title: A highly conserved tRNA modification contributes to C. albicans filamentation and virulence

city: Zürich

first: Puneet

last: Sharma

state: Zürich

address: Otto-Stern-Weg 3

department: Institute of Biochemistry

institute: ETH

summary: tRNA modifications play important roles in maintaining translation accuracy in all domains of life. Disruptions in the tRNA modification machinery, especially of the anticodon stem loop, can be lethal for many bacteria and lead to a broad range of phenotypes in baker’s yeast. Very little is known about the function of tRNA modifications in host-pathogen interactions, where rapidly changing environments and stresses require fast adaptations. We found that two closely related fungal pathogens of humans, the highly pathogenic Candida albicans and its much less pathogenic sister species, Candida dubliniensis, differ in the function of a tRNA-modifying enzyme. This enzyme, Hma1, exhibits speciesspecific effects on the ability of the two fungi to grow in the hypha morphology, which is central to their virulence potential. We show that Hma1 has tRNA-threonylcarbamoyladenosine dehydratase activity, and its deletion alters ribosome occupancy, especially at 37°C—the body temperature of the human host. A C. albicans HMA1 deletion mutant also shows defects in adhesion to and invasion into human epithelial cells and shows reduced virulence in a fungal infection model. This links tRNA modifications to host-induced filamentation and virulence of one of the most important fungal pathogens of humans.

overall_design: Candida albicans wild type and hma1 knockout strains were grown at 30°C and 37°C and analyzed by ribosome profiling to study the role of ct6A in translation.

first: Bettina

last: Böttcher

first: Sandra

last: Kienast

first: Johannes

last: Leufken

first: Cristian

last: Eggers

first: Christine

first: Bianka

last: Morgner

first: Daniela

last: Schulz

first: Hannes C

middle: A

last: Drexler

first: Stefanie

last: Allert

first: Ilse

last: Jacobsen

first: Slavena

last: Vylkova

first: Sebastian

last: Leidel

first: Sascha

last: Brunke


title: Foxp striatal compensation (RNA-Seq)

status: Public on Apr 04 2024

city: Dallas

first: Genevieve

last: Konopka

state: Texas

address: 5323 Harry Hines Blvd.

department: Neuroscience

country: USA

institute: University of Texas Southwestern Medical Center

type: Expression profiling by high throughput sequencing

summary: Spiny projection neurons (SPNs) of the striatum are critical in integrating neurochemical information to coordinate motor and reward-based behavior. Mutations in the regulatory transcription factors expressed in SPNs can result in neurodevelopmental disorders (NDDs). Paralogous transcription factorsFoxp1andFoxp2, which are both expressed in the dopamine receptor 1 (D1) expressing SPNs, are known to have variants implicated in NDDs. Utilizing mice with a D1-SPN specific loss ofFoxp1,Foxp2, or both and a combination of behavior, electrophysiology, and single-nuclei RNA (snRNA-seq) and single-nuclei Assay for Transposase-Accessible Chromatin sequencing (snATAC-seq), we find that loss of both genes results in impaired motor and social behavior as well as increased firing of the D1-SPNs. Differential gene expression analysis of snRNA-seq data implicates genes involved in autism risk, altered electrophysiological properties, and neuronal development and function. These data indicate complementary roles betweenFoxp1andFoxp2in the D1-SPNs.

overall_design: The transcription factors Foxp1 and Foxp2 are both expressed in the D1-SPNs of the striatum. We used single nuclei RNA sequencing (snRNA-Seq) in both juvenile (P9) and adult (P56) mouse striatum to test the effects of loss of one or both of these genes from the D1-SPNs.

first: Newaz

last: Ahmed

first: Nitin

last: Khandelwal

first: Ashley

last: Anderson

first: Ashwinikumar

last: Kulkarni

first: Karthigayini

last: Sivaprakasam

first: Jay

last: Gibson


title: The Myc-Associated Zinc Finger Protein (MAZ) controls STAT1-mediated antiviral response by reshaping epigenome [RNA-Seq]

city: Bethesda

first: Gary

last: Felsenfeld

address: 9000 Rockville Pike

institute: NIH

summary: The objectives of this study are to understand the regulatory roles of MAZ in biological processes using the NGS-deriveed ChIP-seq, DNA-MEDIP-seq and RNA-seq data in HAP1 control cells and MAZ knockout cells.  Our comparative analysis of these data generated from the HAP1 control and MAZ KO cells shows that MAZ is required for recruiting STAT1 to its target sites by reshaping epigenetic landscape in the human genome, thereby mediating antiviral response cells.

overall_design: Total RNA was extracted from HAP1 control and MAZ knockout cells treated with or without IFN-g. Differemtiated Gene expression levels between control and MAZ knockout cells were evaluated using DeqSeq2.

first: Tiaojiang

last: Xiao

first: Xin

last: Li


title: Discerning the roles of key cytokine signaling pathways in DR Mtb infection [dataset1]

status: Public on Apr 03 2024

city: Chicago

first: Shabaana

last: Khader

state: Illinois

address: 920 East 58th Street

institute: The University of Chicago

summary: We have found that drug-resistant (DR) Mtb infection alters the host pathogen interactions thought to occur during drug-sensitive (DS) Mtb infection. Recent data suggests that lack of both, Type I and IL-1, signaling pathways leads to susceptibility to infection to DR Mtb infection. To understand the pathways involved in maintaining control of DR Mtb infection, we are sequencing the bulk lung cells early in infection.

overall_design: RNA was generated from the lungs of various mice strains (wt, Ifnar-/-, Il1r1-/-, and Ifnar-/- Il1r1-/-) infected with DR Mtb for 14 days.

first: Bobba

last: Suhas


title: Neoantigen-Augmented iPSC-based Cancer Vaccine Combined with Local Radiotherapy Promotes Systemic Antitumor Immunity in Poorly Immunogenic Cancers

city: Taichung City

first: Kevin

middle: Chih-Yang

last: Huang

address: No. 91, Xueshi Road, North District

department: Department of Biomedical Imaging and Radiological Science

country: Taiwan

institute: China Medical University

summary: Although irradiated induced-pluripotent stem cells (iPSCs) as a prophylactic cancer vaccine elicit an antitumor immune response, the therapeutic efficacy of iPSC-based cancer vaccines is not promising due to their insufficient antigenicity and the immunosuppressive tumor microenvironment. Here, we found that neoantigen-engineered iPSC cancer vaccines can trigger neoantigen-specific T cell responses to eradicate cancer cells and increase the therapeutic efficacy of RT in poorly immunogenic colorectal cancer (CRC) and triple-negative breast cancer (TNBC). We generated neoantigen-augmented iPSCs (NA-iPSCs) by engineering AAV2 vector carrying murine neoantigens and evaluated their therapeutic efficacy in combination with radiotherapy. After administration of NA-iPSC cancer vaccine and radiotherapy, we found that ~60% of tumor-bearing mice achieved a complete response in microsatellite-stable CRC model. Furthermore, splenocytes from mice treated with NA-iPSC plus RT produced high levels of IFN secretion in response to neoantigens and had a greater cytotoxicity to cancer cells, suggesting that the NA-iPSC vaccine combined with radiotherapy elicited a superior neoantigen-specific T-cell response to eradicate cancer cells. The superior therapeutic efficacy of NA-iPSCs engineered by mouse TNBC neoantigens was also observed in the syngeneic immunocompetent TNBC mouse model. We found that the risk of spontaneous lung and liver metastasis was dramatically decreased by NA-iPSCs plus RT in the TNBC animal model. Altogether, these results indicated that autologous iPSC cancer vaccines engineered by neoantigens can elicit a high neoantigen-specific T-cell response, promote tumor regression and reduce the risk of distant metastasis in combination with local radiotherapy.

overall_design: Examination of expression and mutation-associated antigens (neoantigens) profile of murine (CT26) colorectal cancer cells treated with  iPSC cancer vaccine (iPSC-Vac). In the experiment, the impact of mouse irradiated iPSC cancer vaccine on neoantigen-bearing gene profiles was monitored. The experiment had 2 samples (CT26, CT26-iPSC-Vac). CT26 samples were used as the reference sample to compare. An analysis was also performed on RNA extracted from tumor samples (CT26 cells grown in the back of Balb/c mice, subcatenous injected with mouse irradiated iPSC cancer vaccine).

middle: C

first: William

middle: T

last: Chen

first: Jia-Yi

first: Chien-Yueh

last: Lee

first: Chia-Hsin

last: Wu

first: Chia-Ying

last: Lai

first: Pei-Chen

last: Yang

first: Ji-An

last: Liang

first: An‑Cheng

last: Shiau

first: K

middle: S

last: Chao

first: Tao-Wei

last: Ke


title: Discerning the roles of key cytokine signaling pathways in DS HN878 Mtb infection [dataset2]

summary: We have found that drug-resistant (DR) Mtb infection alters the host pathogen interactions thought to occur during drug-sensitive (DS) Mtb infection. Recent data suggests that lack of IL-1, but not Type I IFN, signaling pathways leads to susceptibility to infection to DR Mtb infection. To understand the pathways involved in maintaining control of DS Mtb infection, we are sequencing the bulk lung cells early in infection.

overall_design: Lung mRNA profiles of DS Mtb infected wt, Ifnar-/-, Il1r1-/-, and Ifnar-/- Il1r1-/- mice at 14 dpi


title: A rewiring of DNA replication mediated by MRE11 exonuclease underlies primed-to-naive cell de-differentiation

city: Madrid

first: Daniel

last: Gimenez

address: Calle de Melchor Fernández Almagro, 3

country: Spain

institute: CNIO

summary: Mouse embryonic stem cells (mESCs) in the primed pluripotency state, that resembles the post-implantation epiblast, can be de-differentiated in culture to anaivestate that resembles the pre-implantation inner cell mass.We report that primed-to-naive mESC transition entails a significant slowdown of DNA replication forks and the compensatory activation of dormant origins. Using iPOND (“isolation of proteins on nascent DNA”) coupled to mass spectrometry, we identify key changes in replisome composition that are responsible for these effects. Naive mESC forks are enriched in MRE11 nuclease and other DNA repair proteins. MRE11 is recruited to newly synthesized DNA in response to transcription-replication conflicts, and its inhibition or genetic downregulation in naive mESCs is sufficient to restore the fork speed observed in primed cells. Notably, MRE11 is required for the efficient primed-to-naive mESC transition, demonstrating a direct link between DNA replication dynamics and the mESC de-differentiation process

overall_design: RNA-Seq of untreated primed mESCs and after a 6-day 2i treatment, alone or in combination with two different concentrations of a MRE11 inhibitor.

first: Patricia

last: Ubieto-Capella

first: Pilar

last: Ximénez-Embún

last: Giménez-Llorente

first: Ana

last: Losada

first: Javier

last: Muñoz

first: Juan

last: Méndez


title: UBE2N function in skin homeostasis and immunity

city: Durham

first: Jennifer

middle: Yunyan

last: Zhang

state: NC

address: 3 Genome Ct, rm4130

department: Dermatology

institute: Duke University

summary: Epidermal keratinoyctes consitute the primary physical and biological barrier of the skin. The goal of this study is to determine the functions of keratinocyte UBE2N in regulation of skin homeostasis and immunity.

overall_design: We subjected C57BL6 WT and Rosa26CreER:Ube2nFlox/Flox mice back skin with topical treatments of 4-hydroxytamoxifen, and then collected WT and mutant skin samples for single-cell RNA-sequencing analysis.

first: Min

middle: J

first: Wanying

last: Miao

first: Vaibhav

last: Jain

first: Simon

last: Gregory

middle: Y


title: Linking CRISPR/Cas9 double-strand break profiles to gene editing precision with BreakTag [ampliseq 3]

city: Mainz

first: Sergi

last: Sayols

address: Ackermannweg 4

country: Germany

institute: Institute of Molecular Biology, Mainz

summary: Here we developed BreakTag, a versatile, highly parallel and scissile-aware methodology for the profiling of Cas9-induced DNA double strand break (DSBs), to identify molecular determinants influencing Cas9 incisions

overall_design: Map the off-target landscape of 1 sgRNAs targeting VEGFAsite2 with Streptococcus pyogenes (Sp)Cas9 using TTISS-seq

first: Gabriel

middle: M

last: Longo

first: Vassilis

last: Roukos


title: Linking CRISPR/Cas9 double-strand break profiles to gene editing precision with BreakTag [breaktag 2]

overall_design: Characterization of the scissile profile of 4 target regions with Cas12


title: Linking CRISPR/Cas9 double-strand break profiles to gene editing precision with BreakTag [ampliseq 2]

overall_design: Characterization of indels following Cas9-induced breaks in selected amplicons predicted to cut blunt or staggered.


title: Linking CRISPR/Cas9 double-strand break profiles to gene editing precision with BreakTag [ampliseq 2nd]

overall_design: Validation of bona fide off-targets nominated with BreakTag and or TTISS-seq via amplicon sequencing


title: Linking CRISPR/Cas9 double-strand break profiles to gene editing precision with BreakTag [breaktag 1]

overall_design: Map the off-target landscape and characterization of the scission profile of 1491 genomic gRNA-targets (HiPlex1 library) coupled with LZ3 using BreakTag


title: Linking CRISPR/Cas9 double-strand break profiles to gene editing precision with BreakTag [ampliseq 1]


title: Mapping the chromatin accessibility landscape of zebrafish embryogenesis at single-cell resolution by SPATAC-seq

status: Public on Apr 14 2024

city: beijing

first: Keyong

last: Sun

address: Hai Dian street

department: School of Medicine

country: China

institute: tsinghua university

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Zebrafish is a popular model for studies of embryonic development. Recent surveys of zebrafish embryogenesis with single-cell transcriptomics have uncovered hundreds of cell types, but the dynamic accessible regions of the genome that determine regulatory programs responsible for the unique identity and function of each cell type during embryogenesis lack detailed study. Here, we present SPATAC-seq: a split-pool ligation-based high-throughput single-cell assay for transposase-accessible chromatin using sequencing. Using SPATAC-seq, we profiled chromatin accessibility in more than 808,046 individual nuclei across 20 developmental stages spanning the sphere stage to the early larval protruding mouth stages in a single experiment. Using this chromatin accessibility map, we identified 604 cell types and inferred their developmental relationships. We also identified 959,040 candidate cis-regulatory elements (cCREs) and delineate development-specific cCREs, as well as transcription factors defining diverse cell identities. Importantly, transient reporter assays confirmed that a majority of tested cCREs exhibited robust enhanced EGFP expression in restricted cell types or tissues. Finally, we explored gene regulatory programs that drive pigment and notochord cell differentiation. Our work provides a comprehensive and valuable open resource for exploring transcriptional regulators that determine cell fate specification in zebrafish embryogenesis.

overall_design:
Wild-type zebrafish strain Tübingen (TU) line were maintained in standard zebrafish units at Core Facilities, school of life sciences, Tsinghua University. Naturally spawned fish embryos were collected at the 1-cell stage and the time of fertilization was used to stage each clutch. Embryos were maintained in Holtfreter’s solution at 28.5 °C for time course experiments and processed for bulk ATAC-seq and SPATAC-seq at the indicated times. Embryo nuclei of 20 developmental stages were extracted via previously described methods (Corces MR, et al. Nat Methods. 2017; Cao J, et al. Nature. 2019). The isolated nuclei were used for bulk ATAC-seq with TruePrep DNA Library Prep Kit V2 for Illumina (TD501, Vazyme), SPATAC-seq and 10x Genomics scATAC-seq (See more detailed method procedures from our paper). SPATAC-seq and bulk ATAC-seq libraries were sequenced with paired-end 150-bp reads on Illumina NovaSeq 6000 platform or DNBSEQ-T7. 10x Genomics scATAC-seq were sequenced with paired-end 100-bp reads on MGISEQ-2000RS, and i5 16 bp were linked at the end of Read 2.
Please note that the Series supplementary *bw files were genearated from all 535 samples together.

first: Xin

last: Liu

first: Anming

last: Meng

first: Xun

last: Lan


title: RNASeq, multiome, and genomic profiling of hematopoietic progenitors and B cells from mice with a point mutation in MYC [CUT&RUN]

city: Seattle

first: Brian

last: Freie

state: WA

address: 1100 Fairview Ave N

department: Basic Sciences

institute: Fred Hutchinson Cancer Research Center

summary: We generated mice with a mutation at threonine 58 of MYC in the mouse germline (T58A mutant). These mice ultimately develop AML or B cell lymphomas. We profiled changes in gene expression and genomic occupation of MYC in single cells of sorted, immature bone marrow progenitor cells, mature (spleen derived) and immature (marrow derived) B cells. B cells were stimulated with LPS and IL7, respectively.

overall_design: Lineage negative, ckit+, Sca1+ cells were sorted from marrow of WT or T58A littermate mice, and profiled for expression and DNA accessibility by single cell RNA-Seq and ATAC-Seq using the 10X genomic system. Bulk RNA-Seq was also performed on immature B cells isolated from bone marrow, and stimulated with IL7. RNA-Seq was also done on mature B cells isolated from spleens of these animals, and stimulated with lipopolysaccharide (LPS). Genomic profiling was done using CUT&RUN to determine MYC occupation, and histone modifications (H3K27Ac and H3K4Me2) were carried out in both the LPS and IL7 treated cells. For scRNA-Seq, 2 paired, biological replicate experiments (LSK_1 and LSK_2, comparing sorted cells from WT and T58A mutant mice) were done for each genotype. Each replicate consisted of LSK cells from 2 mice of each genotype, for a total of 4 mice per genotype for this experiment. Genomic profiling was done using CUT&RUN to determine MYC occupation, and histone modifications (H3K27Ac and H3K4Me2) were carried out in both the LPS and IL7 treated cells. For all genomic profiling experiments (CUT&RUN), 2 biological replicates per genoype (mice) were performed.

first: Bob

last: Eisenman


title: RNASeq, multiome, and genomic profiling of hematopoietic progenitors and B cells from mice with a point mutation in MYC [Multiome]

overall_design: Lineage negative, ckit+, Sca1+ cells were sorted from marrow of WT or T58A littermate mice, and profiled for expression and DNA accessibility by single cell RNA-Seq and ATAC-Seq using the 10X genomic system. Bulk RNA-Seq was also performed on immature B cells isolated from bone marrow, and stimulated with IL7. RNA-Seq was also done on mature B cells isolated from spleens of these animals, and stimulated with lipopolysaccharide (LPS). Genomic profiling was done using CUT&RUN to determine MYC occupation, and histone modifications (H3K27Ac and H3K4Me2) were carried out in both the LPS and IL7 treated cells. For scRNA-Seq, 2 paired, biological replicate experiments (LSK_1 and LSK_2, comparing sorted cells from WT and T58A mutant mice) were done for each genotype. Each replicate consisted of LSK cells from 2 mice of each genotype, for a total of 4 mice per genotype for this experiment. For all genomic profiling experiments (CUT&RUN), 2 biological replicates per genoype (mice) were performed.


title: Expression data from quadriceps of myofiber mineralocorticoid receptor conditional knockout (MRcko)/mdx and myeloid cell MRcko/mdx mice compared to Cre-/mdx control mice

city: Columbus

first: Jill

last: Rafael-Fortney

state: OH

address: 460 W. 12th Ave

department: Physiology & Cell Biology

institute: The Ohio State University

summary: To identify the gene expression differences in skeletal muscles resulting from conditional knockout of the mineralocorticoid receptor in myofibers and myeloid cells in dystrophin-deficient mdx mice

overall_design: 9 total samples were analyzed; 3 biological replicates for each Cre -/mdx control, myofiber MRcko/mdx, and myeloid cell MRcko/mdx groups; a pairwise comparison between groups was carried out and fold-changes ≥2 were selected.

first: Chetan

middle: K

last: Gomatam


title: Mitochondrial signals mediate paternal effects on offspring metabolism (scRNA-Seq)

city: München

first: Raffaele

last: Teperino

address: Ingolstädter Landstraße 1

department: Institute of Experimental Genetics (IEG)

institute: Helmholtz Zentrum München

summary: Spermatozoa deliver a complex and environment sensitive pool of small non-coding RNAs (sncRNA) to the oocyte at fertilisation, which influences offspring development and adult phenotypic trajectories. Whether mature spermatozoa in the epididymis can directly sense the environment is still not fully understood. Here, we used two distinct paradigms of preconception acute High Fat Diet challenge to dissect epididymal vs spermatogenic contributions to the sperm sncRNA pool and offspring health. We show that epididymal spermatozoa, but not developing germ cells, are sensitive to the environment and identify mitochondrial tRNA fragments as sperm-born sensors. In human spermatozoa, we found mt-tsRNAs in linear association with BMI and showed that paternal overweight at conception is sufficient to double offspring obesity risk and compromise metabolic health. Using mouse genetics and metabolic phenotypic data, we show that alterations of mt-tsRNAs are downstream of mitochondrial dysfunction in mice. Most importantly, single embryo transcriptomics of genetically hybrid two-cell embryos demonstrated  sperm-to-oocyte transfer of mt-tsRNAs at fertilisation and implied them in the control of early embryo metabolism.  Our study supports the importance of paternal health at conception for offspring metabolism, propose mt-tsRNAs as sperm-born environmental effectors of paternal inheritance and demonstrate, for the first time in a physiological and unperturbed setting, father-to-offspring transfer of sperm mt-tsRNAs at fertilisation.

overall_design: We analysed single cell transcriptome profile of testicular cells from 8 weeks old mice after 2 weeks of High fat diet (HFD) and Low fat diet (LFD) treatment


title: Mitochondrial signals mediate paternal effects on offspring metabolism (Smart-Seq)

summary: Spermatozoa deliver a complex and environment sensitive pool of small non-coding RNAs (sncRNA) to the oocyte at fertilisation [ref], which influences offspring development and adult phenotypic trajectories [refs]. Whether mature spermatozoa in the epididymis can directly sense the environment is still not fully understood [ref]. Here, we used two distinct paradigms of preconception acute High Fat Diet challenge to dissect epididymal vs spermatogenic contributions to the sperm sncRNA pool and offspring health. We show that epididymal spermatozoa, but not developing germ cells, are sensitive to the environment and identify mitochondrial tRNA fragments as sperm-born sensors. In human spermatozoa, we found mt-tsRNAs in linear association with BMI and showed that paternal overweight at conception is sufficient to double offspring obesity risk and compromise metabolic health. Using mouse genetics and metabolic phenotypic data, we show that alterations of mt-tsRNAs are downstream of mitochondrial dysfunction in mice. Most importantly, single embryo transcriptomics of genetically hybrid two-cell embryos demonstrated  sperm-to-oocyte transfer of mt-tsRNAs at fertilisation and implied them in the control of early embryo metabolism.  Our study supports the importance of paternal health at conception for offspring metabolism, propose mt-tsRNAs as sperm-born environmental effectors of paternal inheritance and demonstrate, for the first time in a physiological and unperturbed setting, father-to-offspring transfer of sperm mt-tsRNAs at fertilisation.

overall_design: Oocytes from C57BL/6N-mtST were fertilized with sperm from HFD or LFD treated C57BL/6N animals. Early 2cell embryos were collecetd after overnight incubation and snap frozen in lysis buffer according to SMARTseq2 protocol.


title: Mitochondrial signals mediate paternal effects on offspring metabolism (small RNA-Seq)

type: Non-coding RNA profiling by high throughput sequencing

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Small rna have been studied from cauda spermatozoa and spermatids from mice exposed to HFD and LFD.


title: Linking CRISPR/Cas9 double-strand break profiles to gene editing precision with BreakTag [indels]

overall_design: Characterization of indels following Cas9-induced breaks in selected amplicons containing a SNP in the protospacer.


title: Foxp striatal compensation

first: Genevieve

last: Konopka

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

overall_design: Refer to individual Series


title: Ptch1/Shh signaling control cochlear marginal cell differentiation and stria vascularis formation

city: Shatin

first: Karl Kam Hei

last: So

state: New Territories

address: 608A, Lo Kwee-Seong Integrated Biomedical Sciences Building, The Chinese University of Hong Kong

department: School of Biomedical Sciences

country: Hong Kong

institute: The Chinese University of Hong Kong

summary: The stria vascularis (SV) is an intricate non-sensory structure composed of multiple cell layers of different origins in the mammalian cochlea. It generates and maintains the endolymph, and therefore is essential for hearing functions. Here, using single cell transcriptomics and transgenic mouse models, we discover that the Shh receptor Ptch1 is required for marginal cell (MC, the epithelial cell layer of SV) differentiation and SV development. During cochlear development, MC differentiation gradually proceeds from cochlear base to apex, resembling the spatiotemporal differentiation pattern of sensory hair cells. Inactivation of Ptch1 leads to an impaired MC differentiation through elevated Gli2 levels, and inhibition of Gli2/Shh signaling activity is a prerequisite for MC differentiation. Single cell transcriptome analyses reveal that MCs are specified as early as E14. Moreover, in the absence of Ptch1, MC precursors are maintained in the progenitor state and differentiation is blocked consequently.

overall_design: Cochleae were dissected out from E16.5 WT and Emx2Cre;Ptch1f/f inner ears, digested into single cell suspension and subjected to 10X Genomics library construction and high-throughput sequencing

first: Karl

first: Tianli

last: Qin

first: Mai

middle: H

last: Sham


title: Mitochondrial signals mediate paternal effects on offspring metabolism (RNA-Seq)

overall_design: Study to understand transcriptional changes in germ cells and somatic tissues in male mice exposed to preconceptional High Fat Diet (HFD) and Low Fat Diet (LFD) for 2 weeks and their offspring.


title: Linking CRISPR/Cas9 double-strand break profiles to gene editing precision with BreakTag [cas9_variants]

first: Sergi

overall_design: Characterization of the scissile profile of seven previously described engineered variants with reduced off-target activity: HiFiCas9, xCas9, SniperCas9, HypaCas9, EvoCas9 and LZ3Cas9


title: Transcriptomic remodeling of gastric cells by Helicobacter pylori Outer Membrane Vesicles

city: Porto

first: Marina

last: Leite

state: Porto

address: Rua Alfredo Allen, 208

country: Portugal

institute: i3S - Instituto de Investigação e Inovação em Saúde

summary: Helicobacter pylori, a bacterium that colonizes the human stomach, like all Gram-negative bacteria spontaneously shed outer membrane vesicles (OMVs). OMVs, which act as a delivery system for bacterial components, are involved in bacterial-host interactions and thus contribute to pathogenesis. In this study, to understand the gene expression changes that human gastric epithelial cells might undergo when exposed to H. pylori-OMVs, we profiled the transcriptomic changes of the MKN74 gastric cell line induced by OMVs compared to control cells and H. pylori-infected cells, using the Ion AmpliSeq™ Transcriptome Human Gene Expression Panel. The top enriched pathways in the OMVs challenge condition included amino acid-related metabolic pathways, mitogen-activated protein kinase signaling, autophagy, and ferroptosis. The cell cycle, DNA replication, and repair pathways were the top diminished pathways. The transcriptomic changes induced by OMVs were largely consistent with those of the bacteria, although often at low expression levels, suggesting that their effects will mostly reinforce those of the bacterium itself. Our data provide a valuable portrayal of the transcriptomic remodeling of gastric cells by H. pylori-OMVs, which can be further dissected regarding the underlying molecular mediators and explored to understand the pathobiology of the full-spectrum of H. pylori-mediated diseases.

overall_design:
Transcriptomic analysis of MKN74 gastric cells upon exposure to OMVs isolated from Helicobacter pylori strain 26695 compared to saline-treated control and H. pylori-infected cells.
Submitter states that raw files are not available

first: Joana

last: Melo

first: Bruno

last: Cavadas

first: Luisa

last: Pereira

first: Ceu

last: Figueiredo


title: Linking CRISPR/Cas9 double-strand break profiles to gene editing precision with BreakTag [48sgRNA]

last: Sayols

overall_design: Map the off-target landscape of 46 sgRNAs targeting 12 clinically relevant genes with Streptococcus pyogenes (Sp)Cas9 using BreakTag

first: Gabriel

last: Longo

first: Vassilis

last: Roukos


title: Linking CRISPR/Cas9 double-strand break profiles to gene editing precision with BreakTag [hiplex3]

overall_design: HiPlex BreakTag dataset targeting 300 sites with SNPs targeting the reference and mutant allele (HiPlex3)


title: Linking CRISPR/Cas9 double-strand break profiles to gene editing precision with BreakTag [hiplex2]

overall_design: Characterization of the scissile profile of 1400 genomic gRNA-targets (HiPlex2 library) using BreakTag


title: Linking CRISPR/Cas9 double-strand break profiles to gene editing precision with BreakTag [hiplex1]

overall_design: Map the off-target landscape and characterization of the scissile profile of 1491 genomic gRNA-targets (HiPlex1 library) using BreakTag


title: Linking CRISPR/Cas9 double-strand break profiles to gene editing precision with BreakTag


title: RNASeq, multiome, and genomic profiling of hematopoietic progenitors and B cells from mice with a point mutation in MYC [RNA-seq]

first: Brian

last: Freie

first: Bob


title: Omental preadipocytes stimulate matrix remodeling and IGF signaling to support ovarian cancer metastasis

status: Public on Apr 02 2024

city: San Diego

first: Carrie

last: House

state: CA

address: 5500 Campanile Dr

institute: San Diego State University

summary: Ovarian cancer can metastasize to the omentum, which is associated with a complex tumor microenvironment. Omental stromal cells facilitate ovarian cancer colonization by secreting cytokines and growth factors. Improved understanding of the tumor supportive functions of specific cell populations in the omentum could identify strategies to prevent and treat ovarian cancer metastasis. Here, we showed that omental preadipocytes enhance the tumor initiation capacity of ovarian cancer cells. Secreted factors from preadipocytes supported cancer cell viability during nutrient and isolation stress and enabled prolonged proliferation. Co-culturing with pre-adipocytes led to upregulation of genes involved in extracellular matrix (ECM) organization, cellular response to stress, and regulation of insulin-like growth factor (IGF) signaling in ovarian cancer cells. IGF-1 induced ECM genes and increased alternative NF-κB signaling by activating RelB. Inhibiting the IGF-1 receptor (IGF1R) initially increased tumor omental adhesion but decreased growth of established preadipocyte-induced subcutaneous tumors as well as established intraperitoneal tumors. Together, this study shows that omental preadipocytes support ovarian cancer progression, which has implications for targeting metastasis.

overall_design: ACI-23 ovarian cancer cells were co-cultured with omental preadipocytes and differentiated lipid-storing adipocytes for 6 days. Co-cultures were set up to replicate the low cancer cell to high stromal cell density of early metastasis at a 1:40 ratio with 500 cancer cells and 20,000 preadipocytes/adipocytes. Co-cultures were performed using transwell chambers to allow the for exchange of secreted factors, and all RNA sequencing data was relativized to mono-culture controls.

first: Jennifer

last: Waters

first: Mikella

last: Robinson

first: Samuel

middle: F

last: Gilbert

middle: D


title: The retinoid X receptor has a critical role in synthetic rexinoid-induced increase in cellular all-trans-retinoic acid

city: New York

first: Laszlo

last: Halasz

state: NY

address: 1470 Madison Avenue

department: Department of Oncological Sciences

institute: Mount Sinai Icahn School of Medicine

summary: Rexinoids are agonists of nuclear rexinoid X receptors (RXR) that heterodimerize with other nuclear receptors to regulate gene transcription. A number of selective RXR agonists have been developed for clinical use but their application has been hampered by the unwanted side effects associated with the use of rexinoids and a limited understanding of their mechanisms of action across different cell types. Our previous studies showed that treatment of organotypic human epidermis with the low toxicity UAB30 and UAB110 rexinoids resulted in increased steady-state levels of all-trans-retinoic acid (ATRA), the obligatory ligand of the RXR-RAR heterodimers. Here, we investigated the molecular mechanism underlying the increase in ATRA levels using a dominant negative RXRα that lacks the activation function 2 (AF-2) domain. The results demonstrated that overexpression of dnRXRα in human organotypic epidermis markedly reduced signaling by resident ATRA, suggesting the existence of endogenous RXR ligand, diminished the biological effects of UAB30 and UAB110 on epidermis morphology and gene expression, and nearly abolished the rexinoid-induced increase in ATRA levels. Global transcriptome analysis of dnRXRα-rafts in comparison to empty vector-transduced rafts showed that over 95% of the differentially expressed genes in rexinoid-treated rafts constitute direct or indirect ATRA-regulated genes. Thus, the biological effects of UAB30 and UAB110 are mediated through the AF-2 domain of RXRα with minimal side effects in human epidermis. As ATRA levels are known to be reduced in certain epithelial pathologies, treatment with UAB30 and UAB110 may represent a promising therapy for normalizing the endogenous ATRA concentration and signaling in epithelial tissues.

overall_design: Transcriptomic profiling of control (CTR) and dominant negative RXRa (dnRXR) human skin rafts treated with UAB30 and UAB110 selective RXR agonists, using high throughput sequencing (Lexogen 3' QuantSeq).

first: Wilhelm

last: Berger

last: Nagy


title: Macrophage

first: Aderem

last: Lab

address: 307 Westlake Avenue North, Suite 500

institute: Seattle Children's Research Institute


title: the role of HPV11 E7 in modulating sting-dependent interferon β response in recurrent respiratory papillomatosis

city: 南京市

first: lijuan

last: chen

state: 江苏省

address: Nanjing City, 建邺区

institute: Nanjing BenQ hospital

summary: recurrent respiratory papillomatosis is a rare benign tumor caused mainly by the infection of the respiratory tract epithelial cells by the human papillamavirus type 6/11.the  specific mechanisms underlying the inhibition of the host's innate immune response by HPV remian unclear.For this purpose,we employed single cell RNA sequencing to analyze the states of various immune cells in RRP samples post HPV infection.

overall_design: four papilloma and three normal muscosal samples were subjected to transcriptome profiling by single cell RNAsequencing

first: yufei

last: pan

first: zhnekun

last: yu


title: Optimized RNA Interference Therapeutics Combined with Interleukin-2 mRNA for Treating Hepatitis B Virus Infection

city: Shanghai

first: Wenjing

last: Zai

state: 上海

address: Room 306, Fuxing Building, 131 Dongan Road, Shanghai 200032, China

institute: Fudan University

summary: BACKGROUND & AIMS: This study aimed to develop a pan-genotypic and multifunctional small interfering RNA (siRNA) against hepatitis B virus (HBV) with an efficient delivery system for treating chronic hepatitis B (CHB), and explore combined RNA interference (RNAi) and immune modulatory modalities for better viral control. METHODS: Twenty synthetic siRNAs targeting consensus motifs distributed across the whole HBV genome were designed and evaluated. The lipid nanoparticle (LNP) formulation was optimized by adopting HO-PEG2000-DMG lipid and modifying the molar ratio of traditional polyethylene glycol (PEG) lipid in LNP prescriptions. The efficacy and safety of this formulation in delivering siHBV (tLNP/siHBV) along with the mouse IL-2 (mIL-2) mRNA (tLNP/siHBVIL2) were evaluated in the rAAV-HBV1.3 mouse model. RESULTS: A siRNA combination (terms “siHBV”) with a genotypic coverage of 98.55% was selected, chemically modified, and encapsulated within an optimized LNP (tLNP) of high efficacy and security to fabricate a therapeutic formulation for CHB. The results revealed that tLNP/siHBV significantly reduced the expression of viral antigens and DNA (up to 3log10 reduction) in dose- and time-dependent manners at single-dose or multi-dose frequencies, with satisfactory safety profiles. Further studies showed that tLNP/siHBVIL2 enables additive antigenic and immune control of the virus, via introducing potent HBsAg clearance through RNAi and triggering strong HBV-specific CD4+ and CD8+ T cell responses by expressed mIL-2 protein. CONCLUSIONS: By adopting tLNP as nucleic acid nanocarriers, the co-delivery of siHBV and mIL-2 mRNA enables synergistic antigenic and immune control of HBV, thus offering a promising translational therapeutic strategy for treating CHB.

overall_design: HepG2 cells were incubated with tLNP/siNC or tLNP/siHBV at the dose of 50 nM for 16 h then applied for cDNA extraction and RNA sequencing. Mice were intravenously injected with tLNP/siNC or tLNP/siHBV at the dose of 1 mg/kg for 72 h, then liver samples were collected and snap frozen. RNA was extracted by TRIzol and cDNA libraries were prepared for following RNA sequencing, according to manufactures’ instructions.

first: Min

last: Yang

first: Kuan

last: Jiang

first: Juan

last: Guan

first: Huijing

last: Wang

first: Kongying

last: Hu

first: Chao

last: Huang

first: Jieliang

last: Chen

first: Wei

last: Fu

first: Changyou

last: Zhan

first: Zhenghong

last: Yuan


title: Evaluation of transcriptional changes following stimulation of naïve, TCM, TEM and TEMRA CD8+ T cell subsets with AB248 compared to  IL-2

city: South San Francisco

first: Manu

last: Kumar

state: California

address: 650 Gateway Blvd Suite 100

institute: Asher Biotherapeutics

summary: Evaluation of transcriptional changes following stimulation of naïve, TCM, TEM and TEMRA CD8+ T cell subsets confirmed that AB248 recapitulated the expected IL-2 signature with high fidelity

overall_design: Human PBMCs from 3 donors were sorted into the following subsets based on the expression of CD45RO and CD62L markers: CD8+-naïve T cells (CD62L+CD45RO-), CD8+ central memory T cells (CD62L+CD45RO+), CD8+ effector memory T cells (CD62L-CD45RO+), and CD8+ effector cells (CD62L-CD45RO-). Briefly, cells were stained with antibodies to sort the individual CD8+ T cell subsets, sorted via FACS, and flow sorted cells were collected in AIM V™ medium (Gibco) and then treated at concentrations corresponding to the EC99 by pSTAT5 for of rhIL-2 (R&D Systems) or AB248 for 24h. Briefly, 70,000 to 200,000 cells were added to 96 well plates in 100 μL AIM V medium (Thermo). At the end of stimulation, cells were pelleted, and RNA isolated per manufacturer’s instructions (Arcturus PicoPure RNA Isolation Kit, Applied Biosystems). RNA was sent to Genewiz for sequencing using Ultra-Low Input RNA-Seq protocol (detects ≥10 pg RNA). Sequencing data was aligned against the human reference genome (Ensembl GrCh38) using the STAR software package and quantified using the featureCounts package. Differential expression analysis was performed using the DESeq2 Bioconductor package.

first: Kelly

last: Moynihan

first: Jeffrey

last: Houser

first: Ruth

last: Lan

first: Yik

last: Yeung

first: Ivana

last: Djuretic


title: Redefining the molecular rejection states in 3230 heart transplant biopsies: relationships to parenchymal injury and graft survival

city: Edmonton

first: Jessica

last: Chang

address: 250 Heritage Medical Research Centre

department: Medicine

country: Canada

institute: University of Alberta

summary: The first-generation Molecular Microscope® (MMDx) system for heart transplant endomyocardial biopsies used expression of rejection-associated transcripts (RATs) to diagnose T cell-mediated rejection (TCMR) and antibody-mediated rejection (ABMR) but also detected acute injury. However, the ideal system should detect rejection without being influenced by injury, to permit analysis of the relationship between rejection and parenchymal injury. To achieve this, we developed a new rejection classification in an expanded cohort of 3230 biopsies: 1641 from INTERHEART (ClinicalTrials.gov #NCT02670408), plus 1589 service biopsies added to improve the power of the machine learning algorithms. The new system used six rejection classifiers instead of RATs and generated seven rejection archetypes: No Rejection 48%; Minor 24%; TCMR1 2.3%; TCMR2 2.7%; TCMR/Mixed 2.7%; EABMR 3.9%; and FABMR 16%. Using rejection classifiers eliminated cross-reactions with acute injury, permitting separate assessment of rejection and injury. TCMR was associated with severe recent injury and late atrophy-fibrosis and rarely had normal parenchyma. ABMR was better tolerated, seldom producing severe injury, but in later biopsies was often associated with atrophy-fibrosis, indicating long term risk. Graft survival and LVEF were reduced in hearts with TCMR, but also in hearts with severe-recent injury and atrophy-fibrosis, even without rejection.

overall_design: We developed a new rejection classification in an expanded cohort of 3230 biopsies: 1641 from INTERHEART (ClinicalTrials.gov #NCT02670408), plus 1589 service biopsies added to improve the power of the machine learning algorithms.


title: Modulation of the microRNA-induced silencing complex loading through a specific interaction with a J-domain co-chaperone

city: Québec

first: Pierre-Marc

last: Frédérick

state: Québec

address: 9 Rue McMahon

institute: Centre de recherche du CHU de Québec - Université Laval

summary: MicroRNAs (miRNAs) are essential regulators of several biological processes. To achieve their repressive function, they are loaded onto Argonaute (AGO) proteins, forming the microRNA Induced Silencing Complex (miRISC). Thus, regulating this loading step is essential for miRNA-dependent gene regulation. In this study, we identified the co-chaperone DNJ-12 as a new interactor of ALG-1, one of the two major miRNA-specific AGOs in Caenorhabditis elegans (C. elegans). Importantly, DNJ-12 does not interact with other AGOs, making it a specific regulator of miRNA function. Furthermore, the loss of DNJ-12 leads to developmental phenotypes previously shown to be dependent on miRNA function. Using the Auxin Inducible Degron (AID) system, a powerful tool to study the spatiotemporal effects of DNJ-12 depletion on miRNA function, we show that DNJ-12 depletion hampers ALG-1 interaction with HSP70, a chaperone required for miRISC loading in vitro. Moreover, DNJ-12 depletion leads to a decrease in the levels of several miRNAs and prevents their loading onto ALG-1. This study highlights for the first time the requirement of a co-chaperone for the loading of miRNAs in vivo, provides a potential mechanism explaining AGO differential loading by small RNA, and therefore helps expand our understanding of the miRISC function in animals.

overall_design: We compared the effect of DNJ-12 depletion on microRNA expression in different C. elegans tissues

first: Guillaume

last: Jannot

first: Isabelle

last: Banville

first: Martin

last: Simard


title: Dynamic Changes in Lymphocyte Populations Establish Zebrafish as a Thymic Involution Model

city: Oklahoma City

first: John

middle: Kimble

last: Frazer

state: Oklahoma

address: 941 Stanton L Young Blvd

department: Pediatrics

institute: University of Oklahoma Health Sciences Center

summary: The thymus is the site of T lymphocyte development and T cell education to recognize foreign, but not self, antigens. B cells also reside and develop in the thymus, although their function(s) are less clear. During ‘thymic involution,’ a process of lymphoid atrophy and adipose replacement linked to sexual maturation, thymic cells decline. However, thymic B cells decrease far less than T cells, such that B cells comprise ~1% of neonatal thymocytes, but up to ~10% in adulthood in humans. All jawed vertebrates possess a thymus, and we and others have shown that zebrafish (Danio rerio) also have thymic B cells. Here, we investigated the precise identities of zebrafish thymic T and B cells and how they change with involution. We assessed the timing and specific details of zebrafish thymic involution using multiple lymphocyte-specific, fluorophore-labeled transgenic lines, quantifying changes in thymic lymphocytes pre- vs. post-involution. Our results prove that, as in humans, zebrafish thymic B cells increase relative to T cells post-involution. We also performed RNA sequencing (RNA-seq) on D. rerio thymic and marrow lymphocytes of four novel double-transgenic lines, identifying distinct populations of immature T and B cells. Collectively, this is the first comprehensive analysis of zebrafish thymic involution, demonstrating its similarity to human involution, and establishing the highly genetically-manipulatable zebrafish model as a template for involution studies.

overall_design: This experimental study aimed to investigate the gene expression profiles of lymphocytes in zebrafish using bulk RNA sequencing (RNA-seq) analysis. Specifically, four different double-transgenic zebrafish lines were utilized, each expressing specific fluorescent markers to label distinct cell populations. The double-transgenic lines included rag2:RFP × cd79a:GFP, rag2:RFP × cd79b:GFP, cd79a:GFP × lck:mCherry, and cd79b:GFP × lck:mCherry. These transgenic lines allowed for the identification and isolation of specific cell types for subsequent transcriptomic analysis.

first: Ameera

last: Hasan

first: Jose

last: Macias

first: Brashe

last: Wood

first: Megan

last: Malone-Perez

first: Gilseung

last: Park

first: Clay

last: Foster

first: J


title: Single-cell RNA sequencing of T3 sarcoma tumors following treatment with CD8-mIL2, CTRL-not-alpha-mIL2, or anti-PD-1

first: Manu

last: Kumar

summary: To assess treatment-induced changes of intratumoral CD8+ T cells in an unbiased manner, CD8+ TILs from T3 sarcomas were analyzed using single-cell RNA-sequencing (scRNAseq) 2 or 4 days after therapy with CD8-mIL2, CTRL-not-α-mIL2, anti-PD-1, or combinations thereof. Barcoded peptide:MHC reagents were included to identify mLama4-specific CD8+ T cells. CD8-mIL2 therapy promotes the generation of a new stem-like effector CD8+ T cell cluster with a signature indicative of polyfunctionality and with features of T cell memory.

overall_design: T3 tumor-bearing mice were treated on day twelve post-tumor transplant. Two and four days later, single-cell suspensions were prepared from TILs. Total CD8+ T cells were enriched using a CD8+ T cell positive selection kit (Miltenyi). Tumors from individual mice were labeled with one of the following antibodies before pooling in order to identify cells from individual tumors in downstream analysis: 1) TCR (TotalSeq™-C0120 anti-mouse TCR β chain antibody, Cat# 109259, BioLegend), 2) CD90.2 (TotalSeq™-C0075 anti-mouse CD90.2 (Thy1.2) antibody, Cat#, 105353, BioLegend), and 3) CD45 (TotalSeq™-C0096 anti-mouse CD45 antibody, Cat# 103169, BioLegend). Samples were submitted to the Genome Institute at Washington University in St. Louis to generate 10x libraries using 10x 5'v2 Single Cell RNAseq, V(D)J enrichment, feature barcoding, and Barcode Enabled Antigen Mapping kits.   Fastq files were processed using the Cell Ranger (v7.1.0) multi pipeline with the default parameters. Gene expression libraries were aligned to the mouse transcriptome (mm10-2020-A) and TCR sequences were aligned to the VDJ reference dataset (GRCm38-alts-ensembl-5.0.0).

first: Kelly

last: Moynihan

first: Hussein

last: Sultan

first: Robert

last: Schreiber


title: Iron boosts anti-tumor type 1 T-cell responses and anti-PD1 immunotherapy

city: Paris

first: Bruno

last: Martin

state: Île-de-France

address: 22, rue Mechain

country: France

institute: Institut Cochin, INSERM U1016, CNRS UMR 8104

summary: Cancers only develop if they escape immunosurveillance, and the success of cancer immunotherapies relies in most cases on their ability to restore effector T-cell functions, particularly IFN-γ. Revolutionizing the treatment of many cancers, immunotherapies targeting immune checkpoints such as PD1 may increase survival and cure patients. Unfortunately, although immunotherapy has greatly improved the prognosis of patients, not all respond to anti-PD1 immunotherapy, making it crucial to identify alternative treatments that could be combined with current immunotherapies to improve their effectiveness. Here, we show that iron supplementation significantly boosts T-cell responses in vivo and in vitro. Such a phenomenon could be the consequence of a metabolic reprogramming of T cells by iron. The "adjuvant" effect of iron results in a strong slowdown of tumor-cell growth after tumor-cell line transplantation in mice. Specifically, our results suggest that iron supplementation promotes anti-tumor responses by increasing IFN-γ production by T cells. In addition, iron supplementation considerably improves the efficacy of anti-PD1 cancer immunotherapy in mice. Finally, our study suggests that, in cancer patients, the quality and efficacy of the anti-tumor response following anti-PD1 immunotherapy may be modulated by plasma ferritin levels. In summary, our results suggest the benefits of iron supplementation on the reactivation of anti-tumor responses and support the relevance of a fruitful association between immunotherapy and iron supplementation.

overall_design: To understand how iron may stimulate T-cell responses, we compared the transcriptome of naive CD4+ T cells from peripheral lymph-nodes of C57BL/6 mice, stimulated for 20 hours with coated anti-CD3 and anti-CD28 antibodies in the presence or absence (Ctrl.) of iron supplementation to cell-culture (FAC). 4 ctlr. samples vs. 4 iron supplemented samples have been compared.

last: Lucas


title: The unusual mode of action of the polyketide glycoside antibiotic cervimycin C [I]

city: Bonn

first: Alina

last: Hoffmann

address: Venusberg Campus 1

institute: University Hospital Bonn

summary: The resistance to antibiotics is an emerging problem, and necessitates novel antibacterial therapies. Cervimycins A–D are bi-glycosylated polyketides produced by Streptomyces tendae HKI 0179 with promising activity against Gram-positive bacteria. Microscopically, cervimycin C (CmC) treatment caused a spaghetti-like phenotype in Bacillus subtilis 168, with elongated curved cells, which stayed joined after cell division, and exhibited a chromosome segregation defect. Electron microscopy of cervimycin treated S. aureus revealed swelling of some cells, misshaped septa, and cell wall thickening and a rough cell wall surface. Incorporation tests in B. subtilis indicated an effect on DNA metabolism at high cervimycin-concentrations. Indeed, the down-regulation of the DNA gyrase subunit B (gyrB) acted synergistically with cervimycin, and the antibiotic inhibited the in vitro DNA gyrase supercoiling activity. To get a more global view on the mode of action of CmC, transcriptomics and proteomics of cervimycin treated versus untreated S. aureus cells were performed. Interestingly, and in contrast to the previous results, cervimycin did not induce the SOS response in S. aureus, which would indicate disturbance of the DNA gyrase. Instead, cervimycin induced the expression of the CtsR/HrcA heat shock operon and the expression of autolysins. Taken together, we identified the DNA gyrase as one target of cervimycin, but omics data revealed massive alterations in cervimycin treated S. aureus, involving cell wall modifying enzymes and protein stress response, indicating a complex mode of action of cervimycin, that is distinct from other antibiotics.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for S. aureus SG511 cells treated with three-times minimal inhibitory concentration cervimycin C versus untreated cells.

first: Ursula

last: Steffens

first: Boris

last: Macek

first: Mirita

last: Franz-Wachtel

first: Kay

last: Nieselt

first: Theresa

last: Harbig

first: Kirstin

last: Scherlach

first: Christian

last: Hertweck

first: Hans-Georg

last: Sahl

first: Gabriele

last: Bierbaum


title: The unusual mode of action of the polyketide glycoside antibiotic cervimycin C

overall_design: Refer to individual Series


title: Single-cell transcriptional profiling of wildtype Rosa26(YFP) (Y) and Kras(G12D) p53(-/-) Rosa26-YFP (KPY) lung AT2 epithelial cells, AT2 organoids, and co-cultured mesenchymal cells with temporal resolution

city: Boston

first: Aaron

middle: L.

last: Moye

state: MA

address: 1 Blackfan Circle

department: Stem Cell Program

institute: Boston Children's Hospital

summary: The aim of this study was to examine the transcriptional dynamics of AT2 progenitor cells and their surrounding mesenchyme in lung adenocarcinoma across model systems, using temporal and single-cell resolution.

overall_design: Examination of the progression of wildtype Rosa26(YFP) and Kras(G12D) p53(-/-) Rosa26(YFP) AT2 cells in organoids, their co-cultured wildtype mesenchymal cells, and GEMMs at various time points after induction using Ad-Cre.

middle: L

first: Carla

last: Kim


title: IL-6-mediated endothelial injury impairs anti-viral humoral immunity

status: Public on Apr 01 2024

first: Geoffrey

middle: R

last: Hill

address: 1100 Fairview Ave N

institute: Fred Hutch Cancer Center

summary: Endothelial function and integrity are compromised after allogeneic bone marrow transplantation (BMT) but how this affects immune responses broadly remains unknown. Using a preclinical model of cytomegalovirus (CMV) reactivation after BMT, we found compromised antiviral humoral responses induced by IL-6 signaling. IL-6 signaling in T cells maintained Th1 cells resulting in sustained IFN secretion which promoted endothelial cell (EC) injury, loss of the neonatal Fc receptor (FcRn) responsible for immunoglobulin G (IgG) recycling, and rapid IgG loss. In contrast, recipient IgG producing cells are rapidly eliminated after BMT. T cell-specific deletion of IL-6R led to persistence of recipient-derived CMV-specific IgG and inhibited CMV reactivation. Deletion of IFN in donor T cells also eliminated EC injury and FcRn loss. In a phase III clinical trial, blockade of IL-6R with tocilizumab promoted CMV-specific IgG persistence and significantly attenuated early HCMV reactivation. In sum, IL-6 invokes IFN-dependent EC injury and consequent IgG loss leading to CMV reactivation. Hence, cytokine inhibition represents a logical strategy to prevent endothelial injury thereby preserving humoral immunity after immunotherapy.

overall_design: Non-infected B6D2F1 recipients were transplanted with WT TCD BM (5x106) ± T cells (2x106) from WT or IFNg ko donors, thus there are 3 groups: TCD BM, WT BM+T and IFNg KO BM+T. Endothelial cells were isolated from liver on day +7, pooled from 3 mice per group, FACS sort to CD31+ and processed for single cell RNAseq.

first: Ping

last: Zhang

first: Olivia

middle: G

last: Waltner

first: Shruti

last: Bhise

first: Scott

middle: N

last: Furlan

first: Simone

last: Minnie


title: The comparison of iPSCs and fibroblasts in Mus minutoides

city: Kagoshima

first: Sumito

last: Matsuya

address: 1-21-24 Korimoto

department: Joint Faculty of Veterinary Medicine

country: Japan

institute: Kagoshima university

summary: Compare gene expression status of iPSCs and fibroblasts in Mus minutoides.

overall_design: We performed RNA-seq analysis on Mus minutoides iPSCs and fibroblasts, two samples each.

first: Kaoru

last: Fujino

first: Hiroyuki

last: Imai

first: Ken

last: Kusakabe

first: Wataru

last: Fujii

first: Kiyoshi

last: Kano


title: Discovery and Validation of a 15-Gene Prognostic Signature for Clear Cell Renal Cell Carcinoma

city: Ann Arbor

first: Simpa

last: Salami

state: MI

address: 1500 E Medical Center Dr

department: Urology

institute: University of Michigan

summary: We report a kidney cancer tissue-based prognostic biomarker encompassing 15 genes (15G score) to classify patients into low versus high risk for recurrence after curative nephrectomy. The 15G score was independently associated with disease free survival adjusting for clinicopathologic variables as well as existing clinical risk calculators or nomograms. By improving risk stratification of patients with ccRCC, the 15G score has the capacity to facilitate selection of biopsy confirmed small renal cancers (T1a) for treatment versus surveillance; inform intensity and duration of surveillance after curative nephrectomy; and to potentially facilitate patient selection for adjuvant systemic therapy. We retrospectively identified 110 patients who underwent radical nephrectomy for ccRCC (discovery cohort). Patients who recurred were matched based on grade/stage to patients without recurrence. Capture whole transcriptome sequencing was performed on RNA isolated from archival tissue using the Illumina platform. We developed a gene-expression signature to predict recurrence/disease-free survival (DFS) using a 15-fold lasso and elastic-net regularized linear Cox model. We derived the 31-gene cell cycle progression (mxCCP) score using RNAseq data for each patient. Kaplan-Meier (KM) curves and multivariable Cox proportional hazard testing were used to validate the independent prognostic impact of the gene-expression signature on DFS, disease specific survival (DSS) and overall survival (OS) in two validation datasets (combined n=761).

overall_design: We developed and validated a novel 15G prognostic signature to improve risk stratification of patients with ccRCC. Pending further validation, this signature has the potential to facilitate optimal treatment allocation.

first: Rohit

last: Mehra

first: Srinivas

last: Nallandhighal

first: Brittney

last: Cotta

first: Zayne

last: Knuth

first: Fengyun

last: Su

first: Amy

last: Kasputis

first: Yuping

first: Rui

last: Wang

first: Xuhong

last: Cao

last: Udager

first: Saravana

last: Dhanasekaran

first: Marcin

middle: P

last: Cieslik

first: Todd

last: Morgan


title: Trascriptional profiling of proliferating and resting fracntions in ibrutinib-treated CLL

city: Aviano

first: Federico

last: Pozzo

state: PN

address: via Franco Gallini 2

department: Department of Translational Research

country: Italy

institute: Centro di Riferimento Oncologico (CRO) di Aviano IRCCS

summary: Bruton’s tyrosine kinase (BTK) inhibitors such as ibrutinib represent an effective strategy for treatment of chronic lymphocytic leukemia (CLL), although ~30% of patients eventually undergo disease progression. Here we investigated the long-term modulation of the CXCR4dim/CD5bright proliferative fraction (PF) and the CXCR4bright/CD5dim resting fraction (RF) in CLL samples, and their correlation with therapeutic outcome and emergence of ibrutinib resistance. Longitudinal tracking by flow cytometry revealed that PF, initially suppressed by ibrutinib, reappeared upon early disease progression suggesting that PF evaluation could represent a sensitive and specific marker of CLL progression upon ibrutinib treatment. Transcriptomic analyses of PF at progression revealed similar proliferation signatures between pre- and post-treatment PF, demonstrating the emergence upon progression of a newly proliferating cell population.

overall_design: PF and RF fractions were cell-sorted from thawed PBMCs and nuclei acids immediately purified. 30-50ng of RNA were processed using mRNA library prep kit

first: Valter

last: Gattei

first: Riccardo

last: Bomben

first: Filippo

last: Vit


title: The unusual mode of action of the polyketide glycoside antibiotic cervimycin C [II]

first: Alina

last: Hoffmann

summary:
Cervimycins A-D are bis-glycosylated polyketide antibiotics produced by Streptomyces tendae HKI 0179 with bactericidal activity against Gram-positive bacteria. In this study, cervimycin C (CmC) treatment caused a spaghetti-like phenotype in Bacillus subtilis 168, with elongated curved cells, which stayed joined after cell division, and exhibited a chromosome segregation defect, resulting in ghost cells without DNA. Electron microscopy of cervimycin treated S. aureus (3 x MIC) revealed swollen cells, misshapen septa, cell wall thickening, and a rough cell wall surface. Incorporation tests in B. subtilis indicated an effect on DNA biosynthesis at high cervimycin concentrations. Indeed, artificial down-regulation of the DNA gyrase subunit B gene (gyrB) increased the activity of cervimycin in agar diffusion tests, and, in high concentrations (starting at 62.5 x MIC), the antibiotic inhibited S. aureus DNA gyrase supercoiling activity in vitro. To obtain a more global view on the mode of action of CmC, transcriptomics and proteomics of cervimycin treated versus untreated S. aureus cells were performed. Interestingly, 3 x MIC of cervimycin did not induce the SOS response, which would indicate disturbance of the DNA gyrase activity in vivo. Instead, cervimycin induced the expression of the CtsR/HrcA heat shock operon and the expression of autolysins, exhibiting similarity to the ribosome-targeting gentamicin. In summary, we identified the DNA gyrase as a target, but at low concentrations electron microscopy and omics data revealed a more complex mode of action of cervimycin, that comprised induction of the heat shock response, indicating protein stress in the cell.

A synergistic effect was seen when sodium chloride (NaCl) was added during cervimycin MIC testing of S. aureus SG511. Thus, transcriptomics of S. aureus cells under osmotic stress (1 M NaCl) were performed. The stress response was compared to the cervimycin stress response to identify factors which might contribute to the synergistic antibacterial effect of the two molecules.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for S. aureus SG511 cells grown in TSB supplemented with 1 M NaCl versus cells grown in TSB without NaCl.

first: Ursula

last: Steffens

first: Boris

last: Macek

first: Mirita

last: Fanz-Wachtel

first: Kay

last: Nieselt

first: Theresa

last: Harbig

first: Kirstin

last: Scherlach

first: Christian

last: Hertweck

first: Hans-Georg

last: Sahl

first: Gabriele

last: Bierbaum


title: Effect of Rosa roxburghii fruit extracts to gene expression of mesenchymal stem cells

city: Guangzhou

first: Yan

last: Huang

address: No. 135, XIngang Street

institute: Sun Yat-sen University

summary: Here we found Rosa roxburghii fruit extracts effectively increase TERT expression and telomerase activity in cultured human mesenchymal stem cells. Both Rosa roxburghii fruit extracts by freeze drying and spray drying methods increase the activity of telomerase. Rosa roxburghii fruit freeze drying extracts is able to reduce reactive oxygen species levels, enhance SOD activity and resistance to oxidative stress, and reduce DNA damage caused by oxidative stress or radiation. Rosa roxburghii fruit extracts promoted cell proliferation, improved senescent cell morphology, delayed replicative cellular senescence, attenuated cell cycle supressors and alleviated the senescence-associated secretory phenotype. Transcriptome and metabolic profilings found that Rosa roxburghii fruit extract promote cell proliferation and DNA repair pathways, decreased triglycerides as well. Overall, we provided a theoretical basis for the application of Rosa roxburghii fruit as an anti-aging natural product.

overall_design: To investigate the role of Rosa roxburghii fruit extracts to gene expression of mesenchymal stem cells, we applied Rosa roxburghii fruit extracts to the medium. 3 repeats were done.


title: Pro-oxidant therapy by a vitamin K precursor targets PI 3-Kinase VPS34 function

city: Dunedin

first: Sarah

last: Diermeier

address: 710 Cumberland Street

department: Biochemistry

country: New Zealand

institute: University of Otago

summary: Men taking anti-oxidant vitamin E supplements have increased prostate cancer (PC) risk. Whether pro- oxidants protect from PC remained unclear. We show that a pro-oxidant vitamin K precursor (MSB) suppresses PC progression in mice killing cells through an oxidative cell death: MSB antagonizes the essential class III PI 3-Kinase VPS34 the regulator of endosome identity and sorting through oxidation of key cysteines pointing to a redox checkpoint in sorting. Testing MSB in a myotubular myopathy model that is driven by loss of MTM1 the phosphatase antagonist of VPS34 we show that dietary MSB improved muscle histology function and extended life span. These findings enhance our understanding of pro-oxidant selectivity and show how definition of the pathways they impinge on can give rise to unexpected therapeutic opportunities.

overall_design: Mouse embryonic fibroblasts (p53 -/- PTEN -/-) were treated with MSB (0, 10, 20 µM) or erastin (0, 0.15, 0.3 µM), in four replicates. RNA was extracted and transcriptome profiles generated.

first: Manojit

last: Swamynathan

first: Lloyd

middle: C

last: Trotman


title: Single cell RNA sequencing analysis of vestibular schwannoma reveals functionally distinct macrophage subsets

city: Birmingham

first: Adam

last: Croft

state: Please Select

address: Birmingham

country: United Kingdom

institute: University of Birmingham

summary: Background: Vestibular schwannomas (VS) remain a challenge due to their anatomical location and propensity to growth. Macrophages are present in VS but their roles in VS pathogenesis remains unknown. Objectives: Assess phenotypic and functional profile of macrophages in VS with single cell RNA sequencing (scRNAseq). Methods: scRNAseq was carried out in three VS samples to examine characteristics of macrophages in the tumor. RT-qPCR was carried out on ten VS samples for CD14, CD68 and CD163 and a panel of macrophage associated molecules. Results: scRNAseq revealed macrophages to be a major constituent of VS microenvironment with three distinct subclusters based on gene expression.  The subclusters were also defined by expression of CD163, CD68 and IL-1β. AREG and PLAUR were expressed in the CD68+CD163+IL-1β+ subcluster, PLCG2 and NCKAP5 were expressed in CD68+CD163+IL-1β- subcluster and AUTS2 and SPP1 were expressed in the CD68+CD163-IL-1β+ subcluster. RT-qPCR showed expression of several macrophage markers in VS of which CD14, ALOX15, Interleukin-1β, INHBA and Colony Stimulating Factor-1R were found to have a high correlation with tumor volume. Conclusions: Macrophages form an important component of VS stroma. scRNAseq reveals three distinct subsets of macrophages in the VS tissue which may have differing roles in the pathogenesis of VS.

overall_design: single cell RNA sequencing analysis of vestibular schwannoma

first: Chris

last: Mahony

first: Paramita

last: Baruah


title: The epigenetic regulator BAP1 is required before birth for differentiation and maintenance of the postnatal enteric nervous system in mice.

city: Philadelphia

first: Katherine

last: Beigel

state: PA

address: 2716 South Street

department: Department of Biomedical and Health Informatics

institute: Children's Hospital of Philadelphia

summary: Epigenetic regulatory mechanisms are underappreciated but critical for enteric nervous system (ENS) development and maintenance. We discovered that fetal loss of the epigenetic regulator Bap1 in the ENS lineage causes severe postnatal bowel dysfunction and early death in Tyrosinase-Cre; Bap1fl/fl mice. Bap1-depleted ENS appears normal in neonates, however, by postnatal day 15 (P15), Bap1-deficient enteric neurons are largely absent from the small and large intestine of Tyrosinase-Cre; Bap1fl/fl mice. Bowel motility becomes markedly abnormal with disproportionate loss of cholinergic neurons. Single-cell RNA sequencing at P5 shows that fetal Bap1 loss inTyrosinase-Cre; Bap1fl/fl mice markedly alters the composition and relative proportions of enteric neuron subtypes. In contrast, postnatal deletion of Bap1 did not cause enteric neuron loss or impaired bowel motility. These findings suggest that BAP1 is critical for postnatal enteric neuron differentiation and for enteric neuron survival.

overall_design: Whole cell enteric neuron isolation from P5 mouse colon was performed by extracting the muscularis layer from full-length colons from P5 Bap1 fl/fl; Tyr-Cre +; R26 TdT/TdT and Bap1 wt/wt; Tyr-Cre +; R26 TdT/TdT pups. P5 Tyr-Cre lineage cells from the colon muscularis were sorted using fluorescence-activated cell sorting (FACS) for presence of TdTomato signal and analyzed using scRNAseq.

first: Sabine

last: Schneider

first: Jessica

middle: B

last: Anderson

first: Rebecca

last: Bradley

first: Christina

last: Wright

first: Beth

middle: A

last: Maguire

first: Guang

last: Yan

first: Deanne

last: Taylor

first: J

middle: W

last: Harbour

first: Robert

middle: O

last: Heuckeroth


title: Chromatin landscape instructs precise transcription factor regulome during embryonic lineage specification

first: Liping

last: Wang

address: 1239 Siping Road

institute: Tongji University

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


title: Circulating microRNAs in association with pancreatic cancer risk within 5 years

city: Nashville

first: Cong

state: TN

address: 2525 West End Ave

institute: Vanderbilt University

summary: miRNAs levels were measured in plasma samples of pancreatic cancer cases and controls collected within 5 years prior to diagnosis using the NanoString nCounter Human v3 miRNA expression panel

overall_design: A two-stage study of pancreatic cancer cases and controls with plasma samples within 5 years prior to diagnosis to identify miRNAs in association with pancreatic cancer risk.


title: Single-nucleus RNA-seq identifies one galanin neuronal subtype in mouse preoptic hypothalamus activated during recovery sleep after sleep deprivation

first: Allan

middle: I

last: Pack

address: 125 South 31st St

institute: University of Pennsylvania

summary: The preoptic area of hypothalamus plays an important role in sleep homeostasis. We used snRNA-seq to identify neurons involved in homeostatic sleep response.

overall_design: We compared mice under high sleep pressure (recovery sleep following sleep deprivation) to mice under low sleep drive (undisturbed sleep toward the end of lights-on).

first: Xiaofeng

last: Guo

first: Brendan

last: Keenan

first: Benjamin

last: Reiner

first: Jie

last: Lian


title: Analysis of a non-obese diabetes-experimental autoimmune encephalomyelitis (NOD-EAE)

city: Takatsuki, Murasaki-cho

first: Toshiki

last: Yoshimizu

state: Osaka

institute: Japan Tobacco inc.

summary: The purpose of this study was to analyze and elucidate the mechanisms of non-obese diabetes-experimental autoimmune encephalomyelitis (NOD-EAE), an animal model of progressive multiple sclerosis (MS), and to compare the pathological features with those observed in human progressive MS. To achieve this, pathological analysis, flow cytometry analysis, immunohistochemical staining, and transcriptome analysis were performed at each pathological stage of the NOD-EAE mice to characterize each pathological stages in the lesion. In the chronic phase of the NOD-EAE mice, fibrosis and lymph follicle formation, characteristic of progressive human MS, were observed. We describe the pathological profile and transcriptome analysis of the NOD-EAE mice and verify that this model has similar features to those of human progressive MS. Our findings suggest that this model recapitulates lymph follicle formation, a disease hallmark of progressive MS, and fibrosis, a feature complicating the pathogenesis of MS in the chronic phase.

overall_design: To compare the NOD-EAE stage, we performed gene expression profiling analysis using data obtained from RNA-seq of 5 different disease phases, normal, 1st peak, remission, relapse, and chronic.

first: Satoshi

last: Hamano

first: Toshihide

last: Yamashita


title: Single-cell gene expression profiling of CD4+ T cells from the murine small intestine lamina propria [steady-state_SI-Treg_10XscRNAseq]

city: Oberschleißheim

first: Caspar

last: Ohnmacht

address: Ingolstädter Landstraße 1

department: Institute for Allergy Research

institute: Helmholtz Munich

summary: Intestinal regulatory T cells consist of at least two distinct populations in flow cytometry. We used single-cell RNA sequencing to determine the transcriptional differences in peripherally and thymically induced Tregs.

overall_design: We sorted all living CD4+ T cells (CD45+, CD3+, CD4+) from small intestine lamina propria of four wildtype mice via fluorescence-activated cell sorting.

first: Amelie

last: Köhler


title: Single-cell gene expression profiling of CD4+ T cells from small intestine lamina propria of mixed Bcl3-/- CD45.2+ and WT CD45.1+ bone marrow recipients [Mixed-bone-marrow_10XscRNAseq]

summary: Loss of Bcl3 influences the abundance and functionality of regulatory T cells in several organs but especially in the intestine. We used single-cell RNA sequencing (scRNAseq) to determine the influence of Bcl3 on cellular basis in a competitive environment.

overall_design: We sorted all living CD4+ T cells (CD45+, CD3+, CD4+) from small intestine lamina propria of four mixed Bcl3-/- CD45.2+ and WT CD45.1+ bone marrow recipients via fluorescence-activated cell sorting.


title: Loss of Bcl3 influences gene expression in Tregs and Treg subsets [Bcl3-WT_Treg_bulkRNAseq]

summary: Peripherally induced regulatory T cells (pTregs) expressing the retinoic acid receptor-related orphan-receptor gamma t (RORγt) are indispensable for intestinal immune homeostasis. Previous studies have determined the role of the atypical NFκB inhibitor Bcl3 in the development of colitis. In this study we analyzed the influence of Bcl3 on pTreg development and functionality in healthy mice and under colitogenic conditions. Our findings reveal that expression of Bcl3, in health, limits not just the development of pTregs in a T cell intrinsic manner but also the formation of a RORγt+Helios+ double positive subset (DPTreg) expressing an activated transcriptional profile. Consequently, loss of Bcl3 in Tregs does not impede their suppressive capacity in a model of T cell transfer colitis but moreover leads to increased production of anti-inflammatory cytokines IL-10 and TGFβ. We further demonstrate that lack of Bcl3 in Tregs results in trans-differentiation towards Th17-like cells. Finally, we provide a Bcl3 dependent gene signature in pTregs including altered signaling of the cytokines IL-2, IL-6 and TNFα. We therefore conclude that expression of Bcl3 regulates the sensitivity of Tregs towards homeostatic cytokines and consequently limits the formation of an unphysiological number of RORγt+ Tregs.

overall_design:
Cells from mesenteric lymph nodes (mLN) and spleen were isolated from wildtype mice (WT), Bcl3-/- Bcl3 deficient mice (KO) and mice conditionally overexpressing Bcl3 in Tregs (OE). Via fluorescence-activated cell sorting (FACS) we isolated CD45+CD3+CD4+Foxp3+ Tregs and CD45+CD3+CD4+Foxp3- Thelper cells (Th).
We FACS sorted Foxp3+ T cells (CD45+, CD3+, CD4+) from mLN, spleen and small intestine lamina propria (SI) of Bcl3 sufficient and Bcl3 deficient mice, both with reporters for Foxp3-RFP and RORgt-GFP. We sorted peripherally induced Foxp3+RORgt+ pTregs and thymically induced Foxp3+RORgt- tTregs.


title: Human γδ T cells in diverse tissues exhibit site-specific maturation dynamics across the life span

city: New York City

first: Joshua

middle: Ian

last: Gray

state: New York

address: 630 168th Street

department: CCTI

institute: Columbia University

summary: Early in ontogeny, gd T cells emerge from the thymus and directly seed peripheral tissues for in situ immunity, though their function in human tissues is not clear. Through analysis of gd T cells in mucosal and lymphoid tissues across the human lifespan, we reveal age- and site-specific differentiation states, functions, and dynamics. Pediatric tissues uniquely contain newly generated gd T cells with diverse repertoires, exhibiting tissue repair and effector functions. In adult tissues, gd T cells persist in lower frequencies, exhibiting highly expanded repertoires and cytolytic profiles. Over childhood, gd T cells convert to adult-like differentiation profiles in mucosal sites, revealing that gd T cells provide diverse recognition and functions for numerous antigenic challenges during early life but persist as terminally differentiated, innate-like cells thereafter.

overall_design: gd T cells were isolated by Fluorescence-activated cell sorting from five tissue sites (spleen, lung, LLN, jejunum, MLN) of four human organ donors between the ages of 1.3 and 2 years of age. Individual samples i.e. single organs from each donor, were stained with a unique CITE-seq hashtag and analyzed using scRNAseq

first: Daniel

last: Caron

first: Donna

last: Farber


title: Epithelial-Immune Metabolic Codependency Fuels Inflammatory Disease [Bulk RNA-seq]

city: New York

first: Shruti

last: Naik

address: 1 Gustave L. Levy Pl

department: Immunology & Immunotherapy

institute: Icahn School of Medicine at Mount Sinai

summary: Inflammatory skin diseases are spurred by unchecked immune-epithelial circuits. Although the function of cytokine and chemokine signals in facilitating this crosstalk is well established, the specific metabolic mediators involved and their simultaneous contribution to dysregulation of these two distinct cellular compartments is unclear. Here, we employed scRNA-seq, spatial transcriptomics, and immunofluorescence across multiple disease indications to elucidate a dysfunctional epithelial state marked by Hypoxia Inducible Factor 1 alpha (HIF1⍺). Ex vivo HIF1⍺ blockade of human Psoriasis (PsO) lesions led to a reduction in pathological gene expression via modulation of glucose metabolism. Epidermal-specific loss of HIF1⍺ or its transcriptional target, glucose transporter 1, curtailed both epidermal pathology and the cutaneous immune response in murine PsO. Glycolytic metabolism sustained epithelial hyperproliferation and differentiation, while lactate, its terminal product, was crucial for sustaining Type 17 response. Notably, inhibition of lactate dehydrogenase A or the lactate transporters MCT1/4 selectively attenuated the Type 17 response, underscoring a divergent requirement for glucose and lactate in epithelial and immune cells, respectively. Collectively, these findings identify therapeutically targetable immune-epithelial circuits by unveiling a remarkable coordination of metabolic processed between the epithelial and immune compartments in inflammatory skin disease.

overall_design: Bulk RNA sequencing from two experimental studies assessing effect of dampening HIF1⍺ in inflammatory disease: (1) Human samples: 4 mm full-thickness punch biopsies from human lesional psoriatic skin and healthy skin were subjected to DMSO vehicle, 10µM of BAY87-2243, and 500nM of calcipotriol and 5µM of betamethasone dipropionate (standard of care) for 24 hours and submitted for sequencing to assess transcriptional differences between treatment conditions. (2) Murine samples: backskin taken after three days of imiquimod-treated WT or epithelial-specific HIF1⍺ knockout mice for comparative gene expression analyses.

first: Ipsita

last: Subudhi

first: Piotr

last: Konieczny

first: Aleksandr

last: Prystupa


title: CD4+ T cells and Bcl3

overall_design: Refer to individual Series


title: Epithelial-Immune Metabolic Codependency Fuels Inflammatory Disease [Spatial Transcriptomics]

summary: Inflammatory skin diseases are spurred by unchecked immune-epithelial circuits. However, the specific metabolic factors involved in crosstalk and their impact on the dysregulation of these two cellular components are not well understood. To better decipher the  metabolic factors involved in psoriasis, we employed spatial transcriptomics (ST), a ground-breaking technology that precisely maps gene expression from histologically-intact tissue sections. These findings identify therapeutically targetable metabolic vulnerabilities in inflammatory skin disease by unveiling a remarkable coordination of metabolic processes between the epithelial and immune compartments.

overall_design: This is a cross-sectional study involving the accrual of 3-4-mm skin punch biopsy samples from pre- and post-treatment lesional and non-lesional psoriatic skin. The samples were then subjected to the protocols entailed by spatial transcriptomics platform.


title: Epithelial-Immune Metabolic Codependency Fuels Inflammatory Disease


title: Hypoxia-inducible PRMT2 addiction in cancers

city: Tianjin

first: feng

last: dong

address: Qixiangtai Road 22

institute: Tianjin Medical University

summary: Hypoxia-inducible transcription factors (HIFs) are crucial transcription factors for cellular response to low oxygen levels, but the key mediators for their downstream transcription activation are not well characterized. We previously found that PRMT2 activates target gene expression through its methyltransferase activity on histone H3R8. Here we find that PRMT2 expression is activated by HIF1 at hypoxic conditions. And PRMT2 activity is widely required for hypoxia-induced transcription activation. Accordingly, PRMT2 inactivation alleviates hypoxia-induced glioblastoma progression and chemotherapeutic resistance. And PRMT2 expression is associated with hypoxia signature

overall_design: The H3R8me2a profiles of U87 shScr-Normoxia, U87 shScr-Hypoxia and U87 shPRMT2-Hypoxia

first: Xudong

last: Wu

first: Feng

last: Dong

first: Xiaoyu

last: Sun


title: Comparative gene expression profiling analysis of RNA-seq data for mCherryHigh and DTC populations compared to mCherryLow metastatic populations [mCherry Pops and DTCs]

first: Lai Ping

last: Wong

address: 185 Cambridge St

department: Molecular Biology

institute: Massachusetts General Hospital

summary:
To investigate characteristics of the Gstt1high metastatic subppulation in pancreatic cancer, lung metastatic cell line was generated to express mCherry from the Gstt1 endogenous promoter. Mice were injected via tail vein with mixed populations until evidence of metastatic disease, or 5 days after injection for DTC sorting.
Upon evidence of metastatic disease, mice were euthanized and lung tissues were digested followed by sorting of mCherry tumor cell populations (top 15-20%) gated on GFP. DTCs were collected 5 days after injection by gating and sorting on GFP. We then performed gene expression profiling analysis using data obtained from RNA-seq comparing mCherryHigh and low populations.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for tumor cells sorted for mCherryHigh and DTC populations compared to mCherryLow populations.

first: Christina

last: Ferrer

first: Raul

last: Mostoslavsky


title: Effect of depletion of Gstt1 on gene expression of metastatic derived cell lines [Gstt1 KD in Metastatic Cells]

summary:
To investigate the function of Gstt1 in the regulation metastatic phenotypes, we established liver and lung derived metastatic cell lines in which either NT control or Gstt1 has been knocked down by shRNA.
We then performed gene expression profiling analysis using data obtained from RNA-seq of 6 different cell lines

overall_design: Comparative gene expression profiling analysis of RNA-seq data for liver and lung metastatic cells and its KD derivative (Gstt1, sh1 and sh2).


title: Differential expression gene signature between primary tumors and metastatic tumors in PDAC and BC mouse models devoid of stromal contamination [GFP Sorted Cells RNA-Seq]

summary:
To identify a common differentially expressed gene signature between metastatic tumors and primary tumors, we utilized both p48-Cre/p53F/+KrasL/+/ROSA-LSL-YFP  and SIRT6/p48-Cre/p53F/+KrasL/+/ROSA-LSL-YFP as well as orthotopically injected 4T1-eGFP Balb/c mice to generate spontaneous. metastases
We then sorted YFP and GFP positive cells from primary and metastatic sites in mice with evidence of metastatic disease. Comparative gene expression profiling on sorted cells was then performed using RNA-Seq.

overall_design: RNA was isolated from GFP and YFP sorted tumor cells  generated from primary tumors and matched liver and lung metastases from mouse models described above. Comparative gene expression profiling analysis of RNA-seq data for YFP and GFP sorted primary tumor cells and metastatic cells.


title: Differential expression gene signature between primary tumors and metastatic tumors in PDAC and BC mouse models [Bulk Tissue RNA-Seq]

summary:
To identify a common differentially expressed gene signature between metastatic tumors and primary tumors, we utilized both p48-Cre/p53F/+KrasL/+  and SIRT6/p48-Cre/p53F/+KrasL/+ as well as orthotopically injected 4T1 Balb/c mice to generate spontaneous metastases
We then isolated RNA from bulk tissue primary tumors and metastases in mice with evidence of metastatic disease. Gene expression profiling on primary tumors and metastases was then performed using RNA-Seq.

overall_design: RNA was isolated from bulk tumor tissue generated from primary tumors and matched liver and lung metastases from mouse models described above. Comparative gene expression profiling analysis of RNA-seq data for primary tumors and metastatic tumors.


title: The transcriptional co-repressor Runx1t1 is essential for N-Myc-driven neuroblastoma tumorigenesis [ChIP-Seq]

city: Randwick

first: Murray

last: Norris

state: NSW

address: Lowy Cancer Research Centre

country: Australia

institute: Children's Cancer Institute

summary: Changes in epigenetic regulation are believed to be a major contributing factor to neuroblastoma development. Using a large-scale in vivo mutagenesis screen in Th-MYCN transgenic mice, we identified a single point mutation in the transcriptional corepressor Runx1t1, that can block N-Myc-driven neuroblastoma tumorigenesis. The loss of function mutation disrupts a highly conserved zinc finger domain (NHR4) within Runx1t1. Crossing an independent Runx1t1 knockout model with Th-MYCN mice, demonstrated that Runx1t1 haploinsufficiency is enough to prevent neuroblastoma development and reverse ganglia hyperplasia. Silencing RUNX1T1 in human neuroblastoma cells resulted in decreased colony formation in vitro, and significant inhibition of tumor growth in vivo. Our results show that RUNX1T1 forms part of a transcriptional LSD1-CoREST3-HDAC repressive complex that regulates the epigenomic landscape and chromatin accessibility, to control neuron-specific pathway genes and maintain an undifferentiated state. Runx1t1 thus represents an entirely novel and highly promising target not previously described in neuroblastoma.

overall_design: KELLY cells containing doxycycline-inducible shRNA to Runx1t1 were cultured with and without treatment. After 72 hours, 12 × 106 cells were fixed per condition to perform ChIP assays and sequencing for K4 monomethyl-histone H3 (H4K4me1), K4 dimethyl-histone H3 (H3K4me2), K4 trimethyl-histone H3 (H3K4me3), k27 acetyl-histone H3 (H3K27AC), and K27 trimethyl-histone H3 (H3K27me3) based on ChIP-seq method as previously described ( Valdés-Mora F et, al, 2017).

first: Emanuele

last: Valli

first: Jayne

middle: E

last: Murray

first: Chelsea

last: Mayoh

first: Nisitha

last: Jayatilleke


title: Targeting the intestinal circadian clock by meal timing ameliorates gastrointestinal inflammation

city: Freising

first: Yunhui

last: Niu

address: Gregor-Mendel-Str. 2

institute: Technische Universität München

summary: Investigating the function of intestinal Bmal1 in regulating the colonic transcriptome

overall_design: We performed gene expression analysis obtained from colonic tissues at 6 timepoints over the 24-h day (in constant darkness)

first: Marjolein

last: Heddes

first: Kinga

last: Balázs


title: Differential expression gene signature

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


title: Effects of KRAS (G13C) expression on transcriptome profiles assessed in HPCs differentiated from patient-derived iPS cells

city: Sagamihara

first: Makoto

last: Otsu

state: Kanagawa

address: 1-15-1 Kitasato, Minami-ku

institute: Kitasato University School of Medicine

summary: How a single oncogenic RAS impacts non-transformed hematopoietic progenitor cells (HPCs) remains largely unknown. We therefore investigated how a monoallelic KRAS mutation (G13C) affected HPCs utilizing induced pluripotent stem cells (iPSCs) derived from two unrelated patients with RAS-associated autoimmune lymphoproliferative syndrome-like disease (RALD).

overall_design: We performed gene expression profiling analysis of 12 HPC samples in total, with 6 samples being from each patient. The two clones from each patient constituted an isogenic pair of different genotypes (WT/WT and WT/G13C). Three samples obtained from each clone were subjected to the analysis, one with no stimulation, and the other two each stimulated with different cytokine combinations.

first: Huan-Ting

last: Lin

first: Masatoshi

last: Takagi

first: Kenji

last: Kubara

first: Kazuto

last: Yamazaki

first: Fumiko

last: Michikawa

first: Takashi

last: Okumura

first: Takuya

last: Naruto

first: Tomohiro

last: Morio

first: Koji

last: Miyazaki

first: Hideki

last: Taniguchi


title: The transcriptional co-repressor Runx1t1 is essential for N-Myc-driven neuroblastoma tumorigenesis [RNA-Seq 1]

first: Murray

last: Norris

overall_design: KELLY cells containing doxycycline-inducible shRNA to Runx1t1 were cultured with and without treatment. After 72 hours, Total RNA was isolated using the RNeasy Micro kit (50), # 74004 kit and RNA quality was assessed prior to being used (Agilent 2100 Bioanalyzer, Agilent Technologies). Only RNA with an RNA Integrity Number (RIN) of >9.0 was used. Whole transcriptome sequencing was performed at the Ramaciotti Centre for Genomics with SMARTer Stranded Total RNA-seq v2  preparation kit. Prepared libraries were pooled and sequenced on NovaSeq 6000 S1 2x100bp lane generating an average of 60 million reads per sample.

first: Emanuele

last: Valli

first: Jayne

last: Murray


title: Peroxisome dynamics determines host-derived ROS accumulation and infectious growth of the rice blast fungus

city: Nanjing

first: jun

last: Zhang

address: 1 Weigang, Nanjing City, Jiangsu Province

institute: Nanjing Agricultural University

summary: To investigate the underlying mechanisms that which genes involving in M. oryzae peroxisome dynamics in response to oxidative stress.

sample_organism:
Oryza sativa
Pyricularia oryzae

platform_organism:
Oryza sativa
Pyricularia oryzae

overall_design: The transcriptome analysis of Guy11 with or without 5  mM H2O2 treatment and the infected leaves at 24 hpi was performed by RNA-Seq experiment.

first: Jun

first: Haifeng


title: Improving vitrification efficiency of human in vitro matured oocytes by the addition of LEA proteins

city: taiyaun

first: Lu

last: Li

address: Wuyi Road, Xinghualing District, Taiyuan City, Shanxi Province

institute: The Second Hospital of Shanxi Medical University

summary: Vitrification cryopreservation of oocytes is an enabling technology for assisted reproductive technology. However, many factors in the vitrification process, such as the toxicity of cryoprotectant, osmotic stress, ice crystal formation during rewarming, will cause fatal damage to oocytes, thus affecting the embryo developmental potential and  subsequent clinical outcomes. Therefore, oocyte vitrification still faces significant challenges. Recent studies have shown that LEA protein has the potential to improve oocyte cryopreservation, while the molecular mechanism by which it exerts protective effects is still unclear. Therefore, we have systematically investigated the effects of LEA proteins on the vitrification cryopreservation and their molecular mechanisms. Results revealed that the proteins improved the developmental potential of human oocytes following cryopreservation, mostly by downregulating FOS genes and reducing oxidative stress. Moreover, the synergistic cryoprotection of LEA proteins by inhibiting the formation of ice crystals was given full play. This study provides a new strategy for high-quality cryopreservation of human oocytes.

overall_design: The SMART (Switching Mechanism at 5' End of RNA Template) technique was used to build the library. Using the total RNA of a single cell as a template, the first-strand cDNA was synthesized and amplified to obtain a sample sufficient for sequence analysis. According to the experimental requirements, a control group (0M), 100 µg/mL AfrLEA2 group, 100 µg/mL AfrLEA3m group and fresh group were set up with 3 replicates in each group, and a total of 12 human oocytes were collected from the corresponding condition groups. After washing each oocyte twice in PBS, a volume of less than 5 µl of cells and fluid was aspirated and quickly transferred under a microscope to a clean 200 µl PCR (RNase-free) tube containing lysate, and each sample was placed on dry ice and frozen immediately after the addition of lysate for the next sample. The PCR tubes were grouped and numbered with a marker, and the centrifuge tubes were tightly wrapped with Parafilm sealing film to keep the samples in a frozen state. This step was performed on dry ice as much as possible.

first: Xingyu

last: Bi

first: Xueqing

last: Wu

first: Zhongrong

last: Chen

first: Yunxia

last: Cao

first: Gang

last: Zhao


title: The transcriptional co-repressor Runx1t1 is essential for N-Myc-driven neuroblastoma tumorigenesis


title: Multiome (GEX + ATAC) profile at single cell level of Foxf1 positive endothelial cells and mesenchymal cells from mouse lung at E18.5

city: Cincinnati

first: Guolun

last: Wang

state: OH

address: 3333 Burnet Avenue

institute: Cincinnati Children's Hospital Medical Center

summary: Foxf1 is enriched and differentially expressed in lung cells derived from mesoderm. We used single cell RNA sequencing (snRNAseq + snATAC) to analyze the dynamic transcriptional regulation in various lung cell types. The single cell analysis show the cell specific regulation of Foxf1 by distal enhancers that contribute to the molecular pathology of ACDMPV.

overall_design: Foxf1 positive cells from lung of mice were isolated by Fluorescence-activated cell sorting (FACS) according to the presence or absence of GFP signal and analyzed using 10X multiome-seq. The cells derived from mESC cells via blastocyst complementation are processed and analysed by single cell RNA sequencing.

first: Vladimir

last: Kalinichenko


title: Clinicopathological and molecular landscape of 5-year IDH-wild-type glioblastoma survivors: a multicentric retrospective study

status: Public on Mar 31 2024

city: Rome

first: Evelina

last: Miele

address: Piazza di Sant'Onofrio 4

department: Oncoematologia

institute: Ospedale Pediatrico Bambino Gesù

summary: Background: Less than 5% of glioblastoma multiforme (GBM) patients survive more than 5-years (long-term survivors, LTS). Nevertheless, the molecular fingerprint of LTS remains uncharted. We aimed to characterize LTS by clinicopathological assessment, DNA methylation (DNAm)/Copy Number Variation (CNV) and mutation profiling.   Methods: This multicentric project proposed by Alliance Against Cancer was coordinated by Fondazione IRCCS Istituto Neurologico Carlo Besta. One-hundred forty-two LTS patients were screened, and samples centrally reviewed; 95 patients were enrolled. DNAm profiles and sequencing were performed on 27 LTS and 24 standard survival GBM (STS).    Results: We focused on 73 IDHwt-LTS. All patients underwent at least partial resection, followed by Stupp protocol (48/70). The median time to progression was 43 months (4-186), and almost all received a second treatment. Morphological and routine diagnostic molecular features of LTS did not deviate from the classic findings. MGMT promoter methylation was detected in 92% of cases.    No significant differences were observed between LTS and STS in terms of prevalence of mutated genes, with TP53 as the most commonly mutated gene (26% of all samples). DNAm showed a more heterogeneous group for LTS, with several cases classified as pediatric high-grade glioma and peculiar CNVs.  We identified 18 differentially methylated regions, 4 associated with survival probability (NR2F2, MME, WDR66 and ENPP4). Methylation levels of individual probes in IGFBP3 gene and corresponding protein expression showed a significant correlation with survival.   Conclusions: We found no significant different morphology nor mutational profile of LTS. DNAm confirmed as a valuable tool for GBM classification. No specific episignature was detected, but methylation levels of specific genes had prognostic value in LTS.

overall_design: 27 LTS (long-term survival) glioblastoma patients were compared to 24 standard survival (control) glioblastoma patients

first: Sara

last: Patrizi

first: Andrea

last: Ciolfi

first: Lucia

last: Pedace

first: Sabina

last: Barresi

first: Angela

last: Mastronuzzi

first: Marco

last: Tartaglia

first: Franco

last: Locatelli