Genome-wide Transcriptome Studies Research Articles

Higher Maternal Body Mass Index Is Associated With Lower Placental Expression of EPYC: A Genome-Wide Transcriptomic Study.

Elevated body mass index (BMI) in pregnancy is associated with adverse maternal and fetal outcomes. The placental transcriptome may elucidate molecular mechanisms underlying these associations. We examined the association of first-trimester maternal BMI with the placental transcriptome in the Gen3G prospective cohort. We enrolled participants at 5 to 16 weeks of gestation and measured height and weight. We collected placenta samples at delivery. We performed whole-genome RNA sequencing using Illumina HiSeq 4000 and aligned RNA sequences based on the GTEx v8 pipeline. We conducted differential gene expression analysis of over 15 000 genes from 450 placental samples and reported the change in normalized gene expression per 1-unit increase in log2 BMI (kg/m2) as a continuous variable using Limma Voom. We adjusted models for maternal age, fetal sex, gestational age at delivery, gravidity, and surrogate variables accounting for technical variability. We compared participants with BMI of 18.5 to 24.9 mg/kg2 (N = 257) vs those with obesity (BMI ≥30 kg/m2, N = 82) in secondary analyses. Participants' mean ± SD age was 28.2 ± 4.4 years and BMI was 25.4 ± 5.5 kg/m2 in early pregnancy. Higher maternal BMI was associated with lower placental expression of EPYC (slope = -1.94, false discovery rate [FDR]-adjusted P = 7.3 × 10-6 for continuous BMI; log2 fold change = -1.35, FDR-adjusted P = 3.4 × 10-3 for BMI ≥30 vs BMI 18.5-24.9 kg/m2) and with higher placental expression of IGFBP6, CHRDL1, and CXCL13 after adjustment for covariates and accounting for multiple testing (FDR < 0.05). Our genome-wide transcriptomic study revealed novel genes potentially implicated in placental biologic response to higher maternal BMI in early pregnancy.

Role of TRDMT1/DNMT2 in stress adaptation and its influence on transcriptome and proteome dynamics under osmotic stress in Physcomitrium patens.

Early land plants such as the moss Physcomitrium patens lack several morphological traits that offer protection to tracheophytes from environmental stresses. These plants instead have evolved several physiological and biochemical mechanisms that facilitate them to adapt to terrestrial stresses such as drought. We have previously shown that loss-of-function mutants of tRNA (cytosine(38)-C(5))-methyltransferase TRDMT1/DNMT2 in P. patens are highly sensitive to oxidative and osmotic stress. To gain insight into the role of PpTRDMT1/PpDNMT2 in modulating genetic networks under osmotic stress, genome-wide transcriptome and proteome studies were undertaken in wild-type and ppdnmt2 plants. Transcriptome analysis revealed 375 genes to be differentially expressed in the ppdnmt2 under stress compared to the WT. Most of these genes are affiliated with carbohydrate metabolic pathways, photosynthesis, cell wall biogenesis, pathways related to isotropic and polarised cell growth and transcription factors among others. Histochemical staining showed elevated levels of reactive oxygen species in ppdnmt2 while transmission electron microscopy revealed no distinct defects in the ultrastructure of chloroplasts. Immunoprecipitation using PpDNMT2-specific antibody coupled with mass spectrometry revealed core proteins of the glycolytic pathway, antioxidant enzymes, proteins of amino acid biosynthetic pathways and photosynthesis-related proteins among others to co-purify with PpTRDMT1/PpDNMT2 under osmotic stress. Yeast two-hybrid assays, protein deletion and α-galactosidase assays showed the cytosol glycolytic protein glyceraldehyde 3-phosphate dehydrogenase to bind to the catalytic motifs in PpTRDMT1/PpDNMT2. Results presented in this study allow us to better understand genetic networks linking enzymes of energy metabolism, epigenetic processes and RNA pol II-mediated transcription during osmotic stress tolerance in P. patens.

Environmentally realistic concentrations of chlorinated, brominated, and fluorinated persistent organic pollutants induce the unfolded protein response as a shared stress pathway in the liver of Atlantic cod (Gadus morhua).

In the North Sea and North Atlantic coastal areas, fish experience relatively high background levels of persistent organic pollutants. This study aimed to compare the mode of action of environmentally relevant concentrations of mixtures of halogenated compounds in Atlantic cod. Juvenile male cod with mean weight of 840 g were exposed by gavage to dietary mixtures of chlorinated (PCBs, DDT analogs, chlordane, lindane, and toxaphene), brominated (PBDEs), and fluorinated (PFOS) compounds for 4 weeks. One group received a combined mixture of all three compound groups. The results showed that the accumulated levels of chemicals in cod liver after 4weeks of exposure reflected concentrations found in wild fish in this region. Pathway analysis revealed that the treatment effects by each of the three groups of chemicals (chlorinated, brominated, and fluorinated) converged on activation of the unfolded protein response (UPR). Upstream regulator analysis predicted that almost all the key transcription factors (XBP1, ERN1, ATF4, EIF2AK3, and NFE2L2) regulating the UPR were significantly activated. No additive effect was observed in cod co-treated with all three compound groups. In conclusion, the genome-wide transcriptomic study suggests that the UPR pathway is a sensitive common target of halogenated organic environmental pollutants in fish.

Transcriptome- and proteome-wide association studies nominate determinants of kidney function and damage

BackgroundThe pathophysiological causes of kidney disease are not fully understood. Here we show that the integration of genome-wide genetic, transcriptomic, and proteomic association studies can nominate causal determinants of kidney function and damage.ResultsThrough transcriptome-wide association studies (TWAS) in kidney cortex, kidney tubule, liver, and whole blood and proteome-wide association studies (PWAS) in plasma, we assess for effects of 12,893 genes and 1342 proteins on kidney filtration (glomerular filtration rate (GFR) estimated by creatinine; GFR estimated by cystatin C; and blood urea nitrogen) and kidney damage (albuminuria). We find 1561 associations distributed among 260 genomic regions that are supported as putatively causal. We then prioritize 153 of these genomic regions using additional colocalization analyses. Our genome-wide findings are supported by existing knowledge (animal models for MANBA, DACH1, SH3YL1, INHBB), exceed the underlying GWAS signals (28 region-trait combinations without significant GWAS hit), identify independent gene/protein-trait associations within the same genomic region (INHBC, SPRYD4), nominate tissues underlying the associations (tubule expression of NRBP1), and distinguish markers of kidney filtration from those with a role in creatinine and cystatin C metabolism. Furthermore, we follow up on members of the TGF-beta superfamily of proteins and find a prognostic value of INHBC for kidney disease progression even after adjustment for measured glomerular filtration rate (GFR).ConclusionIn summary, this study combines multimodal, genome-wide association studies to generate a catalog of putatively causal target genes and proteins relevant to kidney function and damage which can guide follow-up studies in physiology, basic science, and clinical medicine.

Breast cancer cell mesenchymal transition and metastasis directed by DAP5/eIF3d-mediated selective mRNA translation

Cancer cell plasticity enables cell survival in harsh physiological environments and fate transitions such as the epithelial-to-mesenchymal transition (EMT) that underlies invasion and metastasis. Using genome-wide transcriptomic and translatomic studies, an alternate mechanism of cap-dependent mRNA translation by the DAP5/eIF3d complex is shown to be essential for metastasis, EMT, and tumor directed angiogenesis. DAP5/eIF3d carries out selective translation of mRNAs encoding EMT transcription factors and regulators, cell migration integrins, metalloproteinases, and cell survival and angiogenesis factors. DAP5 is overexpressed in metastatic human breast cancers associated with poor metastasis-free survival. In human and murine breast cancer animal models, DAP5 is not required for primary tumor growth but is essential for EMT, cell migration, invasion, metastasis, angiogenesis, and resistance to anoikis. Thus, cancer cell mRNA translation involves two cap-dependent mRNA translation mechanisms, eIF4E/mTORC1 and DAP5/eIF3d. These findings highlight a surprising level of plasticity in mRNA translation during cancer progression and metastasis.

A deep transcriptome meta-analysis reveals sex differences in multiple sclerosis

BackgroundMultiple sclerosis (MS), a chronic auto-immune, inflammatory, and degenerative disease of the central nervous system, affects both males and females; however, females suffer from a higher risk of developing MS (2–3:1 ratio relative to males). The precise sex-based factors influencing risk of MS are currently unknown. Here, we explore the role of sex in MS to identify molecular mechanisms underlying observed MS sex differences that may guide novel therapeutic approaches tailored for males or females. MethodsWe performed a rigorous and systematic review of genome-wide transcriptome studies of MS that included patient sex data in the Gene Expression Omnibus and ArrayExpress databases following PRISMA statement guidelines. For each selected study, we analyzed differential gene expression to explore the impact of the disease in females (IDF), in males (IDM) and our main goal: the sex differential impact of the disease (SDID). Then, for each scenario (IDF, IDM and SDID) we performed 2 meta-analyses in the main tissues involved in the disease (brain and blood). Finally, we performed a gene set analysis in brain tissue, in which a higher number of genes were dysregulated, to characterize sex differences in biological pathways. ResultsAfter screening 122 publications, the systematic review provided a selection of 9 studies (5 in blood and 4 in brain tissue) with a total of 474 samples (189 females with MS and 109 control females; 82 males with MS and 94 control males). Blood and brain tissue meta-analyses identified, respectively, 1 (KIR2DL3) and 13 (ARL17B, CECR7, CEP78, IFFO2, LOC401127, NUDT18, RNF10, SLC17A5, STMP1, TRAF3IP2-AS1, UBXN2B, ZNF117, ZNF488) MS-associated genes that differed between males and females (SDID comparison). Functional analyses in the brain revealed different altered immune patterns in females and males (IDF and IDM comparisons). The pro-inflammatory environment and innate immune responses related to myeloid lineage appear to be more affected in females, while adaptive responses associated with the lymphocyte lineage in males. Additionally, females with MS displayed alterations in mitochondrial respiratory chain complexes, purine, and glutamate metabolism, while MS males displayed alterations in stress response to metal ion, amine, and amino acid transport. ConclusionWe found transcriptomic and functional differences between MS males and MS females (especially in the immune system), which may support the development of new sex-based research of this disease. Our study highlights the importance of understanding the role of biological sex in MS to guide a more personalized medicine.

Abstract 3704: The essential role of an alternate mRNA translation initiation in the regulation of breast cancer cell metabolism

Abstract Translational control and metabolic reprogramming are hallmarks of advanced cancers. Important genes involved in cancer progression express mRNAs that are selectively translated, including regulators of cancer cell metabolism. Cancer cells acquire an altered metabolism, switching from oxidative phosphorylation (OXPHOS) to a glycolytic phenotype (Warburg effect), to increase reliance on alternate metabolic pathways to support growth, proliferation, and metastasis. Triple-negative breast cancer (TNBC), one of the most aggressive and highly metastatic subtypes with the poorest outcome, is characterized by elevated glycolysis and low OXPHOS. Still, the exact mechanism for this metabolic switch is largely unknown. Using a TNBC cell model it has been shown an alternate mechanism of cap-dependent but mTORC1/eIF4Eindependent mRNA translation by the eIF4G homolog, DAP5, which directly binds the cap-binding protein eIF3d. Our research indicates that this alternate translation initiation is essential in regulating several mRNAs involved in breast cancer metabolism. DAVID analysis from genome-wide transcriptomic and translatomic studies, metabolomic assays including Glycolysis Assay [Extracellular Acidification], Oxidative Phosphorylation, and Mitotracker Assay, as well as RT-PCR and western blot analysis was performed from well-characterized triple-negative aggressive breast cancer cell lines,4T1 and MDA-231, Non-silencing (Nsi) control and silencing DAP5 (shDAP5) samples. Several mRNAs related to glucose metabolism decreased after silencing DAP5 and on the opposite, critical mRNAs associated with OXPHOS increased, independent of their steady-state mRNA abundance. Silencing DAP5 increased Oxidative Phosphorylation activity and decrease the glycolytic rate in TNBC breast cancer cells. Proteins correlated with aberrant glycolytic metabolism including the E-cadherin transcriptional repressors Snail (SNAI1) were reduced after silencing of DAP5. Our data indicates that the alternate mRNA translation initiation mechanism by DAP5 of specific mRNAs correlated to glycolysis and OXPHOS could play an essential role in the mechanism by which TNBC breast cancer cells orchestrate a metabolic switch required for malignant progression and metastasis. Citation Format: Hanna Rosenstock, Erna Mitaishvili, Columba de la Parra. The essential role of an alternate mRNA translation initiation in the regulation of breast cancer cell metabolism. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3704.

Abstract P240: Higher Body Mass Index is Associated With Lower Placental Expression of EPYC: Results From a Genome-Wide Transcriptomic Study in the Gen3G Cohort

Background: Elevated maternal body mass index (BMI) in pregnancy is associated with adverse maternal and fetal outcomes. Identification of alterations in the placental transcriptome may elucidate molecular markers of the biological pathways underlying these associations. Methods: We investigated the association of first trimester maternal BMI with the placental transcriptome in Gen3G, a large population-based pregnancy cohort. We completed RNA sequencing of 449 placental samples and performed differential gene expression analysis of over 15,000 genes (Limma Voom). We used maternal BMI as a continuous variable, with estimates reported as change in normalized gene expression per 1-unit increase in BMI (kg/m2), as well as a categorical variable (participants with normal weight (N=270) vs. obesity (N=82)), with RNA expression estimates reported as log2 fold changes. We adjusted models for maternal age, fetal sex, gestational age and 6 surrogate variables inferred from the expression data to adjust for batch effects and other unmeasured sources of variability. We accounted for multiple testing using False Discovery Rate (FDR). Sensitivity analyses were performed by removing cases of gestational diabetes and hypertensive disorders in pregnancy. Results: At enrollment in early pregnancy, participants had a mean ± SD age of 28.2 ± 4.4 years and measured BMI of 25.4 ± 5.5 kg/m 2 ; 57% of participants were categorized in normal weight, 22% in overweight and 18% in obesity BMI categories. Maternal BMI was significantly associated with decreased expression of EPYC , a gene that regulates fibrillogenesis (slope= -0.14, FDR-adjusted P=1.68x10 -5 for continuous BMI; log2 fold change= -1.71, FDR-adjusted P=1.06x10 -2 for obesity vs. normal weight). Both sensitivity analyses with the removal of gestational diabetes and hypertensive disorders revealed similar associations for EPYC . Conclusions: We identified an association between elevated maternal BMI and downregulation of EPYC expression in the placenta. Additional studies are required to examine the role of placental EPYC expression in maternal and fetal health.

Genome-wide association and transcriptome studies reveal the segregation mechanism of mantle markings in three-way cross hybrid abalone

Trait separation which often appears in shellfish progeny, has been commercially used in shellfish breeding projects. A three-way cross hybrid abalone was produced with heterosis in growth performance and thermal tolerance, and with segregation in mantle marking. However, the inheritance mechanism of mantle marking is unclear. In this study, mantle marking was demonstrated to be a qualitative trait, following simple Mendelian inheritance, through inheritance pattern analysis of the presence of black markings on the mantle in five families of three-way cross of hybrid abalone. Thermal tolerance of abalone was assessed by the Arrhenius breakpoint temperature (ABT) of cardiac performance and a correlation between mantle marking and thermal tolerance of abalone was verified. A genome-wide association study (GWAS) for mantle marking was conducted using 603,067 high-quality single nucleotide polymorphisms (SNPs) derived from 57 individuals with mantle markings (Y group) and 58 individuals without mantle markings (N group). A total of 493 SNPs that were significantly (p &amp;lt; 2.32E–07) associated with mantle markings were mainly distributed on chromosome 15. In the potential significantly associated region, 80 genes, including pigmentation-related genes PTPRT, PTPRC, PNCA, and CALM4 were annotated. Transcriptome analysis of the two groups showed that the expression levels of these genes (PTPRT, DDT-b, ATOX1, SLC6A3, and GSTO1) were significantly different, and they may play important roles in the formation of mantle markings in the three-way cross hybrid abalone. Overall, our data provide valuable information for deciphering the phenotype differences of mantle marking in three-way cross hybrid abalone and help in the molecular marker-assisted breeding in abalone.

Co-option of the SHOOT MERISTEMLESS network regulates protocorm-like body development in Phalaenopsis aphrodite.

The protocorm is a structure that is formed upon germination of an orchid seed. It lacks cotyledons and is ovoid in shape. The protocorm-like body (PLB), on the other hand, is a protocorm-like organ induced from somatic tissues. PLBs have been widely used for orchid micropropagation. Because of its unique structure and its application in the orchid industry, PLB development has drawn considerable interest from orchid and developmental biologists. Our previous genome-wide comparative transcriptome study demonstrated that protocorms and PLBs share similar molecular signatures and suggested that SHOOT MERISTEMLESS (STM)-dependent organogenesis is important for PLB development. Here, we show that overexpression of Phalaenopsis aphrodite STM (PaSTM) greatly enhances PLB regeneration from vegetative tissue-based explants of Phalaenopsis orchids, confirming its regulatory role in PLB development. Expression of PaSTM restored shoot meristem function of the Arabidopsis (Arabidopsis thaliana) stm-2 mutant. Moreover, we identified class S11 MYB transcription factors (TFs) as targets downstream of PaSTM. A cis-acting element, TTGACT, identified in the promoters of S11 MYB TFs was found to be important for PaSTM binding and activation. Overexpression of PaSTM or its downstream targets, PaMYB13, PaMYB14, and PaMYB17, enhanced de novo shoot regeneration in Arabidopsis, indicating the active role of the PaSTM-S11 PaMYB module in organogenesis. In summary, our data demonstrate that PaSTM is important for PLB development. The STM-S11 MYB regulatory module is evolutionarily conserved and may regulate shoot or shoot-related organ development in plants.

Co-expression analysis of placental genes in the search for key signaling pathways and biomarkers of the great obstetrical syndromes

Objective. To study the molecular mechanisms responsible for the development of diseases grouped within the great obstetrical syndromes (GOS) at the level of the transcriptome of human maternal placenta.Material and Methods. We gathered the results of genome-wide transcriptome studies of the human placental tissue using Gene Expression Omnibus (GEO) data repository for the following phenotypes: physiological pregnancy, preeclampsia (PE), premature birth, and intrauterine growth restriction (IUGR). Eleven data sets were selected and supplemented with our experimental data; a total of 481 samples of human placental tissue were included in the integrative analysis. Bioinformatic data processing and statistical analyses were performed in the R v3.6.1 software environment using the Bioconductor packages. The pooled dataset was used to search for common molecular targets for GOS via weighted gene co-expression network analysis (WGCNA). The functional annotation of genes and the resulting clusters was carried out with the DAVID database; protein-protein interaction network was built using the STRING software; and the hub genes for the network were identified using the MCC analysis with plugin cytoHubba in Cytoscape software 3.7.2.Results. We obtained a table of expression levels for 15,167 genes in 246 samples. Hierarchical clustering of this network allowed to find 55 modules of co-expressed genes in the group with PE, 109 modules in the group with PB, 75 modules in patients with IUGR, and 56 modules in the control group. The preservation analysis of co-expressed modules for the studied phenotypes suggested the presence of a common cluster comprising eight genes specific only for patients with PE and IUGR, as well as the module of 23 co-expressed genes typical only for patients with PB and IUGR. Protein-protein interaction network was built for these gene sets, and the SOD1, TXNRD1, and UBB genes were the central nodes in the network. Based on network topology evaluation with cytoHubba, six hub genes (rank ˂ 5) were identified as follows: SOD1, TKT, TXNRD1, GCLM, GOT1, and ACO1.Conclusion. The obtained results allowed to identify promising genetic markers for preeclampsia, intrauterine growth restriction, and miscarriage. Moreover, the study also made it possible to identify the most important overlapping molecular mechanisms of these diseases occurring in the placental tissue.

The Role of Long Non-Coding RNAs in Endometriosis.

Endometriosis is a chronic gynecological disorder affecting the quality of life and fertility of many women around the world. Heterogeneous and non-specific symptoms may lead to a delay in diagnosis, with treatment options limited to surgery and hormonal therapy. Hence, there is a need to better understand the pathogenesis of the disease to improve diagnosis and treatment. Long non-coding RNAs (lncRNAs) have been increasingly shown to be involved in gene regulation but remain relatively under investigated in endometriosis. Mutational and transcriptomic studies have implicated lncRNAs in the pathogenesis of endometriosis. Single-nucleotide polymorphisms (SNPs) in lncRNAs or their regulatory regions have been associated with endometriosis. Genome-wide transcriptomic studies have identified lncRNAs that show deregulated expression in endometriosis, some of which have been subjected to further experiments, which support a role in endometriosis. Mechanistic studies indicate that lncRNAs may regulate genes involved in endometriosis by acting as a molecular sponge for miRNAs, by directly targeting regulatory elements via interactions with chromatin or transcription factors or by affecting signaling pathways. Future studies should concentrate on determining the role of uncharacterized lncRNAs revealed by endometriosis transcriptome studies and the relevance of lncRNAs implicated in the disease by in vitro and animal model studies.

Mapping epigenetic divergence in the massive radiation of Lake Malawi cichlid fishes

Epigenetic variation modulates gene expression and can be heritable. However, knowledge of the contribution of epigenetic divergence to adaptive diversification in nature remains limited. The massive evolutionary radiation of Lake Malawi cichlid fishes displaying extensive phenotypic diversity despite extremely low sequence divergence is an excellent system to study the epigenomic contribution to adaptation. Here, we present a comparative genome-wide methylome and transcriptome study, focussing on liver and muscle tissues in phenotypically divergent cichlid species. In both tissues we find substantial methylome divergence among species. Differentially methylated regions (DMR), enriched in evolutionary young transposons, are associated with transcription changes of ecologically-relevant genes related to energy expenditure and lipid metabolism, pointing to a link between dietary ecology and methylome divergence. Unexpectedly, half of all species-specific DMRs are shared across tissues and are enriched in developmental genes, likely reflecting distinct epigenetic developmental programmes. Our study reveals substantial methylome divergence in closely-related cichlid fishes and represents a resource to study the role of epigenetics in species diversification.

Activation of Fibroblasts in Skin Cancer

Fibroblasts have emerged as a dominant component of the tumor microenvironment, but despite the surging interest in the activation of fibroblasts and their role in cancer, they remain an elusive and complex cell type. In this perspective, we discuss the phenotypic plasticity of cancer-associated fibroblasts (CAFs) in melanoma and nonmelanoma skin cancer identified by genome-wide transcriptomic studies and focus on the molecular pathways underlying their activation. These studies reveal distinct fibroblast activation profiles depending on tumor type and stage. A better understanding of skin CAF heterogeneity in origin and function will guide novel therapeutic approaches targeting this cell type in clinical cancer care.

Overexpression of Histone Demethylase KDM6B in the Hematopoietic Compartment Promotes the Effect of Innate Immune Activation and Results in Myelodysplastic Syndrome (MDS)-like Phenotype in Mice

The histone demethylase KDM6B regulates transcriptional activity of genes involved in innate immune signals and controls differentiation of hematopoietic cells. Our laboratory previously reported that KDM6B is overexpressed in bone marrow (BM) hematopoietic stem and progenitor cells (HSPCs) of over 50% MDS pts (Leukemia 2013). Pharmacologic targeting of KDM6B has shown pre-clinical activity in several neoplasms such as glioma and ALL. It still remains to be elucidated whether KDM6B overexpression directly alters hematopoiesis, in order to test the hypothesis that overexpression of KDM6B contributes to aberrant epigenetic and innate immune signal in MDS and thus forms an important pathological mechanism and potential therapeutic target.Here we report an animal model of KDM6B overexpression in mouse hematopoietic compartment (Vav-KDM6B). In 20-wk old mice, KDM6B overexpression led to mild phenotype including reduced white cell count (WBC) in peripheral blood (PB) (15% p<0.05 N=47) and decreased LSK in BM (20% p<0.05 N=12). When exposed to chronic low dose Toll-like receptor ligand lipopolysaccharide (LPS) (6mg every other day, i.p. 6 wks), phenotype severity was more profound in Vav-KDM6B mice than in wildtype (WT) mice. First, hematopoietic cell numbers in both PB and BM were significantly reduced: LPS led to a 40% decrease of PB WBC (p<0.01 N=17) and a 38% decrease of BM cell (p<0.01 N=13 Fig 1A) in KDM6B mice, compared to 20% and 28% reduction respectively in WT mice treated by LPS. Second, both number and repopulating function of BM long term (LT) HSC were severely compromised: LPS led to decrease of BM LT-HSC by 52% (p<0.01 N=13) in KDM6B mice compared to a 35% decrease in WT mice. Functionally, colonies formed in methylcellulose by LSKs of LPS treated KDM6B mice decreased by 75% (p<0.01 N=13) at 2nd plating, whereas LPS led to a reduction of 60% for WT-LSK colony formation at 2nd plating. Similar effect was also observed when BM repopulating capacity was evaluated by chimerism (CD45.2:CD45.1) in competitive transplantation assay (p<0.05 N=11). Third, increase of megakaryocytic dysplasia was documented in BM of LPS treated KDM6B mice compared to WT counterparts (Fig 1B).To identify KDM6B overexpression mediated molecular signals under innate immune stimuli, we performed genome-wide transcriptome study using BM LSK of treated mice. Results reveal that over 130 genes were more profoundly activated by LPS in LSK of Vav-KDM6B than in WT LSK. Gene set enrichment assays (GSEA) indicated that major molecular pathways related to these genes include MDS-relevant signals, such as inflammatory and complement innate immune signals, MAPK pathway, as well as activated gene signature of RPS14 knockout in human BM CD34+ cell. We also performed RNA-Seq analysis in BM CD34+ HSPCs of MDS (N=26) and healthy controls (N=10). Results confirmed that a significant proportion of these genes (N>20) are overexpressed in MDS patients. Furthermore, we demonstrated in mouse BM cells that KDM6B overexpression mediates histone H3K4 and H3K27 methylation at promoters of several key innate immune genes towards transcription activation.We next evaluated effect of pharmacological KDM6B targeting using a highly specific KDM6B inhibitor (GSK-J4). At relatively low-dose (0.5mM), GSK-J4 could increase in vitro colony formation of LSKs of LPS treated KDM6B mice by 80% (p<0.05) at 2nd plating. Injection of GSK-J4 (160mg once a week, 5 weeks) resulted in 50% increase of WBM cells in LPS treated Vav-KDM6B mice (p<0.05). These results suggest that GSK-J4 can improve compromised hematopoiesis of Vav-KDM6B mice. We next treated primary BM HSPCs isolated from patients with low risk MDS (N=11) with GSK-J4. Results indicated that at 0.5mM, GSK-J4 could improve colony formation of MDS CD34+ cells (p=0.05). Flow cytometry analysis using cells from colonies indicated that GSK-J4 led to increased myeloid mark CD11b (p=0.08) and reduced apoptotic mark annexin-V (p<0.05). The same concentration of GSK-J4 did not affect colony formation of normal BM CD34+ HSPCs isolated from healthy donors (N=4).Taken together, our results indicate that overexpression of KDM6B mediates the hyperactivation of innate immune signals and has a functional impact in BM HSPCs, supporting role for KDM6B overexpression in MDS pathogenesis. Furthermore, our study also indicates that pharmacological inhibition of KDM6B may have therapeutic potential in MDS. [Display omitted] DisclosuresKhoury:Kiromics: Research Funding; Angle: Research Funding; Stemline Therapeutics: Research Funding; Pfizer: Research Funding.

Genome-wide association and transcriptome studies identify candidate genes and pathways for feed conversion ratio in pigs

BackgroundThe feed conversion ratio (FCR) is an important productive trait that greatly affects profits in the pig industry. Elucidating the genetic mechanisms underpinning FCR may promote more efficient improvement of FCR through artificial selection. In this study, we integrated a genome-wide association study (GWAS) with transcriptome analyses of different tissues in Yorkshire pigs (YY) with the aim of identifying key genes and signalling pathways associated with FCR.ResultsA total of 61 significant single nucleotide polymorphisms (SNPs) were detected by GWAS in YY. All of these SNPs were located on porcine chromosome (SSC) 5, and the covered region was considered a quantitative trait locus (QTL) region for FCR. Some genes distributed around these significant SNPs were considered as candidates for regulating FCR, including TPH2, FAR2, IRAK3, YARS2, GRIP1, FRS2, CNOT2 and TRHDE. According to transcriptome analyses in the hypothalamus, TPH2 exhibits the potential to regulate intestinal motility through serotonergic synapse and oxytocin signalling pathways. In addition, GRIP1 may be involved in glutamatergic and GABAergic signalling pathways, which regulate FCR by affecting appetite in pigs. Moreover, GRIP1, FRS2, CNOT2, and TRHDE may regulate metabolism in various tissues through a thyroid hormone signalling pathway.ConclusionsBased on the results from GWAS and transcriptome analyses, the TPH2, GRIP1, FRS2, TRHDE, and CNOT2 genes were considered candidate genes for regulating FCR in Yorkshire pigs. These findings improve the understanding of the genetic mechanisms of FCR and may help optimize the design of breeding schemes.

Synergy of oxytocin and citalopram in modulating Itgb3/Chl1 interplay: Relevance to sensitivity to SSRI therapy

Intranasal treatment with oxytocin showed beneficial effects in post-traumatic stress disorder and autism spectrum disorders; however, it was not investigated as much in depression. Keeping in mind the favorable effects of oxytocin on animal models of anxiety and depression, we postulated that synergy between prescribed first choice drugs, selective serotonin reuptake inhibitors (SSRIs) and oxytocin could improve the treatment outcome compared with SSRI monotherapy. Our previous in vitro genome-wide transcriptomic study on human lymphoblastoid cell lines exposed to paroxetine resulted in increase of integrin β3 (ITGB3) gene expression, and further, ITGB3/CHL1 expression ratio was hypothesized to influence the sensitivity to SSRIs. The aim of this report was to explore molecular mechanisms behind the antidepressant-like oxytocin effect, alone and in synergy with citalopram, on behavioral and molecular level in corticosterone treated rats, a paradigm used to model anxiety and depression in animals. Oxytocin treatment (1) ameliorated corticosterone-induced reduction of neurogenesis and number of parvalbumin-positive interneurons in the hippocampal CA1 region, (2) enhanced anxiolytic- and antidepressant-like effects of citalopram in the open field test, and (3) the SSRI/oxytocin synergy persisted in reversing the reduction of the Itgb3 gene expression and increased Itgb3/Chl1 ratio in the prefrontal cortices. These results support the existence of synergy between citalopram and oxytocin in reversing the molecular and behavioral changes induced by corticosterone treatment and point to possible molecular mechanisms behind antidepressant-like effect of oxytocin.

Genome-wide transcriptome study in skin biopsies reveals an association of E2F4 with cadasil and cognitive impairment

CADASIL is a small vessel disease caused by mutations in NOTCH3 that lead to an odd number of cysteines in the EGF-like repeat domain, causing protein misfolding and aggregation. The main symptoms are migraine, psychiatric disturbances, recurrent strokes and dementia, being executive function characteristically impaired. The molecular pathways altered by this receptor aggregation need to be studied further. A genome-wide transcriptome study (four cases paired with three healthy siblings) was carried out, in addition to a qRT-PCR for validation purposes (ten new cases and eight new controls). To study the expression profile by cell type of the significant mRNAs found, we performed an in situ hybridization (ISH) (nine cases and eight controls) and a research in the Single-nuclei Brain RNA-seq expression browser (SNBREB). Pathway analysis enrichment was carried out with Gene Ontology and Reactome. Neuropsychological tests were performed in five of the qRT-PCR cases. The two most significant differentially expressed mRNAs (BANP, p-value = 7.23 × 10–4 and PDCD6IP, p-value = 8.36 × 10–4) were selected for the validation study by qRT-PCR. Additionally, we selected two more mRNAs (CAMK2G, p-value = 4.52 × 10–3 and E2F4, p-value = 4.77 × 10–3) due to their association with ischemic neuronal death. E2F4 showed differential expression in the genome-wide transcriptome study and in the qRT-PCR (p = 1.23 × 10–3), and it was upregulated in CADASIL cases. Furthermore, higher E2F4 expression was associated with worse executive function (p = 2.04 × 10–2) and attention and information processing speed (IPS) (p = 8.73 × 10–2). In situ hibridization showed E2F4 expression in endothelial and vascular smooth vessel cells. In silico studies indicated that E2F4 is also expressed in brain endothelial cells. Among the most significant pathways analyzed, there was an enrichment of vascular development, cell adhesion and vesicular machinery terms and autophagy process. E2F4 is more highly expressed in the skin biopsy of CADASIL patients compared to controls, and its expression is present in endothelial cells and VSMCs. Further studies are needed to understand whether E2F4 could be useful as a biomarker, to monitor the disease or be used as a therapeutic target.

Posttraumatic stress disorder: from gene discovery to disease biology.

Posttraumatic stress disorder (PTSD) is a complex mental disorder afflicting approximately 7% of the population. The diverse number of traumatic events and the wide array of symptom combinations leading to PTSD diagnosis contribute substantial heterogeneity to studies of the disorder. Genomic and complimentary-omic investigations have rapidly increased our understanding of the heritable risk for PTSD. In this review, we emphasize the contributions of genome-wide association, epigenome-wide association, transcriptomic, and neuroimaging studies to our understanding of PTSD etiology. We also discuss the shared risk between PTSD and other complex traits derived from studies of causal inference, co-expression, and brain morphological similarities. The investigations completed so far converge on stark contrasts in PTSD risk between sexes, partially attributed to sex-specific prevalence of traumatic experiences with high conditional risk of PTSD. To further understand PTSD biology, future studies should focus on detecting risk for PTSD while accounting for substantial cohort-level heterogeneity (e.g. civilian v. combat-exposed PTSD cases or PTSD risk among cases exposed to specific traumas), expanding ancestral diversity among study cohorts, and remaining cognizant of how these data influence social stigma associated with certain traumatic events among underrepresented minorities and/or high-risk populations.

Genome-wide transcriptome study using deep RNA sequencing for myocardial infarction and coronary artery calcification

BackgroundCoronary artery calcification (CAC) is a noninvasive measure of coronary atherosclerosis, the proximal pathophysiology underlying most cases of myocardial infarction (MI). We sought to identify expression signatures of early MI and subclinical atherosclerosis in the Framingham Heart Study (FHS). In this study, we conducted paired-end RNA sequencing on whole blood collected from 198 FHS participants (55 with a history of early MI, 72 with high CAC without prior MI, and 71 controls free of elevated CAC levels or history of MI). We applied DESeq2 to identify coding-genes and long intergenic noncoding RNAs (lincRNAs) differentially expressed in MI and high CAC, respectively, compared with the control.ResultsOn average, 150 million paired-end reads were obtained for each sample. At the false discovery rate (FDR) < 0.1, we found 68 coding genes and 2 lincRNAs that were differentially expressed in early MI versus controls. Among them, 60 coding genes were detectable and thus tested in an independent RNA-Seq data of 807 individuals from the Rotterdam Study, and 8 genes were supported by p value and direction of the effect. Immune response, lipid metabolic process, and interferon regulatory factor were enriched in these 68 genes. By contrast, only 3 coding genes and 1 lincRNA were differentially expressed in high CAC versus controls. APOD, encoding a component of high-density lipoprotein, was significantly downregulated in both early MI (FDR = 0.007) and high CAC (FDR = 0.01) compared with controls.ConclusionsWe identified transcriptomic signatures of early MI that include differentially expressed protein-coding genes and lincRNAs, suggesting important roles for protein-coding genes and lincRNAs in the pathogenesis of MI.