianjin

first: kan

last: li

address: Tianjin Medical University, 22 Qixiangtai Road, Tianjin, China

institute: Tianjin Medical University

summary: Background: Abdominal aortic aneurysm (AAA) is a potentially life-threatening condition, but approved medical therapies to prevent AAA progression and rupture are currently lacking. Sphingolipids metabolism disorders are associated with the occurrence and development of AAA. It has been discovered that ganglioside GM3, a sialic acid-containing type of glycosphingolipid, plays a protective role in atherosclerosis which is an important risk factor for AAA, but the potential contribution of GM3 to AAA development has not been investigated.  Methods: We performed a metabolomics study to evaluated GM3 level in plasma of human AAA patients. We profiled GM3 synthase (ST3GAL5) expression in the mouse model of aneurysm and human AAA tissues through western blotting and immunofluorescence staining. RNA sequencing, proteomic analysis, affinity purification and mass spectrometry, surface plasmon resonance (SPR) analysis, and functional studies were used to dissect the molecular mechanism of GM3-regulating ferroptosis. We conditionally deleted and overexpressed St3gal5 in smooth muscle cells (SMCs) in vivo to investigate its role in AAA.  Results: We found significantly reduced plasma levels of the GM3 in human AAA patients. GM3 content and ST3GAL5 expression were all decreased in abdominal aortic VSMCs in AAA patients and mouse model. RNA-sequencing analysis showed that ST3GAL5 silencing in human aortic SMCs (HASMCs) induced ferroptosis. Importantly, we showed that GM3 interacted directly with the extracellular domain of transferrin receptor 1 (TFR1), a cell membrane protein critical for cellular iron uptake, disrupted its interaction with holo-transferrin. SMC-specific St3gal5 knockout exacerbated iron accumulation at lesion sites and significantly promoted AAA development, while GM3 supplementation suppressed lipid peroxidation, reduced iron deposition in aortic VSMCs and markedly decreased AAA incidence.  Conclusions: Together, these results suggest that GM3 dysregulation promotes ferroptosis of VSMCs in AAA. Furthermore, GM3 may constitute a new therapeutic target for the treatment of AAA.

overall_design: To understand how GM3 affected cellular processes in VSMCs, we conducted RNA sequencing (RNA-seq) in human aortic smooth muscle cells (HASMCs) transfected with ST3GAL5 siRNA (siST3GAL5) or control siRNA (siNC).

first: Fangni

last: Zhang

first: Kan

last: Li

first: Kaiwen

last: Bao

first: Chunyong

first: Hongfeng

last: Jiang

first: Ding

last: Ai