title: Reprogramming Muller glia to regenerate ganglion-like cells in adult mouse retina with developmental transcription factors.

first: Thomas

last: Reh

state: Washington

address: 1959 NE Pacific Street, UW School of Medicine, Seattle, WA 98195

department: Department of Neurobiology and Biophysics

institute: University of Washington

summary: Many neurodegenerative diseases cause degeneration of specific types of neurons. For example, glaucoma leads to death of retinal ganglion cells, leaving other neurons intact. Neurons are not regenerated in the adult mammalian central nervous system. However, in non-mammalian vertebrates, glial cells spontaneously reprogram into neural progenitors and replace neurons after injury. We have recently developed strategies to stimulate regeneration of functional neurons in the adult mouse retina by overexpressing the proneural factor Ascl1 in Müller glia. In this report, we test additional transcription factors (TFs) for their ability to direct regeneration to particular types of retinal neurons. We engineered mice to express different combinations of TFs in MG, including Ascl1, Pou4f2, Islet1 and Atoh1. Using IHC, scRNA-seq, scATAC-seq, and electrophysiology we find retinal ganglion-like cells can be regenerated in the damaged adult mouse retina in vivo with targeted overexpression of developmental RGC-transcription factors.

overall_design: Included in this submission are scRNA-seq and scATAC-seq data from overexpression experiments of Islet1/Pou4f2/Ascl1 (IPA) or Ascl1-only in murine Muller glia, both in vivo and in vitro.  For in vivo datasets, overexpression of transgenes alongside GFP in Glast-Cre mice was combined with intravitreal injections of NMDA and TSA.  After >3 weeks, retinas from 4-8 mice were pooled and dissociated and the GFP+ cells were FACS purified and then processed with the 10x genomics platform for scRNA-seq or scATAC-seq.  Additionally, retinas dissociated from E14 mice were processed for scATAC-seq.  For the in vitro experiments, transgenes were under control of rtTA.  Muller glia cultured from 3-5 mice and treated with doxycycline to induce transgenes for 5 days, alongside untreated control glia, were processed with the 10x platform for scRNA-seq.

first: Levi

last: Todd

first: Wesley

last: Jenkins

first: Connor

last: Finkbeiner

first: Marcus

last: Hooper

first: Phoebe

middle: C

last: Donaldson

first: Marina

last: Pavlou

first: Juliette

last: Wohlschlegel

first: Norianne

last: Ingram

first: Fred

last: Rieke


title: BRD8 maintains glioblastoma by epigenetic reprogramming of the P53 network [RNA-seq]

status: Public on Sep 16 2022

city: Cold Spring Harbor

first: Olaf

last: Klingbeil

state: New York

address: 1 Bungtown Road

country: USA

institute: Cold Spring Harbor Laboratory

type: Expression profiling by high throughput sequencing

summary: Transcriptome analysis by high throughput sequencing

overall_design: Gene expression profiles in glioblastoma with gene knockout

first: Xueqin

last: Sun

first: Alea

middle: A

last: Mills


title: Parenteral BCG vaccine induces respiratory mucosal-resident memory macrophages and trained innate immunity via the gut-lung axis I

city: Hamilton

first: Zhou

last: Xing

state: Ontario

address: MDCL building, 1280 Main St W

department: Division of Respirology

country: Canada

institute: McMaster University

summary: Besides centrally induced innate immune memory/trained immunity in the bone marrow/peripheral blood by parenteral vaccination or infection, recently emerging evidence suggests that the barrier mucosal tissue-resident innate immune memory may develop via a local inflammatory pathway following mucosal immunologic exposure. However, it remains unclear whether the mucosal-resident innate immune memory may result from integrating distally generated immunological signals following parenteral vaccination/infection.  We show here that subcutaneous Bacillus Calmette-Guérin (BCG) vaccination is able to induce memory alveolar macrophages (AM) and trained immunity at the respiratory barrier mucosa. Although parenteral BCG vaccine can centrally train bone marrow progenitors and circulating monocytes, induction of memory AM is entirely independent of circulating monocytes. Rather, parenteral BCG vaccination, via distal mycobacterial dissemination, causes a time-dependent alteration in gut microbiome, barrier function and microbial metabolites including short-chain fatty acids, and subsequently the changes in circulating and lung tissue metabolites, leading to induction of tissue-resident memory macrophages and trained innate immunity in the lung. Our study thus reveals a novel gut microbiota-mediated pathway for innate immune memory development at distal barrier mucosal tissues. Our findings have far-reaching implications in developing next-generation parenteral vaccine strategies against respiratory pathogens such as M.tb.

overall_design: A mix of L-carnitine and butyrate hydrochloride was introduced to the drinking water (DW+M) of a group of naïve animals for a period of 3 weeks and as control, other animals received drinking water without supplements (DW). Alveolar macrophages were obtained by bronchoalveolar lavage and cultured in the presence (S) or absence (US) of stimulation with M. tuberculosis whole cell lysate for 16 h. Isolated RNA from the cultured cells were subjected to transcriptional analysis.

first: Mangalakumari

last: Jeyanathan


title: KLF4 inhibits early neural differentiation of ESCs by coordinating specific 3D chromatin structure, a new mechanism of early neural differentiation

status: Public on Sep 20 2022

city: Tianjin

first: Niannian

last: Li

state: Tianjin

address: No. 94 Weijin Road, Nankai District, Tianjin

department: college of Life Sciences

country: China

institute: Nankai University

type: Genome binding/occupancy profiling by high throughput sequencing

summary:
Circular RNA has been reported to be dynamically expressed during embryonic development and regulates human embryonic stem cells (hESCs), but the identification and regulation of functional circular RNA in mouse embryonic stem cells (mESC) remains unclear.
Neural differentiation of embryonic stem cells (ESCs) requires precisely orchestrated gene regulation, a process governed in part by changes in 3D chromatin structure. How these changes regulate gene expression in this context remains unclear. In this study, we observed enrichment of the transcription factor KLF4 at some poised or closed enhancers at TSS-linked regions of genes associated with neural differentiation. Combination analysis of ChIP, HiChIP and RNA-seq data indicated that KLF4 loss in ESCs induced changes in 3D chromatin structure, including increased chromatin interaction loops between neural differentiation-associated genes and active enhancers, leading to upregulated expression of neural differentiation-associated genes and therefore early neural differentiation. This study suggests KLF4 inhibits early neural differentiation by regulation of 3D chromatin structure, which is a new mechanism of early neural differentiation.
Conclusions: Our study  suggests KLF4 inhibits early neural differentiation by regulation of 3D chromatin structure, which is a new mechanism of early neural differentiation.

overall_design: Two biological replicates were analyzed for each experimental condition. WT and KO cells were RNA-seq/HiChIP in ESCs or neuro induction at day 12.

first: Wange

last: Lu

first: Jinfang

last: Bi

first: Wenbin

last: Wang


title: Lower nuclear pore numbers in cardiomyocytes decrease nuclear import of signaling proteins, which restricts adaptive gene regulation, and improves cardiac stress response in vivo

status: Public on Sep 26 2022

city: Pittsburgh

first: Bernhard

last: Kuhn

state: Pennsylvania

address: 4401 Penn Ave

institute: Children's Hospital of Pittsburgh

summary: Nuclear pores are essential pathways for nuclear-cytoplasmic transport of molecules. Whether and how cells change nuclear pores to alter nuclear transport and cellular function is unknown. Here, we show that heart muscle cells (cardiomyocytes) undergo a 63% decrease in nuclear pore numbers during differentiation, which alters their response to extracellular signals. This maturation-associated decline in nuclear pore numbers per nucleus is associated with lower nuclear levels and import of Mitogen-activated Protein Kinase (MAPK) signaling proteins. Experimental reduction of nuclear pore numbers decreased nuclear import of MAPK signaling proteins. In a mouse model of high blood pressure, reduction of cardiomyocyte nuclear pore numbers reduced adverse heart remodeling and gene regulation, which reduced progression to lethal heart failure. The observed decrease in nuclear pore numbers in cardiomyocyte differentiation and resulting functional changes suggest a paradigm by which terminally differentiated cells could permanently alter their handling of information flux across the nuclear envelope and, with that, their behavior.

overall_design: mRNA profiles of control mice (Tnnt2-Cre+/-, Lmnb2fl/fl-Tnnt2 cre+/-) mice that underwent sham or TAC (transverse aortic constriction) sugeries.

last: Kühn

first: Lu

last: Han

first: Dhivyaa

last: Rajasundaram

first: Niyatie

last: Ammanamanchi


title: Differentially regulated miRNAs in skin fibroblasts with constitutively active Nrf2

city: Zurich

first: Paul

last: Hiebert

address: Otto-Stern-Weg 7

country: Switzerland

institute: ETH Zurich

type: Non-coding RNA profiling by high throughput sequencing

summary: The NRF2 transcription factor is a key regulator of the cellular antioxidant response, and pharmacological NRF2 activation is a promising strategy for the prevention of age-related diseases. Here we show, however, that genetic activation of Nrf2 in skin fibroblasts leads to the production of a matrisome, which is similar to aged skin and characterized by reduced synthesis and deposition of selected matrix proteins, including the major skin collagens.  Mechanistically, the production of an aging matrisome results from Nrf2-mediated overexpression of microRNAs (miRs), which directly suppress collagen expression independently of myofibroblast differentiation. This is functionally important, as Nrf2 activation in fibroblasts caused structural abnormalities of the dermal ECM and increased the susceptibility to skin tearing, similar to aged skin. Activation of endogenous NRF2 was observed in human skin from elderly individuals in vitro and in vivo. These data implicate a key role for activated NRF2 in controlling age-related molecular and biomechanical skin alterations.

overall_design: Primary fibroblasts were isolated and cultured from the skin of newborn transgenic mice with Cre-recombinase expressed in fibroblasts (Col1a2Cre). Fibroblasts from 3 mice with Cre plus a constitutively active Nrf2 (caNrf2) mutant and 3 mice with Cre only (ctrl) were used. Fibroblasts were allowed to proliferate in culture for 6 days to allow for sufficient numbers for miRNA sequencing. Total RNA was isolated from cells followed by miRNA-seq analysis.

first: P

first: S

last: Werner


title: Modulation of histone H3K4 dimethylation by spermidine ameliorates motor neuron survival and neuropathology in a mouse model of ALS

status: Public on Sep 13 2022

city: Seoul

first: Hoon

last: Ryu

state: Seoul

address: Seoul

department: Brain Science Institute

country: South Korea

institute: Korea Institute of Science and Technology

summary: Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by progressive paralysis due to motor neuron degeneration. It has been proposed that epigenetic modification and transcriptional dysregulation may contribute to motor neuron death. In this study, we investigate the basis for therapeutic approaches to target lysine-specific histone demethylase 1 (LSD1) and elucidate the mechanistic role of LSD1-histone H3K4 signaling pathway in ALS pathogenesis. Cellular and animal models of ALS were treated by spermidine (SD), then we performed neuropathological, body weight and survival evaluation in order to examine the role of SD in mutant superoxide dismutase 1 (mSOD1) (G93A) ALS mice. Herein, we found that dimethylated histone H3K4 (H3K4me2) levels are decreased in cellular and animal models of ALS, and correlated with motor neuronal dysfunction in ALS (G93A) mice. SD administration modulated the LSD1 activity and restored H3K4me2 levels in ChAT-positive motor neurons in the lumbar spinal cord of ALS mice. SD prevented cellular damage by improving the number and size of motor neurons in ALS mice. SD administration also reduced GFAP-positive astrogliogenesis in the white and gray matter of the lumbar spinal cord, improving the neuropathology of ALS mice. Moreover, SD administration delayed disease onset and prolonged the lifespan of ALS (G93A) transgenic mice. Together, modulating epigenetic targets such as LSD1 by small compounds may be a useful therapeutic strategy for treating ALS.

overall_design:
Gene expression profiling analysis of RNA-seq data for G93A mice treated by spermidine.

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first: Ali

last: Yousefian-Jazi


title: BRD8 maintains glioblastoma by epigenetic reprogramming of the P53 network [ChIP-seq]

first: Olaf

last: Klingbeil

summary: Genome-wide binding/occupancy profiling by high throughput sequencing

overall_design: Genome-wide analysis of BRD8, p53, and H2AZ binding, and H3K27ac/H3K4me3 modification in glioblastoma

first: Xueqin

last: Sun


title: Parenteral BCG vaccine induces respiratory mucosal-resident memory macrophages and trained innate immunity via the gut-lung axis II

first: Zhou

last: Xing

overall_design: Mice were vaccinated sub cutaneously with BCG (BCG) or carrier (PBS). At 5 weeks post-vaccination, airway alveolar macrophages isolated by bronchoalveolar lavage (BAL) was cultured in the presence (S) or absence (US) of stimulation with M. tuberculosis whole cell lysate for 16 h. Isolated RNA from the cultured cells were subjected to transcriptional analysis.


title: Dynamics of regulatory elements in the liver during ruminant evolution

status: Public on Sep 15 2022

city: Beijing

first: Siqian

last: Chen

address: 2rd, Yuanmingyuan West Road

department: College of Animal Science and Technology

institute: China Agricultural University

summary: To investigate the evolutionary changes of  regulatory elements in ruminants, we then performed regulatory elements profiling analysis using ChIP-seq datasets (H3K27ac and H3K4me3) of liver from three ruminants.

sample_organism:
Bos taurus
Capra hircus
Ovis aries

platform_organism:
Bos taurus
Capra hircus
Ovis aries

overall_design: Comparative analysis of regulatory elements in the liver for three ruminants.

first: Ying

last: Yu


title: Aerobic Exercise Training Rejuvenates the Human Skeletal Muscle Methylome Ten years after Breast Cancer Treatment and Survival

status: Public on Sep 12 2022

city: Oslo

first: Adam

middle: P

last: Sharples

address: 220 Sognsveien

country: Norway

institute: Norwegian School of Sport Sciences

summary: Many cancer survivors suffer from impairments in skeletal muscle (SkM), both in terms of reduced mass and function. Interestingly, human SkM possesses an epigenetic memory of earlier stimuli, such as exercise. Long-term retention of epigenetic changes in SkM following cancer survival and/or exercise training have not yet been studied. We therefore investigated genome-wide DNA methylation (the methylome) in SkM following a 5-month, 3/week treadmill-based aerobic training intervention in breast cancer survivors 10-14 years after diagnosis and treatment. These results were compared to breast cancer survivors who remained untrained and to age-matched controls with no history of cancer, who undertook the same training intervention. SkM biopsies were obtained before (pre) and after (post) the 5-month training period and DNA methylation was analysed using InfiniumEPIC 850K Arrays. The breast cancer survivors displayed a significant retention of increased DNA methylation (i.e., hypermethylation) at a larger number of differentially methylated positions (DMPs) compared with healthy age-matched controls pre-training. Training in cancer survivors led to an exaggerated number of DMPs with a hypermethylated signature occurring at more random non-regulatory regions across the DNA compared with training in healthy age-matched controls. However, the opposite occurred in important gene regulatory regions, where training in cancer survivors elicited a considerable reduction in methylation (i.e., hypomethylation) in 99% of the DMPs located specifically in CpG islands within promoter regions. Importantly, training was able to reverse the hypermethylation identified in cancer survivors back towards a hypomethylated signature that was observed pre-training in healthy age-matched controls at 300 (out of 881) of these island and promoter associated CpG sites. Pathway enrichment analysis identified that the training in cancer survivors evoked this predominantly hypomethylated signature in pathways associated with: Cell cycle, DNA replication and repair, transcription, translation, mTOR signalling and proteosome. Differentially methylated region (DMR) analysis also identified genes: BAG1, BTG2, CHP1, KIFC1, MKL2, MTR, PEX11B, POLD2, S100A6, SNORD104 and SPG7 as hypermethylated in breast cancer survivors with training reversing these CpG island / promoter associated DMRs towards a hypomethylated signature. Training also elicited a largely different epigenetic response in healthy individuals than that observed in cancer survivors, with very few overlapping changes. Only one gene, SIRT2, was identified as having altered methylation in cancer survivors at baseline as well as after training in both the cancer survivors and healthy controls. In conclusion, human SkM muscle retains a hypermethylated signature for as long as 10-14 years after breast cancer treatment and survival. Importantly, 5 months of aerobic training rejuvenated the SkM methylome towards signatures identified in healthy age-matched individuals in gene regulatory regions.

overall_design:
Ethical approval:

This study is part of the ongoing CAUSE-trial (Cardiovascular Survivor Exercise) initiated by the Norwegian School of Sport Sciences (NSSS) and Oslo University Hospital. The Regional Committees for Medical and Health Research Ethics (REK) approved the study (reference number 28930; 2019/1318), and the study was pre-registered in clinicaltrials.gov (NCT04307407).

Participants and Experimental Design:

The CAUSE-trial is a two-armed randomised controlled exercise training trial, where breast cancer survivors are randomised to either an exercise group or a control group (usual care), that also includes third arm of age-matched women with no prior history of cancer undertaking the same exercise training program. Breast cancer survivors diagnosed, with stage II-III HER2 negative breast cancer at the age of 60 years or less, between 2008 and 2012 were identified by the Cancer Registry of Norway. The study included survivors that had received anthracycline-based chemotherapy (Epirubicin), were currently exercising less than 90 minutes at moderate to high intensity per week, and that were living within driving distance from NSSS. Major exclusion criteria for the study were stage IV breast cancer diagnosis, adjuvant Trastuzumab-treatment, recurrent breast cancer or presence of secondary cancers, previous major cardiac surgery, pacemaker, chronic atrial fibrillation or any recent or uncontrolled cardiovascular disease. Participants for the healthy age-matched control group were recruited via advertisements. The participants were matched for age with the breast cancer survivors and could not have a history of any cancer and otherwise had to adhere to the same eligibility criteria as the breast cancer survivors (except for the breast cancer diagnosis).

The breast cancer survivors were block randomised with a 1:1 allocation ratio to either an aerobic exercise training group (Cancer Trained) or to a control group (Cancer Untrained) for 5 months (20 weeks). The training intervention is described below. Participants in the healthy age-matched group did not undergo randomization, but all underwent the same exercise protocol as the cancer trained group (Healthy Age-Matched Trained). All participant groups had SkM biopsies taken and underwent other study-related assessments pre and post training. SkM biopsy methods are detailed below.     Aerobic Exercise Training: Both the cancer survivors training group and the healthy age-matched group underwent five months of 3 times per week treadmill-based endurance training, aimed at increasing V̇O2peak. The program followed a non-linear periodization model, typically with one hard (i.e., 80-87% of HRpeak), one moderate (i.e., 70-80% of HRpeak) and one easy (i.e., 60-70% of HRpeak) session per week. Initially, all sessions were continuous in duration, but from week 5, the hard session was an interval session, with 3-5 intervals each lasting 4 minutes (i.e., 90-97% of HRpeak). After week 10, we introduced 8-minute intervals for the moderate session (i.e., 85-91% of HRpeak), progressing from 2-4 intervals throughout the intervention.

Skeletal Muscle Biopsies:

The biopsy procedure was conducted under local anaesthesia (Xylocaine with adrenaline, 10 mg/ml lidocaine + 5 µg/ml adrenaline, AstraZeneca, London, UK) and muscle tissue was obtained using a modified Bergström technique with suction. The tissue was quickly rinsed in physiological saline, before fat, connective tissue and blood were removed and discarded. Subsequently, the samples were quickly frozen in isopentane and cooled on dry ice or liquid nitrogen, before being transferred and stored at −80°C for later isolation of DNA. All biopsies from both pre and post training time points were taken at rest, in the morning after an overnight fast from the lateral portion of vastus lateralis muscle. Post training samples were obtained at least 72 hrs after the end of the training intervention. Pre and post biopsies were taken from the left leg in all participants.

Cardiopulmonary Exercise Test and Physical Activity levels:

Participants body height and mass were recorded using a stadiometer and a scale (SECA 213, Hamburg, Germany). Cardiorespiratory fitness (V̇O2peak) was assessed by a treadmill-based, symptom-limited cardiopulmonary exercise test, using a modified Balke protocol. After familiarisation with the treadmill, participants were encouraged to walk at gradually increasing exercise loads (increased inclination or speed) until voluntary exhaustion. Continuous expired air was measured breath-by-breath, using a gas and volume calibrated metabolic chart (Oxycon Pro, Jarger GmbH, Hoechberg, Germany). Objectively measured physical activity levels were recorded for seven consecutive days, using the ActiGraph™ model GT3X+ (ActiGraph LLC, Pensacola FL, USA). The average minutes spent in light and moderate-to-vigorous physical activity, using cut-off values used previously [1], with light physical activity defined as 100-1951 counts per minute and moderate-to-vigorous physical activity >1952 counts per minute. Baseline characteristics of V̇O2peak and physical activity levels for all participants can be found below. There was no significant difference in V̇O2peak or physical activity levels between the cancer groups and the healthy age-matched group prior to the start of the exercise program.

Tissue Homogenization, DNA isolation, Bisulfite Conversion and Genome-Wide DNA Methylation:

DNA was isolated from SkM tissue derived from a randomly selected subpopulation within each group of the larger trial (n=5 cancer trained, n=5 cancer untrained and n=6 healthy age-matched trained) at both pre and post training. These subpopulations baseline characteristics can be found here in the order of: Cancer Trained (n=5), Cancer Untrained (n=5), Healthy Age-Matched Trained (n=6), Cancer Survivors vs. Healthy Age-matched (p-value) respectively for: Age (years; SD) 60.4 (5.9), 62.0 (5.5), 58.3 (4.8), p = 0.307; Body Height (cm; SD) 167.5 (5,9), 168.1 (5.0), 169.0 (4.8), p = 0.660; Body Mass (Kg; SD) 80.5 (7.9), 74.1 (8.6), 80.0 (16.5), p = 0.670; Body Mass Index (SD) 28.8 (3.8), 26.4 (4.3), 28.0 (5.8), p = 0.861; V̇O2peak (ml/kg/min; SD) 27.2 (3.9),30.9 (6.7), 30.7 (7.7), p = 0.630; Physical activity (PA) levels (minutes; SD) for Light PA 154 (44.9),154 (38.4), 162.9 (28.3), p = 0.652; Moderate to Vigorous PA 43.3 (34.6), 41.8 (16.4), 56.2 (31.6), p = 0.362. Baseline characteristics were well balanced between the groups. There was no difference in age, height, BMI, V̇O2peak or physical activity levels between the cancer groups and the healthy age-matched group. Muscle samples were homogenized for 45 seconds at 6,000 rpm × 3 (5 min on ice in-between intervals) in lysis buffer (180 µl buffer ATL with 20 µl proteinase K) provided in the DNeasy spin column kit (Qiagen, UK) using a Roche Magnalyser instrument and homogenization tubes containing ceramic beads (Roche, UK). The DNA was then bisulfite converted using the EZ DNA Methylation Kit (Zymo Research, CA, United States) as per the manufacturer’s instructions.    Infinium MethylationEPIC BeadChip Array    All DNA methylation experiments were performed in accordance with Illumina manufacturer instructions for the Infinium Methylation EPIC BeadChip Array. Methods for the amplification, fragmentation, precipitation and resuspension of amplified DNA, hybridisation to EPIC beadchip, extension and staining of the bisulfite converted DNA (BCD) can be found in detail in our open access methods paper [2, 3]. EPIC BeadChips were imaged using the Illumina iScan System (Illumina, United States).

DNA methylome analysis, differentially methylated Positions (DMPs), pathway enrichment analysis (KEGG pathways) and differentially methylated region (DMR) analysis:

Following MethylationEPIC BeadChip arrays, raw .IDAT files were processed using Partek Genomics Suite V.7 (Partek Inc. Missouri, USA) and annotated using the MethylationEPIC_v-1-0_B4 manifest file. The mean detection p-value for all samples was 0.0004, which was well below the recommended 0.01 in the Oshlack workflow [4]. We also examined the raw intensities/signals of the probes, that demonstrated an average median methylated and unmethylated signal of 11.77 and 11.69, respectively, with an average of 11.73, which is recommended to be above 11.5 [4]. The difference between the average median methylated and average median unmethylated signal was 0.07, well below the recommended difference of less than 0.5 [4]. Note: Upon import of the data into Partek Genomics Suite we filtered out probes located in known single nucleotide polymorphisms (SNPs) and any known cross-reactive probes using previously defined SNP and cross-reactive probe lists from EPIC BeadChip 850K validation studies [5]. Although the average detection p-value for each sample across all probes was very low (on average 0.0004) we also excluded any individual probes with a detection p-value that was above 0.01 as recommended previously [4]. Furthermore, due to an all-female cohort of breast cancer survivors and age-matched healthy controls, we filtered out any probes on the male only Y chromosome (therefore the analysis included all probes on the X chromosome only). Out of a total of 865,860 probes in the EPIC array, removal of known SNPs, cross-reactive probes, those with a detection p-value above 0.01 and those on the Y chromosome resulted in 808,804 probes being taken forward for downstream analysis. Following this, background normalisation was performed via functional normalisation (with noob background correction) as previously described [6]. After functional normalisation, we also undertook quality control procedures via principal component analysis (PCA). One sample (Sample No.13_205624860016_R05C01_Cancer Untrained_Pre_Participant 10)  was removed due to a larger variation than that expected within that condition (variation defined as values above 2.2 standard deviations (SDs) for that condition – depicted by ellipsoids in the PCA plots. Following normalisation and quality control procedures, we undertook differentially methylated position (DMP) analysis by converting β-values to M-values (M-value = log2(β / (1 - β)), as M-values show distributions that are more statistically valid for the differential analysis of methylation levels [7]. We then performed a two-way ANOVA for group/condition (cancer trained, cancer untrained, healthy age-matched trained) and time (pre and post), with planned contrast/pairwise comparisons of: Cancer Trained-Pre vs. Healthy Trained-Pre (to investigate the effect of Cancer on the methylome named - Cancer Alone), Cancer Trained-Post vs. Cancer Trained-Pre (to investigate the impact of training on the methylome of breast cancer survivors named - Training in Cancer), Healthy Age-Matched Trained-Post vs. Heathy Age-Matched-Pre (to investigate the impact of training on the methylome in age-matched healthy controls named- Training in Healthy), Cancer Untrained-Post vs. Cancer Untrained-Pre (to help assess if any methylation changes occur over the time of the intervention period in cancer survivors, named - Experimental Time), and finally Cancer Trained Pre vs. Cancer Untrained Pre (to assess if there were any differences in methylation between the cancer groups at baseline, named - Between Cancer Groups at Baseline). For initial discovery of CpG sites that were deemed statistically significant, DMPs with an unadjusted P value of ≤ 0.01 were accepted for downstream analysis (Kyoto Encyclopedia of Genes and Genomes/KEGG pathway, differentially methylated region/DMR analysis - see below). We then undertook CpG enrichment analysis on these DMPs within KEGG pathways [8-10] using Partek Genomics Suite and Partek Pathway software. DMR analysis was performed to identify where several CpGs were differentially methylated within a short chromosomal locations/regions, undertaken using the Bioconductor package DMRcate (DOI: 10.18129/B9.bioc.DMRcate).

IMPORTANT NOTE for Methylation Array files:

For submission of data to GEO it is recommended to upload all normalized data for ALL samples and ALL probes. Therefore, for the matrix signal intensities we include normalized data for all probes and all samples as the GEO submission. Therefore, it’s important to note, (as described in detail in the methods above), sample no. 13 (Sample No.13_205624860016_R05C01_Cancer Untrained_Pre_Participant 10) was removed from downstream analysis. Also, for any downstream analysis (DMP, DMR, GO / KEGG analysis etc.) we removed cross-reactive probes, probes on SNPs, probes with a detection p value of above 0.01 and due to an all-female cohort, we also removed any probes on the male only Y chromosome (therefore including all probes on the X chromosome only). We also removed 378 DMPs from downstream analysis that included those that were altered over the experimental time period (5 months) in the cancer untrained group as well as those that were significantly different between cancer groups at baseline (specific details can be found in the final publication).

REFERENCES:

1. Ekelund U, Tarp J, Fagerland MW, Johannessen JS, Hansen BH, Jefferis BJ, et al. Joint associations of accelero-meter measured physical activity and sedentary time with all-cause mortality: a harmonised meta-analysis in more than 44 000 middle-aged and older individuals. Br J Sports Med. 2020; 54(24):1499-1506.
2. Seaborne RA, Strauss J, Cocks M, Shepherd S, O'Brien TD, Someren KAV, et al. Methylome of human skeletal muscle after acute & chronic resistance exercise training, detraining & retraining. Scientific Data (Nature). 2018; 5:180213.
3. Seaborne RA, Strauss J, Cocks M, Shepherd S, O'Brien TD, van Someren KA, et al. Human Skeletal Muscle Possesses an Epigenetic Memory of Hypertrophy. Scientific Reports (Nature) 2018; 8(1):1898.
4. Maksimovic J, Phipson B, and Oshlack A. A cross-package Bioconductor workflow for analysing methylation array data [version 1; referees: 3 approved, 1 approved with reservations]. F1000Research. 2016; 5(1281).
5. Pidsley R, Zotenko E, Peters TJ, Lawrence MG, Risbridger GP, Molloy P, et al. Critical evaluation of the Illumina MethylationEPIC BeadChip microarray for whole-genome DNA methylation profiling. Genome Biology. 2016; 17(1):208.
6. Maksimovic J, Gordon L, and Oshlack A. SWAN: Subset-quantile within array normalization for illumina infinium HumanMethylation450 BeadChips. Genome Biol. 2012; 13(6):R44.
7. Du P, Zhang X, Huang C-C, Jafari N, Kibbe WA, Hou L, et al. Comparison of Beta-value and M-value methods for quantifying methylation levels by microarray analysis. BMC Bioinformatics. 2010; 11(1):587.
8. Kanehisa M and Goto S. KEGG: kyoto encyclopedia of genes and genomes. Nucleic Acids Res. 2000; 28(1):27-30.
9. Kanehisa M, Furumichi M, Tanabe M, Sato Y, and Morishima K. KEGG: new perspectives on genomes, pathways, diseases and drugs. Nucleic Acids Res. 2017; 45(D1):D353-d361.
10. Kanehisa M, Sato Y, Kawashima M, Furumichi M, and Tanabe M. KEGG as a reference resource for gene and protein annotation. Nucleic Acids Res. 2016; 44(D1):D457-62.


title: Comparative analyses of DNA methylomes in the liver among three ruminants

type: Methylation profiling by high throughput sequencing

summary: This study is the first to report a genome-wide comparative DNA methylation map of liver in three ruminants using WGBS technology.

overall_design: Investigate the changes of DNA methylomes in the liver using WGBS during ruminant evolution


title: DNA dioxygenases Tet2/3 control epithelial differentiation

status: Public on Sep 09 2022

city: BOSTON

first: Guodong

state: MA

address: 609 Albany Street, ROOM 414

department: Dermatology

institute: Boston University

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

overall_design: Refer to individual Series


title: DNA dioxygenases Tet2/3 control epithelial differentiation [RNA-seq]

summary:
Execution of lineage-specific differentiation programs requires tight coordination between many regulators including Ten-eleven translocation (TET) family enzymes catalyzing 5-methylcytosine oxidation in DNA. Here, by using Keratin 14-Cre-driven ablation of Tet genes in skin epithelial cells, we demonstrate that ablation of Tet2/Tet3 result in marked alterations of hair shape and length followed by hair loss. We show that through DNA demethylation, Tet2/Tet3 control chromatin accessibility, Dlx3 binding and promoter activity of the Krt25 and Krt28 genes regulating hair shape, as well as regulate interactions between the Krt28 gene promoter and distal enhancer. Moreover, Tet2/Tet3 also control three-dimensional chromatin topology in Keratin type I/II gene loci via DNA methylation-independent mechanisms. These data demonstrate the essential roles for Tet2/3 in establishment of lineage-specific gene expression program and control of Dlx3/Krt25/Krt28 axis in hair follicle epithelial cells and implicate modulation of DNA methylation as a novel approach for hair growth control.
We performed RNA-seq from sorted hair matrix keratinocytes of wild type and TetDKO (Krt14-Cre, Tet2 flox/flox, Tet3 flox/flox) mice at P4.5 which are treated or untreated with 5-aza.

overall_design: Hair matrix keratinocytes are isolated throgh FACS from mice at P4.5. For 5-aza treated mice, mice are treated with 5-aza at P2.5, and hair matrix keratinocytes are isolated following the same procedure decribed before. Total RNA is isolated from 100 - 200K sorted cells with RNeasy Micro Kit (QIAGEN, #74004) as the manual described, and eluted in 12 µl of DNase/RNase-free water. Zymo-Seq RiboFree Total RNA Library Kit (Zymo, #R3000) is used for cDNA synthesis and stranded library preparation as the manual described. DNA libraries are sequenced as the paired-end 2 × 150bp sequencing in an Illumina Hiseq4000 system.

first: Iqra

last: Fatima

first: Qin

last: Xu

first: Michael

middle: Y

last: Fessing

first: Andrei

middle: N

last: Mardaryev

first: Andrey

last: Sharov

first: Elena

last: Rozhkova

first: Guoliang

first: Vladimir

last: Botchkarev


title: BRD8 maintains glioblastoma by epigenetic reprogramming of the P53 network


title: Mitochondrial Transfer Regulates Cell Fate Through Metabolic Remodeling in Osteoporosis

status: Public on Sep 11 2022

city: Wuhan

first: yiqian

last: yu

state: Hubei

address: Luoyu Road 237#, 430079, Wuhan, Hubei

institute: wuhan university

summary: Mitochondria are the powerhouse of eukaryotic cells, which regulate cell metabolism and differentiation.  Recently, mitochondrial transfer between cells has been shown to direct recipient cell fate. However, it is unclear whether mitochondria can translocate to stem cells and whether this transfer alters stem cell fate. Here, we examined mesenchymal stem cell (MSC) regulation by macrophages in the bone marrow environment. We found that macrophages promote osteogenic differentiation of MSCs by delivering mitochondria to MSCs. However, under osteoporotic conditions, macrophages with altered phenotypes and metabolic statuses release oxidatively damaged mitochondria. The transfer of dysfunctional mitochondria to MSCs triggers a reactive oxygen species burst, which leads to metabolic remodeling. We showed that abnormal metabolism in MSCs is caused by the abnormal succinate accumulation, which is a key factor in abnormal osteogenic differentiation. These results reveal that mitochondrial transfer from macrophages to MSCs allows metabolic crosstalk to regulate bone homeostasis. This mechanism identifies a potential target for the treatment of osteoporosis.

overall_design:
The mRNA profiles of bone-derived macrophages of  8 weeks old sham and ovx mice
The mRNA profiles of mesenchymal stem cells of 8 weeks mice with no treatment, co-cultured with RAW cells or LPS stimulated RAW cells

first: Wenjin

last: Cai

first: Yufeng

last: Zhang


title: Detailed phenotypic and functional characterization of CMV-associated adaptive NK cells in rhesus macaques

status: Public on Sep 07 2022

city: Göttingen

first: Lutz

last: Walter

address: Kellnerweg 4

department: Primate Genetics

country: Germany

institute: German Primate Center

summary: Previous research on adaptive NK cells in rhesus macaques suffered from the lack of specific antibodies to differentiate between inhibitory CD94/NKG2A and stimulatory CD94/NKG2C heterodimeric receptors. Recently we reported an expansion of NKG2C receptor-encoding genes in rhesus macaques, but their expression and functional role on primary NK cells remained unknown due to this deficit. Thus, we established monoclonal antibodies 4A8 and 7B1 which show identical specificities and bind to both NKG2C-1 and NKG2C-2 but neither react with NKG2C-3 nor NKG2A on transfected cell lines. Using a combination of 4A8 and Z199 antibodies in multicolor flow cytometry we detected broad expression (4-73%) of NKG2C-1 and/or NKG2C-2 (NKG2C-1/2) on primary NK cells in rhesus macaques from our breeding colony. Stratifying our data to CMV-positive and CMV-negative animals, we noticed a higher proportion (23-73%) in primary NK cells expressing NKG2C-1/2 in CMV+ as compared to CMV- macaques (4-5%). These NKG2C-1/2-positive NK cells in CMV+ macaques are characterized by lower expression of IL12RB2, ZBTB16 and SH2D1B as well as high expression of IFN-gamma, indicating that antibody 4A8 detects CMV-associated adaptive NK cells. Single cell RNA seq data of 4A8-positive NK cells from a CMV-positive individual demonstrated that a high proportion of these adaptive NK cells transcribe in addition to NKG2C-1/2 also NKG2C-3, but interestingly NKG2A as well. Remarkably, NKG2C-1 and in particular NKG2C-2 have a higher affinity to Mamu-E as compared to NKG2A. Primary NK cells exposed to Mamu-E-expressing target cells displayed strong degranulation as well as IFN-gamma expression of 4A8+ adaptive NK cells of rhCMV+ animals that was not evident in rhCMV- animals. Thus, despite co-expression of inhibitory and stimulatory CD94/NKG2 receptors the higher number of different stimulatory NKG2C receptors and their higher binding avidity to Mamu-E outreach inhibitory signaling via NKG2A. These data demonstrate the evolutionary conservation of the CMV-driven development of NKG2C-positive adaptive NK cells with particular molecular signatures in primates and with changes in gene copy numbers and regulation and in ligand binding strength of NKG2C isotypes. Thus, rhesus macaques represent a suitable and valuable nonhuman primate animal model to study the CMV-NKG2C liaison in vivo.

overall_design: Rhesus monkey (Macaca mulatta) blood NK cells analyzed using scRNAseq (ICELL8, Takara)

first: Mohammad

middle: Z

last: Hasan

first: Charlotte

last: Hölterman

first: Beatrix

last: Petersen

first: Annette

last: Schrod

first: Kerstin

last: Mätz-Rensing

first: Artur

last: Kaul

first: Gabriela

last: Salinas

first: Ralf

last: Dressel


title: Transcriptome analysis of PK-15 cells expressing CSFV NS4A

city: Zhengzhou

first: Wang

last: Dong

address: No. 6 Longzi Hubei Road

institute: Henan University of Animal Husbandry and Economy

summary: We used RNA-seq to investigate the functional role of CSFV NS4A in PK-15 cells.we used RNA-seq to investigate the functional role of CSFV NS4A in PK-15 cells.

overall_design:
A total of 3893 differentially expressed genes (DEGs) were identified in PK-15 cells expressing NS4A, compared to empty vector (NC) transfectant.
Comparative gene expression profiling analysis of RNA-seq data for PK-15 cells expressing NS4A and PK-15 cells expressing empty vector

first: Huifang

last: Lv


title: Ribosome profiling analysis of mouse brain neocortex during development

status: Public on Sep 06 2022

city: Berlin

first: Matthew

middle: Lee

last: Kraushar

state: Berlin

address: Ihnestrasse 63-73

department: Department of Computational Molecular Biology

institute: Max Planck Institute for Molecular Genetics

summary: We perform Ribosome Profiling (Ribo-seq) analysis of mouse brain neocortex during development embyonic days 12.5, 14, 15.5, 17, and postnatal day 0 in biological duplicate

overall_design: Each biological replicate analyzed for ribosome-protected mRNA fragments after RNAse digestion (Ribosome Profiling, Ribo-seq)

first: Kraushar

last: Matthew


title: Cell types and molecular architecture of the Octopus bimaculoides visual system

status: Public on Sep 04 2022

city: Eugene

first: Cristopher

last: Niell

state: Oregon

institute: University of Oregon

summary: Cephalopods have a remarkable visual system, with a camera-type eye, high acuity vision, and a wide range of sophisticated visual behaviors. However, the cephalopod brain is organized dramatically differently from that of vertebrates, as well as other invertebrates, and little is known regarding the cell types and molecular determinants of their visual system organization beyond neuroanatomical descriptions. Here we present a comprehensive single-cell molecular atlas of the octopus optic lobe, which is the primary visual processing structure in the cephalopod brain. We combined single-cell RNA sequencing with RNA fluorescence in situ hybridization to both identify putative molecular cell types and determine their anatomical and spatial organization within the optic lobe. Our results reveal six major neuronal cell classes identified by neurotransmitter/neuropeptide usage, in addition to non-neuronal and immature neuronal populations. Moreover, we find that additional markers divide these neuronal classes into subtypes with distinct anatomical localizations, revealing cell type diversity and a detailed laminar organization within the optic lobe. We also delineate the immature neurons within this continuously growing tissue into subtypes defined by evolutionarily conserved fate specification genes as well as novel cephalopod- and octopus- specific genes. Together, these findings outline the organizational logic of the octopus visual system, based on functional determinants, laminar identity, and developmental markers/pathways. The resulting atlas presented here delineates the “parts list” of the neural circuits used for vision in octopus, providing a platform for investigations into the development and function of the octopus visual system as well as the evolution of visual processing.

overall_design: Characterization of cell types in the Octopus bimaculoides visual system through both gene expression profiling analysis of single-cell RNA sequencing data and RNA fluorescent in situ hybridization

first: Jeremea

middle: O

last: Songco-Casey

first: Gabrielle

last: Coffing

first: Denise

middle: M

last: Piscopo

first: Judit

middle: R

last: Pungor

first: Andrew

middle: D

last: Kern

first: Adam

last: Miller


title: Transcriptomic analysis of pals-22 and pals-25 mutants in C. elegans

status: Public on Sep 02 2022

city: San Diego

first: Spencer

middle: S

last: Gang

state: California

address: Bonner Hall #4205, 7835 Trade St. Suite 100

department: Division of Biological Sciences

institute: University of California, San Diego

summary: We used RNA-seq to identify gene expression changes in C. elegans pals-22 mutants, pals-22 pals-25 double mutants, and pals-25 mutants

overall_design: Comparative gene expression of pals-22(jy3), pals-22(jy3) pals-25(jy9), and pals-25(jy111) mutants to wild-type animals at the L4 life stage

first: Manish

last: Grover

first: Kirthi

last: Reddy

first: Deevya

last: Raman

first: Ya-Ting

last: Chang

first: Damian

last: Ekiert

first: Michalis

last: Barkoulas

first: Emily

last: Troemel


title: CITE-seq ex vivo engineered human plasma cells

city: Seattle

first: Rene

last: Cheng

state: WA

address: 1900 9th AVE

institute: Seattle children research institute

summary: Due to their unique longevity and capacity to secrete high levels of protein, plasma B cells play have the potential to be used as a cell therapy for protein replacement. Here, we show that ex vivo engineered human plasma cells exhibited transcriptional features of long-lived plasma cells.

overall_design: Ex vivo differentiated B cells day 7 and day13.

first: Richard

last: James


title: 7 dpf RNA Sequencing - Zebrafish FASD Model

status: Public on Sep 01 2022

city: Boston

first: Olivia

last: Weeks

address: 77 Ave Louis Pasteur, New Research Building #458

institute: Brigham and Women's Hospital

summary: Purpose: The transcriptional alterations underlying physiological changes in fetal alcohol spectrum disorder are largely unidentified. Here we perform RNA sequencing on 0% and 1% EtOH (12 hpf - 5 dpf) zebrafish at 7dpf stages in order to identify significant transcriptional alterations.

overall_design: All fish were treated with 0% or 1% EtOH from 12 hpf - 5 dpf and removed from EtOH at 5 dpf. From day 5 - 7, larvae were placed in system water (28C) containing paramecia starter culture. 3 biological samples, each containing a group of pooled whole embryos, were included in the study.

middle: G

first: Wolfram

last: Goessling

first: Isaac

last: Oderberg


title: DNA dioxygenases Tet2/3 control epithelial differentiation [CUT&TAG]

first: Guodong

last: Chen

summary:
Execution of lineage-specific differentiation programs requires tight coordination between many regulators including Ten-eleven translocation (TET) family enzymes catalyzing 5-methylcytosine oxidation in DNA. Here, by using Keratin 14-Cre-driven ablation of Tet genes in skin epithelial cells, we demonstrate that ablation of Tet2/Tet3 result in marked alterations of hair shape and length followed by hair loss. We show that through DNA demethylation, Tet2/Tet3 control chromatin accessibility, Dlx3 binding and promoter activity of the Krt25 and Krt28 genes regulating hair shape, as well as regulate interactions between the Krt28 gene promoter and distal enhancer. Moreover, Tet2/Tet3 also control three-dimensional chromatin topology in Keratin type I/II gene loci via DNA methylation-independent mechanisms. These data demonstrate the essential roles for Tet2/3 in establishment of lineage-specific gene expression program and control of Dlx3/Krt25/Krt28 axis in hair follicle epithelial cells and implicate modulation of DNA methylation as a novel approach for hair growth control.
We performed CUT&Tag from sorted hair matrix keratinocytes of wild type and TetDKO (Krt14-Cre, Tet2 flox/flox, Tet3 flox/flox) mice at P4.5 which are treated or untreated with 5-aza.

overall_design: CUTANA™ CUT&Tag kit is purchased from Epicypher and the assay is carried out as the described manual. Briefly, nuclei of 100K sorted hair matrix keratinocytes are isolated with Nuclear Extraction Buffer (20 mM HEPES-KOH pH 7.9, 10 mM KCl, 0.1% TritonX-100, 20% Glycerol, 0.5 mM Spermidine and 1x cOmplete™ Protease Inhibitor Cocktail) and attached to Concanavalin A linked magnetic beads which are activated with Bead Activation Buffer (20 mM HEPES-KOH pH 7.9, 10 mM KCl, 1mM CaCl2, 1 mM MnCl2), and beads attaching nuclei are incubated with 0.5 µg of primary antibody in the Digitonin150 Buffer (20 mM HEPES pH 7.5, 150 mM NaCl, 0.5 mM Spermidine, 0.01% Digitonin, and 1x cOmplete™ Protease Inhibitor Cocktail) containing 2 mM EDTA at 4 °C with gentle shaking for overnight and 0.5 µg of secondary antibodies in Digitonin150 Buffer at room temperature for 30 minutes successively. After washed twice with Digitonin150 Buffer, beads are incubated with 2.5 μl of pAG-Tn5 in Wash300 Buffer (20 mM HEPES pH 7.5, 300 mM NaCl, 0.5 mM Spermidine, and 1x cOmplete™ Protease Inhibitor Cocktail) containing 0.01% Digitonin at room temperature for 1 hour.  Beads attaching nuclei are washed with Wash300 Buffer containing Digitonin twice and incubated in Wash300 Buffer containing 10 mM MgCl2 at 37 °C for 1 hour to perform chromatin tagmentation. Beads are washed with TAPS Buffer (10 mM TAPS, pH 8.5 and 0.2 mM EDTA), and the DNA is released in 5 μl of SDS Release Buffer (10 mM TAPS, pH 8.5 and 0.1% SDS) at 55 °C for 1 hour and quenched with 15 μl of 0.67% Triton-X 100. DNA libraries are amplified for 18-21 cycles with 25 μl of NEBNext® High-Fidelity 2x PCR Master Mix (NEB, #M0541) and 2.5 μl of unique i5 and i7 index primers for Illumina. Paired-end sequencing is performed in Illumina NextSeq2000 P2 PE50 system (2 x 50bp). Antibodies: Dlx3 (Abcam, #ab66390), Lef1 (Cell Signaling, #2230), Pol2Ser5 (Abcam, #ab5408), CTCF (Diagenode, #C15410210), H3K4me1 (Diagenode, #C1541094), H3K4me3 (Abcam, #ab12209), H3K27ac (Abcam, #ab4729) and H3K27me3 (Millipore, #07-449).

first: Iqra

last: Fatima

first: Qin

last: Xu

first: Michael

last: Fessing

first: Andrei

last: Mardaryev

first: Andrey

last: Sharov

first: Elena

last: Rozhkova

first: Guoliang

first: Vladimir

last: Botchkarev


title: Changes in Treg gene expression upon deletion of Usp22 and Usp21

city: Chicago

first: Deyu

last: Fang

address: 303 E Chicago Avenue

institute: Northwestern University

summary: Usp22 and Usp21 are important for Foxp3 stability. We used RNA sequencing to determine the broad changes upon Usp22 and Usp21 deletionWe then performed gene expression profiling analysis using data obtained from RNA-seq and compared to each other

overall_design: Naïve natural Treg cells were isolated from murine spleens of Usp21-KO and Usp22 and Usp21-double KO mice. RNA isolated and sent for sequencing

last: Montauti

first: Shuvam

last: Chaudhuri

first: Sam

middle: E

last: Weinberg


title: Gene expression changes in beta-blocker treated neonatal hearts

status: Public on Aug 30 2022

city: Cincinnati

first: Mei

last: Xin

state: OH

address: 3333 Burnet Avenue

department: EHCB

institute: Cincinnati Children's Hospital

summary: The regeneration potential of the mammalian heart is limited. We found that treatment of beta-blocker robustly enhanced cardiomyocyte proliferation and promoted cardiac regeneration post myocardial infarction. To investigate the gene expression changes, we performed RNA-seq using the hearts treated with beta-blocker for 7 days.

overall_design: mRNA profiles of neonatal control and beta-blocker treated hearts at P7

first: Masahide

last: Sakabe


title: Chromosome 9p21.3 Coordinates Cell Intrinsic and Extrinsic Tumor Suppression

status: Public on Sep 08 2022

city: New York

first: Yu-Jui

last: Ho

address: 417 E 68th St

department: Cancer Biology & Genetics Program

institute: Memorial Sloan Kettering Cancer Center

summary: Somatic chromosomal deletions are prevalent in cancer, yet their effects are poorly understood. The most prominent homozygous deletions affect chromosome 9p21.3 and eliminate the CDKN2A/B tumor suppressor genes, thus disabling a cell intrinsic barrier to tumorigenesis. However, half of 9p21.3 deletions encompass a cluster of 16 type I interferons (IFNs) whose co-deletion remains unexplored. To functionally dissect 9p21.3 and other genomic deletions, we developed MACHETE (Molecular Alteration of Chromosomes with Engineered Tandem Elements), a genome engineering strategy that enables flexible modeling of megabase-sized deletions. Applying MACHETE to a syngeneic mouse model of pancreatic cancer, we show that concomitant loss of the IFN cluster with Cdkn2a/b enhanced immune evasion, metastasis, and immunotherapy resistance compared to Cdkn2a/b deletions alone. Mechanistically, IFN co-deletion disrupted type I IFN signaling in the tumor microenvironment, leading to marked changes in infiltrating immune cells and escape from CD8+ T cell surveillance, effects largely driven by the poorly understood interferon epsilon (Ifne). These results reveal how IFN-encompassing 9p21.3 deletions disable cell intrinsic and extrinsic tumor suppression, thereby providing a pervasive route for immune evasion, metastasis, and immunotherapy resistance. Our study provides a framework for interrogating large deletion events in cancer and beyond.

overall_design: RNA-seq analysis was performed from KrasG12D; sgP53; dS or KrasG12D; sgP53; dL tumors, which were harvested from immune-competent mice treated with control IgG or anti-IFNAR1 blocking antibody. Bulk tumor pieces were flash frozen on dry ice and stored at -80C. Tissues were then mechanically disrupted in Trizol and RNA was extracted following manufacturer’s instructions. RNA integrity was analyzed with an Agilent 2100 Bioanalyzer. Samples that passed QC were then used for library preparation and sequencing. Samples were barcoded and run on a HiSeq (Ilumina) in 76 bp SE run, with an average of 50 million reads per sample. RNA-Seq data was then trimmed by removing adapter sequences and reads were aligned to the mouse genome (GRCm38.91; mm10), and transcript counts were used to generate an expression matrix.

first: Francisco

last: Barriga

first: Kaloyan

last: Tsanov

first: Yu

middle: H

last: Jo

first: Noor

last: Sohail

first: Amy

last: Zhang

first: Timour

last: Baslan

first: Alexandra

last: Wuest

first: Isabella

last: Del Priore

first: Brigita

last: Meskauskaite

first: Geulah

last: Livshits

first: Direna

last: Alonso-Curbelo

first: Janelle

last: Simon

first: Almudena

last: Chaves-Perez

first: Dafna

last: Bar-Sagi

first: Christine

last: Iacobuzio-Donahue

first: Faiyaz

last: Notta

first: Ronan

last: Chaligne

first: Roshan

last: Sharma

first: Dana

last: Pe'er

first: Scott

middle: W

last: Lowe


title: Gene expression analysis of male and female bone endothelial cells from 2 and 65 weeks old mice

status: Public on Aug 29 2022

city: London

first: LMS

last: Bioinformatics Core

address: Hammersmith Hospital Campus, Du Cane Road

department: Bioinformatics Core

country: United Kingdom

institute: MRC London Institute of Medical Sciences

summary: Bone endothelial cells (ECs) were purified from male and female mice of 2 weeks and 65 weeks old. RNA sequencing was performed to understand sex- dependent gene expression pattern in bone ECs during development and ageing to understand blood vesssel role in sex differences observed in bones. Further comparison of young and old mice would beneficial in understanding sex specific ageing mechanism.

first: Saravana

middle: K

last: Ramasamy

first: Gopuraja

last: Dharmalingham

first: Yi-Fang

last: Wang


title: Parenteral BCG vaccine induces respiratory mucosal-resident memory macrophages and trained innate immunity via the gut-lung axis

city: Hamilton

first: Zhou

last: Xing

address: MDCL building, 1280 Main St W

institute: McMaster University


title: Multimodal chromatin profiling  using nanobody-based single-cell CUT&Tag

city: Stockholm

first: Marek

last: Bartosovic

address: Solnavagen 9

country: Sweden

institute: Karolinska Institute

summary: Probing epigenomic marks such as histone modifications at a single cell level in thousands of cells has been recently enabled by technologies such as scCUT&Tag. Here we developed a multimodal and optimized iteration of scCUT&Tag called nano-CT (for nano-CUT&Tag) that allows simultaneous probing of three epigenomic modalities at single-cell resolution, using nanobody-Tn5 fusion proteins. nano-CT is compatible with starting materials as low as 25 000 cells and has significantly higher resolution than scCUT&Tag, with a 16-fold increase in the number of fragments per cells. We used nano-CT to simultaneously profile chromatin accessibility, H3K27ac and H3K27me3 in a complex tissue - juvenile mouse brain. The obtained multimodal dataset allowed for discrimination of more cell types/states that scCUT&Tag, and inference of chromatin velocity between ATAC and H3K27ac in the oligodendrocyte (OL) lineage. In addition, we used nano-CT to deconvolute H3K27me3 repressive states and infer two sequential waves of H3K27me3 repression at distinct gene modules during OL lineage progression. Thus, given its high resolution, versatility, and multimodal features, nano-CT allows unique insights in epigenetic landscapes in different biological systems at single cell level.

overall_design: Multimodal profiling of two histone modifications and open chromatin in single cells by nano-CT technology

first: Goncalo

last: Castelo-Branco


title: Multiomic Profiling Reveals Unexpected Increased Chromatin Accessibility Downstream from the 3’ LTR OF Active HIV-1 Proviruses (Hi-C)

status: Public on Aug 26 2022

city: La Jolla

first: Jesse

last: Dixon

state: CA

address: 10010 N. Torrey Pines Rd.

institute: Salk Institute for Biological Studies

summary: The three-dimensional structure of the genome is a regulator of transcription and cell function; HIV-1 infection can influence host cell function, but the degree to which this is mediated through changes to host chromatin architecture is unclear. We interrogated genome-wide chromatin organization and the structure of chromatin around latently infected HIV-1 integration sites using Hi-C and ATAC-seq and combined these data with RNA transcriptional analysis of the provirus and neighboring genes in HIV-inducible cellular models. We found chromatin interaction networks around integrated HIV-1 are predominantly preserved with respect to uninfected cells, proving the lack of an obligate association between latent integration and major chromatin remodeling. Instead, we find that induction of proviral transcription may lead to local changes in chromatin accessibility downstream from the 3’ LTR, demonstrating that HIV-1 can alter local cellular chromatin structure post-integration. Using long-read Nanopore RNA-seq, we interrogated the local host and HIV-1 transcriptomes and observed that 1-5% of HIV-1 transcripts initiated at the 5’ LTR promoter extended into the flanking cellular genome, generating chimeric virus-host RNAs. Thus, integration leading to latency (and provirus activation) may not lead to obligate global chromatin rearrangements; we also observed, previously unreported, novel changes in chromatin accessibility during HIV-1 transcription.

overall_design: Analysis of Hi-C data in Jurkat T-cells and Jurkat-derived JLat models during TNF-alpha induced reversal of HIV latency.


title: NCP26 is a potent prolyl-tRNA synthetase inhibitor and induces pro- apoptotic responses with in vivo potency in multiple myeloma

city: Oxford

first: Adam

last: Cribbs

state: Oxfordshire

address: Botnar Research Centre

department: NDORMS

institute: University of Oxford

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


title: NCP26 is a potent prolyl-tRNA synthetase inhibitor and induces pro- apoptotic responses with in vivo potency in multiple myeloma. Bulk RNA-seq data.

summary: Glutamyl-prolyl-tRNA synthetase (EPRS) is a unique bifunctional aminoacyl-tRNA synthetase which catalyzes the ATP-dependent aminoacylation of glutamyl and prolyl residues to cognate tRNAs, constituting fundamental building blocks in protein synthesis. Genomic data suggest that EPRS promotes disease progression and is associated with poor prognosis in multiple myeloma (MM), while downregulation of EPRS triggers MM cell apoptosis implying an amino acid starvation response as a central mechanism. We developed a novel ATP competitive inhibitor, NCP26, targeting the prolyl-tRNA synthetase (PRS) component. The inhibitor demonstrates significant anti-tumor activity against human MM cell lines, regardless of their sensitivity to other therapeutic agents, against patient MM cells and is active in vivo using a MM xenograft mouse model. In contrast to the proline competitive PRS inhibitor halofuginone, NCP26 effects are not altered by exogenous proline levels. NCP26 treatment induces G0/G1 cell cycle arrest followed by apoptotic MM cell death, evidenced by depletion of mitochondrial membrane potential as well as caspase and PARP cleavage. Using combined transcriptomic and proteomic approaches we demonstrate a complex phenotypic response involving multiple pathways in protein quality control which center around the ribosome as integrating hub. Furthermore, we identified multiple proline rich motif containing protein targets of NCP26 as downstream effectors. These include myeloma survival factors such as syndecan 1 (SDC1, CD138), or basic helix-loop-helix transcription factors such as MYC, and transcription factor 3 (TCF3) suggesting that acute blockade of prolyl- aminoacylation evokes a complex pro-apoptotic response beyond the canonical amino acid starvation and integrated stress response. Taken together, our pre-clinical studies validate EPRS as a possible therapeutic target and provide the framework for evaluation of PRS inhibitors in a clinical setting.

overall_design: 22 unique conditions, in triplicate = 6 samples total. 2 cell types (WT AMO-1 and CFZ-resistant L363) and two time points. CFZr treated with DMSO (control), NCP22, NCP26, MAZ1392 (Halofuginone) and MAZ1805 (Halofuginol). WT AMO-1 treated with DMSO (control), NCP22, NCP26, MAZ1392 (Halofuginone) and MAZ1805 (Halofuginol) and carfilzomib.

first: Anna

last: James-Bott


title: Quantitative real-time PCR analysis of tRNA abundance in the developing mouse neocortex

first: Matthew

last: Kraushar

summary: qRT-PCR analysis of mouse brain neocortex during development at embyonic days 12.5, 14, 15.5, 17, and postnatal day 0 in biological duplicate

overall_design: Each biological replicate analyzed for tRNA abundance using a qRT-PCR protocol (Arraystar Inc.)

first: Dermot

last: Harnett


title: Image-seq: spatially-resolved single cell sequencing guided by in situ and in vivo imaging

status: Public on Aug 25 2022

first: Shenglin

last: Mei

address: 10 Shattuck Street

institute: Harvard medical school (DMBI)

summary:
We collected bone marrow samples from mice using the Image-seq technology that we developed for spatial transcriptomics. We isolated samples from the anatomically distinct D, M and R cavities that were recently identified by in vivo imaging (Christodoulou et al, Nature, 2020, 578, 278-283). We compared these samples in aggregated form (i.e. all Image-seq to all WT control) to whole calvarial bone marrow samples from wild-type (WT) mice and mice injected with a combination of tetracycline and alizarin red (used also to identify different cavity types). The transcriptional composition of all samples was assessed using single-cell RNA-seq (scRNA-seq).
We collected bone marrow samples from mice using the Image-seq technology that we developed for spatial transcriptomics. We isolated samples from regions with proliferating, non-proliferating and intermediate AML cells (see manuscript for details), and sorted single GFP+ AML cells into individual wells filled with lysis buffer by flow cytometry. We analyzed the transcriptional composition of each individual cell using single-cell RNA-seq.

overall_design: Bone marrow were disaggregated and profiled by 10X Genomics Single cell 3 gene expression V2.  Smartseq v4 protocol were used  (https://www.takarabio.com) to profile rare (<0.01% leukemic burden) acute myeloid leukemia (AML) cells and BM stromal cells.

first: Christa

last: Haase


title: Metabolic regulation by p53 prevents R-loop-associated genomic instability [ATAC-Seq]

status: Public on Aug 24 2022

city: Rome

first: panatta

last: emanuele

state: RM

address: Via Montpellier, 1

country: Italy

institute: University of Rome "Tor Vergata"

summary: To investigate the role of p53 in maintaining genome integrity we deleted p53 via doxycycline-inducible short hairpin (sh)RNA targeting p53 (KPshp53 cells), and we treat the cells with the DNA demethylating molecule 5-Aza-2′-deoxycytidine (AZA or dC)

overall_design: We performed comparative gene expression profiling (RNAseq, 3 biological replicates/sample) and differential genome-wide accessibility profiling (ATAC-seq, 2 biological replicates/sample) on shCTR_NT, shCTR_AZA, shp53_NT, shp53_AZA

first: Emanuele

last: Panatta

first: Alessio

last: Butera

first: Eleonora

last: Mammarella

first: Consuelo

last: Pitolli

first: Alessandro

last: Mauriello

first: Richard

last: Knight

first: Gerry

last: Melino

first: Ivano

last: Amelio


title: NCP26 is a potent prolyl-tRNA synthetase inhibitor and induces pro- apoptotic responses with in vivo potency in multiple myeloma. Single-cell RNA-seq data BM samples.

overall_design: 3 separate experiments. Experiment 1 = 1 patient, 7 treatment samples. Experiment 2 = 1 patient, 11 treatment samples. Experiment 3= 2 patients, 16 different treatment samples = 32 samples


title: Metabolic regulation by p53 prevents R-loop-associated genomic instability [RNA-Seq]


title: Expression data for tumor and normal tissue samples from deceased sepsis and non-sepsis colorectal cancer patients

city: Honolulu

first: Yurii

last: Shvetsov

state: HI

address: 701 Ilalo St

department: Cancer Center

institute: University of Hawaii

summary:
The mortality risk from cancer-associated sepsis is differentially affected by individual cancer sites and host responses to sepsis are heterogenous. Native Hawaiians have a 2-fold higher mortality risk from cancer-associated sepsis than Whites and also higher mortality risk from colorectal cancer (CRC). We investigated this disparity by examining ethnic variation in the transcriptome of patients with CRC-associated sepsis and its relation to survival and genetic diversity.
We conducted transcriptomic profiling of CRC tumors and adjacent non-tumor tissue obtained from adult patients of Native Hawaiian and Japanese ethnicity who died from cancer-associated sepsis. We examined differential gene expression in relation to patient survival and sepsis disease etiology.

overall_design: FFPE tumor and adjacent normal tissue was obtained from 30 Hawaiian and Japanese CRC patients. 15 patients who died of sepsis were age and ethnicity matched to patients with non-sepsis cause of death.

first: Michelle

middle: L

last: Matter

middle: B


title: TLR7  agonism accelerates disease in a mouse model of primary Sjögren’s syndrome and  drives expansion of age-associated B cells

city: Dallas

first: Chengsong

last: Zhu

state: TX

address: 5323 Harry Hines Blvd

department: Immunology

institute: UT Southwestern Medical Center

summary: Primary Sjögren’s syndrome (pSS) is a systemic autoimmune disease characterized by chronic inflammation of exocrine tissue, resulting in loss of tears and saliva. Patients also experience many extra-glandular disease manifestations. Treatment for pSS is palliative, and there are currently no treatments available that target disease etiology. Previous studies in our lab demonstrated that Myd88 is crucial for pSS pathogenesis in the NOD.B10Sn-H2b (NOD.B10) pSS mouse model, although the way in which Myd88-dependent pathways become activated in disease remains unknown. Based on its importance in other autoimmune diseases, we hypothesized that TLR7 activation accelerates pSS pathogenesis. We administered the TLR7 agonist Imiquimod (Imq) or sham treatment to pre-disease NOD.B10 females for 6 weeks. Parallel experiments were performed in age and sex-matched C57BL/10 controls. Imq-treated pSS animals exhibited cervical lymphadenopathy, splenomegaly, and expansion of TLR7-expressing B cells. Robust lymphocytic infiltration of exocrine tissues, kidney and lung was observed in pSS mice following treatment with Imq. TLR7 agonism also induced salivary hypofunction in pSS mice, which is a hallmark of disease. Anti-nuclear autoantibodies, including Ro (SSA) and La (SSB) were increased in pSS mice following Imq administration.  Cervical lymph nodes from Imq-treated NOD.B10 animals demonstrated an increase in the percentage of activated/memory CD4+ T cells. Finally, aged-associated B cells (ABCs) were expanded in the spleens of Imq-treated pSS mice. Thus, activation of TLR7 accelerates local and systemic disease and promotes expansion of the ABC subset in pSS.

sample_organism:
Mus musculus
synthetic construct

overall_design:
C57BL/10 and NOD.B10H2b/J mice were treated with a sham cream (Con) or a cream containing 5% imiquimod (Imq) for 6 weeks beginning at 6 weeks of age. The cream was administered epicutaneously 3X/week by application to the ear. Animals were euthanized at 13 weeks of age and sera were harvested. Sera were harvested from BL/10 sham-treated ( = 6), BL/10 Imq-treated (n = 6), NOD.B10 sham-treated (n = 9) and NOD.B10 Imq-treated mice (n = 8). PBS was used as negative control for the array.
Each gpr file includes the raw signal data for 16 arrays (samples).  Independent samples are identified in the .gpr file with the "Block" column ID included on each sample page as a characteristic.

first: Jill

last: Kramer


title: mTOR-neuropeptide Y signaling sensitizes nociceptors to drive neuropathic pain development

city: Hangzhou

first: Lunhao

last: Chen

state: Zhejiang

address: 866 Yuhangtang Road

institute: Zhejiang University

summary: Neuropathic pain is a refractory condition that involves de novo protein synthesis in the nociceptive pathway. The mechanistic target of rapamycin (mTOR) is a master regulator of protein synthesis; however, mechanisms underlying its role in neuropathic pain remain elusive. Using spared nerve injury-induced neuropathic pain model, we found mTOR activation in large-diameter dorsal root ganglion (DRG) neurons and spinal microglia. However, selective ablation of mTOR in DRG neurons, rather than microglia, alleviated neuropathic pain. Combining transcriptomic profiling, electrophysiological recording and pharmacologic manipulations, we demonstrated that activated mTOR promoted neuropeptide Y (NPY) induction in mechanoreceptors and that NPY acted on Y2 receptors (Y2R) but not Y1R to enhance nociceptor excitability. Peripheral replenishment of NPY reversed pain alleviation upon mTOR removal, whereas Y2R antagonists prevented its function. Our findings reveal an unexpected link between mTOR and NPY in promoting nociceptor sensitization and neuropathic pain, through NPY/Y2R signaling-mediated intra-ganglionic transmission.

overall_design: DRG mRNA profiles of Mtorfl/fl and Adv-Cre::Mtorfl/fl mice following spared nerve injury

first: Yaling

last: Hu

first: Siyuan

last: Wang

first: Kelei

last: Cao

first: Weihao

last: Mai

first: Weilin

last: Sha

first: Huan

last: Ma

first: Yong-Jing

last: Gao

first: Shumin

last: Duan

first: Yue

first: Zhihua


title: Nuclear corepressors NCOR1/NCOR2 regulate B cell development, maintain genomic integrity, and prevent transformation

city: Minneapolis

first: Todd

middle: P

last: Knutson

state: MN

address: 117 Pleasant St SE

department: Minnesota Supercomputing Institute

institute: University of Minnesota

summary: The nuclear corepressors NCOR1 and NCOR2 interact with transcription factors involved in B cell development and potentially link these factors to alterations in chromatin structure and gene expression. Herein we demonstrate that NCOR1/2 deletion limits B cell differentiation via impaired recombination, attenuates pre-BCR-signaling, and enhances STAT5-dependent transcription. Furthermore, NCOR1/2-deficient B cells exhibited derepression of EZH2-repressed gene modules, including the p53 pathway. These alterations resulted in aberrant Rag1 and Rag2 expression and accessibility in proliferating cells. Whole-genome sequencing of Ncor1/2 DKO B cells identified increased number of structural variants compared to wildtype B cells with cryptic recombination signal sequences. Finally, deletion of Ncor1 alleles in mice facilitated leukemic transformation, while human leukemias with less NCOR1 correlated with worse survival. NCOR1 mutations in human leukemia correlated with increased RAG expression and increased number of structural variants. These studies illuminate how the corepressors NCOR1/2 regulate B cell differentiation and provide insights into how NCOR1/2 mutations may promote B cell transformation.

overall_design: scATAC-Seq and scRNA-Seq WT, Ncor1-KO, Ncor1_Ncor2-DKO

first: Robin

last: Lee

first: Sarah

middle: A

last: Munro

first: Jeffrey

last: Miller

first: Lynn

last: Heltemes-Harris

first: Kristen

last: Jepsen

first: Michael

last: Farrar


title: Association of Presurgical Circulating MicroRNAs with 1-Year Postsurgical Pain Reduction in Lumbar Spinal Stenosis Patients with Spine Facet Osteoarthritis

city: Toronto

first: Mohit

last: Kapoor

state: Ontario

address: 60 Leonard Ave

institute: Schroeder Arthritis Institute

summary:
OBJECTIVE: Our objective was to sequence the plasma miRNome from 56 patients with lumbar spinal stenosis (LSS) to determine whether miRNA signatures at time of surgery predicted pain responses one year later.
MicroRNA libraries were prepared using the QIAseq miRNA Library Kit and sequenced on the Illumina NextSeq550. Counts were produced for microRNAs captured in miRBase

overall_design: MicroRNA sequencing was performed on 56 plasma samples of LSS patients, 30 of which had degenerative spondylolisthesis (DS).

first: Starlee

last: Lively

first: Marie

last: Milliot

first: Pratibha

last: Potla

first: Osvaldo

last: Espin-Garcia

first: Mehdi

middle: Y

last: Layghifard

first: Kala

last: Sundararajan

first: Helal

last: Endisha

first: Akihiro

last: Nakamura

first: Anthony

last: Perruccio

first: Raja

last: Rampersaud


title: CXCR4-CXCL12 axis promotes the efficient T cell reconstitution by the HSPCs immigration

city: Chongqing

first: jiangyong

last: he

address: Tiansheng Li Street of Beibei District

institute: southwest university

summary: T cells are essentially involved in safeguarding homeostasis through fighting against the pathogens and malignant cells. T cell immunodeficiency, especially their perturbation in the severe infection, irradiation, chemotherapy, and thymic atrophy in ageing, is detrimental. Therefore, strategies that enhance T cell reconstitution provide considerable benefit and warrant intensive investigation. Here, we constructed a T cells ablation model in Tg(coro1a:DenNTR) zebrafish via administrating a proper volume of metronidazole (MTZ). T cells completely recovered at 6.5 days post treatment (dpt). The nascent regenerated T cells were mainly derived from the immigration of hematopoietic stem/progenitor cells (HSPCs) in the kidney, the functional homologue of BM. cxcr4b, but not ccr9 nor ccr7, was drastically unregulated in the responsive HSPCs. Functional interference of CXCR4 via both genetic and chemical assays yielded limited influence in T lymphopoiesis but notably delayed T cells regeneration by a destroyed HSPCs migration. In contrast, hematopoietic providing cxcr4b in Tg:(coro1a:cxcr4b) accelerates thymus-replenishment of HSPCs. Correspondingly, Cxcl12b, the ligand of Cxcr4, exhibited impressive increment presentation in the thymic epithelial cells of injured animals. Interfering or overacting Cxcl12b in either cxcl12b-/- mutants or Tg(hsp70:cxcl12b) recapitulated the similarly compromised or promoted T cells recovery as that seen in Cxcr4b scenario. Therefore, CXCR4-CXCL12 axis plays a crucial role in promoting thymocyte reconstitution but did not influence T cell development. Our study discloses a special role of CXCR4-CXCL12 signaling in promoting T cells recovery and provides a promising target to mitigate T cell immunodeficiency.

overall_design: 6  Smart-seq data of kidney hematopoietic cells in DMSO and MTZ treated zebrafish

first: Li

last: Li

first: Fangying

last: Zhao

first: Yafang

last: Lu

first: Zhifan

first: Jiangyong

last: He

first: Nianfei

last: Cui

first: Lingfei

last: Luo


title: Mediator facilitates transcription initiation at most promoters via a Tail-independent mechanism

status: Public on Aug 22 2022

city: Oklahoma City

first: Rafal

last: Donczew

state: OK

address: 825 NE 13th St

department: Cell Cycle and Cancer Biology

institute: Oklahoma Medical Research Foundation

overall_design: Refer to individual Series


title: Pancreatic cancer cells upregulation of LPA receptor 4 in response to stress create a tumor-initiating niche

status: Public on Aug 15 2022

first: Chengsheng

last: Wu

address: 2880 Torrey Pines Scenic Dr,

institute: University of California San Diego

summary: We report that the lysophosphatidic acid receptor 4 (LPAR4) is specifically upregulated on cells exposed to environmental stress or cancer drugs where it promotes stress tolerance, self-renewal, and tumor initiation. We find that ectopic LPAR4-expressing tumors display an enrichment of key extracellular matrix (ECM)-related genes that are established drivers of cancer stemness, and surprisingly do not require stimulation with the canonical LPAR4 ligand, LPA. From this RNAseq data, we observed that LPAR4 promotes a subset of genes, mainly extracellular matrix related genes, independent of its ligand LPA in patient-derived xenograft cells.

overall_design: 79E+EV vs. 79E+LPAR4 cells, treated with or without 1 µM LPA for 2 hours. Each group has 3 replicates, there are 12 samples in total. 79E+EV without LPA stimulation was the control group.

first: David

last: Cheresh


title: Small RNA-seq data of peripheral blood obtained from treatment-receiving IBD patients and control individuals (small RNA-Seq II)

status: Public on Aug 16 2022

city: Kiel

first: Simonas

last: Juzenas

address: Am Botanischen Garten 11

department: Institute of Clinical Molecular Biology

institute: Christian-Albrechts-University of Kiel

summary: Comperhensive analysis of blood transcriptomes obtained from treatment-receiving IBD patients.

overall_design: Total RNA extracted from peripheral blood samples (n = 242) was used to generate small RNA libraries employing TruSeq Small RNA Sample Preparation Kit (Illumina) according to the manufacturer’s instructions.

first: Andre

last: Franke

first: Georg

last: Hemmrich-Stanisak


title: Gene expression analysis of male and female bone endothelial cells from 12 weeks old mice

first: LMS

last: Bioinformatics Core

summary: Bone endothelial cells (ECs) were purified from male and female mice of 12 weeks old. RNA sequencing was performed to understand sex- dependent gene expression pattern in bone ECs during development and ageing to understand blood vesssel role in sex differences observed in bones. Further comparison of young and old mice would beneficial in understanding sex specific ageing mechanism.

first: Saravana

last: Ramasamy

first: Michelle

last: Hendriks

first: Yi-Fang

last: Wang


title: Genome-wide binding sites of Hypoxia inducible factor 1(HI1)  in renal epithelial cells.

city: Tokyo

first: Imari

last: Mimura

address: 7-3-1 Hongo Bunkyo-ku

department: Div. Nephrol and Endocrinol.

country: Japan

institute: The University of Tokyo

summary: We report the high-throughput profiling of HIF1 in human kidney-2 cells (HK-2) under normoxia and 1% 24 hours hypoxia. By obtaining over two billion bases of sequence from chromatin immunoprecipitated DNA, we generated genome-wide chromatin-state maps of HK-2 under normoxia and hypoxia. We found that HIF1 binds to both transcriptional starting sites and enhancer regions. This study provides novel insights into the epigenetic regulation of HIF1 in renal epithelial cells.

overall_design: We exposed HK-2 cells under 1% hypoxia for 24 hours. We collected mRNA of each samples and performed ChIP using HIF1 antibody. The ChIP DNA fragments were analyzed by NGS.

first: Natsuki

last: Kushida

first: Seitaro

last: Nomura

first: Hiroyuki

last: Aburatani


title: Rat ex vivo cataract model using DNA damage inducer

status: Public on Aug 11 2022

city: Fukui

first: Risa

last: Yamaoka

address: 3-9-1 Bunkyo

institute: University of Fukui

summary:
By adding MMS or Bleomycin, a DNA damage inducing agent, to the removed rat lens and culturing it, opacity was formed in the lens.
To investigate genes whose expression fluctuates when DNA damage is inflicted and causes lens opacity, microarray analysis was performed.

overall_design: Genes whose expression was altered by MMS or Bleomycin compared to the control sample were extracted.

first: Fumito

last: Kanada

first: Masaya

last: Nagaya

first: Yoshihiro

last: Takamura

first: Masaru

last: Inatani

last: Oki


title: Whole-transcriptome microarray analysis of Pseudomonas syringae pv. actinidiae CRAFRU 8.43 and Pseudomonas syringae pv. tomato DC3000 grown in apoplast-like conditions supplemented or not with kiwifruit or tomato extract

city: Verona

first: Elodie

last: Vandelle

state: Verona

address: Strada Le Grazie, 15

department: Department of Biotechnology

institute: University of Verona

summary: Purpose: The outcome of host–pathogen interactions is thought to reflect the offensive and defensive capabilities of both players. When plants interact with Pseudomonas syringae, several well-characterized virulence factors contribute to early bacterial pathogenicity, including the type III secretion system (T3SS), which must be activated by signals from the plant and environment to allow the secretion of virulence effectors. The manner in which these signals regulate T3SS activity is still unclear. Conlusion: the analysis revealed that the perception of plant signals from kiwifruit or tomato extracts anticipates T3SS expression in P. syringae pv. actinidiae compared to apoplast-like conditions

sample_organism: Pseudomonas syringae pv. tomato str. DC3000

sample_organism: Pseudomonas syringae pv. actinidiae str. CRAFRU8.43

overall_design: mRNA profiles of Psa and Pto cells incubated in apoplast-mimicking conditions were compared to mRNA profiles of Psa and Pto cells grown in apoplast-mimicking medium supplemented with kiwifruit or tomato extract. All mRNA profiles were generated from three independent biological replicates.

first: Maria

last: Puttilli

first: Davide

last: Danzi

first: Cristina

last: Correia

first: Jessica

last: Brandi

first: Daniela

last: Cecconi

first: Marcello

last: Manfredi

first: Emilio

last: Marengo

first: Conceição

last: Santos

first: Francesco

last: Spinelli

first: Annalisa

last: Polverari


title: Junctional Adhesion Molecule-A is dispensable for myeloid cell recruitment and diversification in the tumor microenvironment

status: Public on Aug 04 2022

city: Brussels

first: Daliya

last: Kancheva

address: Laarbeeklaan 103, Building E

country: Belgium

institute: Vrije Universiteit Brussel - VIB

summary: Junctional Adhesion Molecule-A (JAM-A), expressed on the surface of myeloid cells, is required for extravasation at sites of inflammation and may also modulate macrophage activation. Myeloid cell infiltration is a common feature of tumors that drives disease progression, but the function of JAM-A in this phenomenon and its impact on cancer-associated myeloid cells is little understood. Here we show that systemic cancer-induced inflammation in mice enhanced JAM-A expression in an IL1β-dependent manner specifically on circulating monocytes. However, using myeloid-specific JAM-A-deficient mice, we found that JAM-A was dispensable for recruitment of monocytes and other myeloid cells to tumors, in contrast to its reported role in inflammation. Single-cell RNA sequencing revealed that the loss of JAM-A did not influence the transcriptional reprogramming of tumor-infiltrating myeloid cells. Overall, our results support the notion that cancer-associated inflammation can modulate the phenotype of circulating immune cells and we demonstrate that tumors can bypass the requirement of JAM-A for myeloid cell recruitment.

overall_design: Sorted 7-AAD-CD45+ cells from subcutaneous Lewis lung carcinoma (LLC) tumors of myeloid-specific JAM-A  knockout mice (F11rfl/fl LysM-Cre+)  and their wild-type littermates (F11rfl/fl). Three mice per genotype were pooled for the library preparation.

first: Máté

last: Kiss

first: Aleksandar

last: Murgaski

first: Helena

last: Van Damme

first: Jan

last: Brughmans

first: Isabelle

last: Scheyltjens

first: Ali

last: Talebi

first: Els

last: Lebegge

first: Robin

last: Awad

first: Yvon

last: Elkrim

first: Sana

last: Arnouk

first: Pauline

last: Bardet

first: Cleo

last: Goyvaerts

first: Frank

last: Nana

first: Jo

last: Van Ginderachter

first: Damya

last: Laoui


title: Role of G-protein couple receptor 55 (GPR55) in B cells

status: Public on Aug 23 2022

city: Munich

first: Sabine

last: Steffens

state: Bayern

address: Pettenkoferstrasse 9b

department: Clinical Pathobiochemistry

institute: Institute for Cardiovascular Prevention-LMU Klinikum

summary:
Identifying novel pathways regulating the adaptive immune response in chronic inflammatory diseases such as atherosclerosis is of particular interest in view of developing new therapeutic drugs. Here we report that the lipid receptor GPR55 is highly expressed by splenic B cells and inversely correlates with atheroma plaque size in mice. In human carotid endarterectomy specimen, GPR55 transcript levels were significantly lower in unstable compared to stable carotid plaques. To study the impact of GPR55 deficiency in atherosclerosis, we crossed Gpr55 knockout mice with apolipoprotein E (ApoE) knockout mice and subjected the mice to Western diet for 4 to 16 weeks. Compared to ApoE-/- controls, ApoE-/-Gpr55-/- mice developed larger plaques with increased necrotic core size, associated with elevated circulating and aortic leukocyte counts. Flow cytometry, immunofluorescence and RNA-sequencing analysis of splenic B cells in these mice revealed a hyperactivated B cell phenotype with disturbed plasma cell maturation and immunoglobulin (Ig)G antibody overproduction. Collectively, these discoveries provide new evidence for GPR55 as key modulator of the adaptive immune response in atherosclerosis. Targeting GPR55 could be useful to limit inflammation and plaque progression in patients suffering from atherosclerosis
We  performed gene expression profiling analysis using data obtained from RNA-seq of female splenic CD19+  B cells isolated from 6 different ApoE-knockout mice and 6 ApoE and Gpr55 double knockout mice, all under C57BL/6J background.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for splenic CD19+ cells, comparing when GPR55 is present or not.

first: Guillamat-Prats

last: Raquel

first: Janjic

last: Aleksandar

first: Enard

last: Wolfgang

first: Steffens

last: Sabine


title: Single Cell RNA-Sequence Analyses Reveal Uniquely   Expressed Genes and Heterogeneous Immune Cell   Involvement in the Rat Model of Intervertebral   Disc Degeneration

status: Public on Aug 19 2022

city: St. Louis

first: Simon

last: Tang

state: MO

address: 425 S. Euclid Ave, Campus Box 8233, Tang Lab, Washington University School of Medicine

department: Orthopaedic Surgery

institute: Washington University in St. Louis

summary:
Purpose:  We used single-cell RNA-sequencing analysis of degenerating tissues of the rat IVD following lumbar disc puncture
Methods: Two control, uninjured IVDs (L2-3, L3-4) and two degenerated, injured IVDs (L4-5, L5-6) from each animal were examined either at the two- or eight-week post-operative time points. The cells from these IVDs were extracted and transcriptionally profiled at the single-cell resolution
Results: Unsupervised cluster analysis revealed the presence of four known cell types in both non-degenerative and degenerated IVDs based on previously established gene markers: IVD cells, endothelial cells, myeloid cells, and lymphoid cells. As a majority of cells were associated with the IVD cell cluster, sub-clustering was used to further identify the cell populations of the nucleus pulposus, inner and outer annulus fibrosus
Conclusions: . Differential gene expression analysis revealed multiple distinct cell types from the myeloid and lymphoid lineages, most notably macrophages and B lymphocytes, and demonstrated a high degree of immune specificity during degeneration. In addition to the heterogenous infiltrating immune cell populations in the degenerating IVD, the increased number of cells in the AF sub-cluster expressing Ngf and Ngfr, encoding for p75NTR, suggest that NGF signaling may be one of the key mediators of the IVD crosstalk between immune and neuronal cell populations

overall_design: Rats (n=8, male Sprague-Dawley, 8 weeks) underwent surgery for retroperitoneal exposure of the L4-L6 -lumbar spine. The L4-5 and L5-6 IVDs each received a single stab injury via a 27G needle and allowed to recover postoperatively for either 2 (n=4) or 8 weeks (n=4). Animals were then  euthanized and the IVD tissues were isolated and pooled from L4-5 and L5-6 (LDP= lumbar disc puncture), or from L2-3 and L3-4 (CON= control) via dissection, with care to remove unwanted peripheral tissues by microdissection. The separated tissue was digested via pronase and collagenase (< 4h total) to obtain IVD cells (51,655 and 60,626 total cell yield from CON and LDP respectively).

first: Milad

last: Rohanifar

first: Sade

last: Clayton

first: Garrett

last: Easson

first: Deepanjali

last: Patil

last: Lee

first: Liufang

last: Jing

first: Marcos

last: Barcellona

first: Julie

last: Speer

first: Jordan

last: Stivers

first: Lori

last: Setton


title: p53 ensures adequate S-adenosylmethionine to prevent R-loop-associated epigenetic and genomic instability

first: panatta

last: emanuele

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


title: HDAC inhibition suppresses telomerase activity in neuroblastoma cells harboring TERT rearrangements [array]

city: Berlin

first: January

last: Weiner

address: Charitéplatz 1

department: CUBI

institute: Berlin Institute of Health, Charité Medical University of Berlin

summary: Genome wide DNA methylation profiling of high-risk TERT-rearranged neuroblastoma cells  treated with solvent or HDAC inhibitor panobinostat. The Illumina HumanMethylation EPIC Beadchip was used to obtain DNA methylation profiles. Samples included 3 solvent treated samples and 3 panobinostat-treated samples after 18 h of treatment.

overall_design: Bisulphite converted DNA from the 6 samples were hybridised to the Illumina HumanMethylation EPIC Beadchip

last: Finkler

first: Konstantin

last: Helmsauer

first: Annika

last: Sprüssel

first: Birte

last: Arlt

first: Annette

last: Kuenkele

first: Johannes

last: Schulte

first: Kerstin

last: Haase

first: Benedikt

last: Obermayer

first: Dieter

last: Beule

first: Matthias

last: Fischer

first: Nils

last: Bluethgen

first: Anton

last: Henssen

first: Marco

last: Lodrini

first: Angelika

last: Eggert

first: Hedwig

last: Deubzer


title: RNA-seq analysis of  HeLa cells overexpressing histone methyltransferase KMT2B

status: Public on Aug 03 2022

city: Shanghai

first: Yu

last: Gan

address: 25/Ln. 2200 Xietu Road

institute: Shanghai Cancer Institute

summary: KMT2B，a major H3K4 tri-methyltransferase, contributes to the development of various cancers. However, its role in cervical cancer  is unclear. We found that  KMT2B is upregulated in cervical cancer tissues, which is significantly associated with poor prognosis.To investigate the underlying mechanism of KMT2B in the progression of cervical cancer, we established stable KMT2B-overexpression HeLa cells.Then we compared the whole genome mRNA expression profile of KMT2B overexpression and control HeLa cells using RNA-seq.

overall_design: HeLa cells endogenous overexpressing KMT2B was established by CRISPR/dCas9 synergistic activation mediator (SAM) system. mRNA profiles of the KMT2B-overexpressing Hela cells and control Hela cells were performed by deep sequencing in quadruplicate, using an Illumina, HiseqXTen platform.

first: Dan

last: Zhao

first: Hui

last: Yuan

first: Hong

last: Tu


title: Next Generation Sequencing Facilitates Quantitative Analysis of E. coli K-12 LE392, P. putida KT2440, and Acinetobacter baylyi ADP1 Transcriptomes under various antidepressant exposure

city: Brisbane

first: Jianhua

last: Guo

address: Research Road, Level 4 Gehrmann Building

department: Australian Centre for Water and Environmental Biotechnology

country: Australia

institute: University of Queensland

summary: The goals of this study are to use Next-generation sequencing (NGS)to detect bacterial mRNA profiles of E. coli K-12 LE392, P. putida KT2440 and Acinetobacter baylyi ADP1 in response to various antidepressant concentrations for 2 h, in triplicate, using Illumina HiSeq 2500.The NGS QC toolkit (version 2.3.3) was used to treat the raw sequence reads to trim the 3’-end residual adaptors and primers, and the ambiguous characters in the reads were removed. Then, the sequence reads consisting of at least 85% bases were progressively trimmed at the 3’-ends until a quality value ≥ 20 were kept. Downstream analyses were performed using the generated clean reads of no shorter than 75 bp. The clean reads of each sample were aligned to the E. coli reference genome (NC_000913), Pseudomonas putida KT2440 genome (NCBI:txid160488) and Acinetobacter baylyi ADP1 genome (NCBI:txid62977) using SeqAlto (version 0.5). Cufflinks (version 2.2.1) was used to calculate the strand-specific coverage for each gene, and to analyze the differential expression in triplicate bacterial cell cultures. The statistical analyses and visualization were conducted using CummeRbund package in R (http://compbio.mit.edu/cummeRbund/). Gene expression was calculated as fragments per kilobase of a gene per million mapped reads (FPKM, a normalized value generated from the frequency of detection and the length of a given gene.

sample_organism:
Pseudomonas putida
Escherichia coli
Acinetobacter baylyi

overall_design: Bacterial mRNA profiles of E. coli K-12 LE392, P.putida KT2440 and A.baylyi ADP1 in response to 0 mg/L (control), 10 mg/L Duloxetine, 1 and 10 mg/L Sertraline, 50 mg/L Escitalopram, 10 mg/L Fluoxetine, 100 mg/L Bupropion and 50 mg/L Agomelatine for 2 h, in triplicate, using Illumina HiSeq 2500

first: Ji

last: Lu

first: Pengbo

last: Ding


title: HIF1A ChIP-seq from primary human uterine smooth muscle cells (hMSMCs) in hypoxia

status: Public on Aug 01 2022

city: Guangzhou

first: Kaiyuan

last: Ji

address: No.9 Jinsui Road, Guangzhou, China

department: Guangzhou Women and Children’s Medical Center

institute: Guangzhou Medical University

summary: Uterine contraction, crucial for successful labor, has been proved to be enchanced by hypoxia, which underlying mechanisms are yet to be elucidated. We found blockade of HIF-1a caused contractility decrease in myometrium and myocytes in hypoxic models. ChIP-seq revealed that HIF-1α directly bound to gemone of 2 typical contraction-associated proteins, the promoter of GJA1 and intron of OXTR. Blockade of GJA1 led to significant decrease of myometrial contractions under hypoxic tissue model, whereas atosiban did not influence the contractility. The conducted research suggests that HIF-1α is required for the augmentation of myometrial contractility via regulating GJA1 under hypoxia.

overall_design: Examination of HIF1a binding sites in primary human uterine smooth muscle cells under hypoxia condition

first: Huishu

last: Liu

first: Bolun

last: Wen


title: BeadChip microarray data of peripheral blood obtained from treatment-näive IBD patients and control individuals

first: Simonas

last: Juzenas

summary: Comperhensive analysis of blood transcriptomes obtained from treatment-näive IBD patients.

overall_design: Total RNA extracted from peripheral blood samples (n = 205) was reverse transcribed and biotin-labeled using the TargetAmp-Nano Labeling Kit for Illumina Expression BeadChip (Epicentre) according to the manufacturer’s protocol. The labeled antisense RNA was hybridized to Human HT-12 v4 BeadChip array (Illumina) following the standard producer’s hybridization protocol. The array imaging was performed on an iScan system (Illumina) according to the standard manufacturer’s protocol.

first: Mauro

last: D'Amato

first: Jonas

last: Halfvarsson

first: Mårten

last: Lindqvist

first: Andre

last: Franke


title: Effect of Tubastatin A (HDAC6 specific inhibitor) on gene expression during differentiation of induced regulatory T cells (iTreg cells)

status: Public on Aug 12 2022

city: Seoul

first: Gap Ryol

last: Lee

address: 35 Baekbeom-ro, Mapo-gu

department: Department of Life Science

institute: Sogang University

summary:
To investigate that Tubastain A, a HDAC6 inhibitor, has effect on early stage of iTreg differentiation, we treated naïve CD4+ T cells with DMSO (vehicle control) or 10 uM Tubastatin A and cultured the cells under Treg-skewing condition for 1 day.
We then performed gene expression profiling analysis using data from RNA-seq of 2 different samples.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for iTreg cells treated with DMSO (vehicle control) or 10 uM Tubastatin A

first: Gap