Genes and environment: novel, functional polymorphism in the human cathepsin L (CTSL1) promoter disrupts a xenobiotic response element (XRE) to alter transcription and blood pressure

Cathepsin L (CTSL1) catalyzes the formation of peptides that influence blood pressure (BP). Naturally occurring genetic variation or targeted ablation of the Ctsl1 locus in mice yield cardiovascular pathology. Here, we searched for genetic variation across the human CTSL1 locus and probed its functional effects, especially in the proximal promoter

“Omics” data integration and functional analyses link Enoyl-CoA hydratase, short chain 1 to drug refractory dilated cardiomyopathy

Abstract Background Large-scale “omics” datasets have not been leveraged and integrated with functional analyses to discover potential drivers of cardiomyopathy. This study addresses the knowledge gap. Methods We coupled RNA sequence (RNA-Seq) variant detection and transcriptome profiling with pathway analysis to model drug refractory dilated cardiomyopathy (drDCM) using the BaseSpace sequencing hub and Ingenuity Pathway Analysis. We used RNA-Seq case-control datasets (n = 6 cases, n = 4 controls), exome sequence familial DCM datasets (n = 3 Italians, n = 5 Italians, n = 5 Chinese), and controls from the HapMap project (n = 5 Caucasians, and n = 5 Asians) for disease modeling and putative mutation discovery. Variant replication datasets: n = 128 cases and n = 15 controls. Source of datasets: NCBI Sequence Read Archive. Statistics: Pairwise differential expression analyses to determine differentially expressed genes and t-tests to calculate p-values. We adjusted for false discovery rates and reported q-values. We used chi-square tests to assess independence among variables, the Fisher’s Exact Tests and overlap p-values for the pathways and p-scores to rank network. Results Data revealed that ECHS1(enoyl-CoA hydratase, short chain 1(log2(foldchange) = 1.63329) hosts a mirtron, MIR3944 expressed in drDCM (FPKM = 5.2857) and not in controls (FPKM = 0). Has-miR3944-3p is a putative target of BAG1 (BCL2 associated athanogene 1(log2(foldchange) = 1.31978) and has-miR3944-5p of ITGAV (integrin subunit alpha V(log2(foldchange) = 1.46107) and RHOD (ras homolog family member D(log2(foldchange) = 1.28851). There is an association between ECHS1:11 V/A(rs10466126) and drDCM (p = 0.02496). The interaction (p = 2.82E-07) between ECHS1:75 T/I(rs1049951) and ECHS1:rs10466126 is associated with drDCM (p < 2.2e-16). ECHS1:rs10466126 and ECHS1:rs1049951 are in linkage disequilibrium (D’ = 1). The interaction (p = 7.84E-08) between ECHS1:rs1049951 and the novel ECHS1:c.41insT variant is associated with drDCM (p < 2.2e-16). The interaction (p = 0.001096) between DBT (Dihydrolipoamide branched chain transacylase E2):384G/S(rs12021720) and ECHS1:rs10466126 is associated with drDCM (p < 2.2e-16). At the mRNA level, there is an association between ECHS1 (log2(foldchange) = 1.63329; q = 0.013927) and DBT (log2(foldchange) = 0.955072; q = 0.0368792) with drDCM. ECHS1 is involved in valine (−log (p = 3.39E00)), isoleucine degradation (p = 0.00457), fatty acid β-oxidation (−log(p) = 2.83E00), and drug metabolism:cytochrome P450 (z-score = 2.07985196) pathways. The mitochondria (−log(p) = 8.73E00), oxidative phosphorylation (−log(p) = 5.35E00) and TCA-cycle II (−log(p) = 2.70E00) are dysfunctional. Conclusions We introduce an integrative data strategy that considers the interplay between the DNA, mRNA, and associated pathways, which represents a possible diagnostic, prognostic, biomarker, and personalized treatment discovery approach in genomically heterogeneous diseases

Single cell transcriptomic analyses reveal diverse and dynamic changes of distinct populations of lung interstitial macrophages in hypoxia-induced pulmonary hypertension.

INTRODUCTION: Hypoxia is a common pathological driver contributing to various forms of pulmonary vascular diseases leading to pulmonary hypertension (PH). Pulmonary interstitial macrophages (IMs) play pivotal roles in immune and vascular dysfunction, leading to inflammation, abnormal remodeling, and fibrosis in PH. However, IMs response to hypoxia and their role in PH progression remain largely unknown. We utilized a murine model of hypoxia-induced PH to investigate the repertoire and functional profiles of IMs in response to acute and prolonged hypoxia, aiming to elucidate their contributions to PH development. METHODS: We conducted single-cell transcriptomic analyses to characterize the repertoire and functional profiles of murine pulmonary IMs following exposure to hypobaric hypoxia for varying durations (0, 1, 3, 7, and 21 days). Hallmark pathways from the mouse Molecular Signatures Database were utilized to characterize the molecular function of the IM subpopulation in response to hypoxia. RESULTS: Our analysis revealed an early acute inflammatory phase during acute hypoxia exposure (Days 1-3), which was resolved by Day 7, followed by a pro-remodeling phase during prolonged hypoxia (Days 7-21). These phases were marked by distinct subpopulations of IMs: MHCIIhiCCR2+EAR2+ cells characterized the acute inflammatory phase, while TLF+VCAM1hi cells dominated the pro-remodeling phase. The acute inflammatory phase exhibited enrichment in interferon-gamma, IL-2, and IL-6 pathways, while the pro-remodeling phase showed dysregulated chemokine production, hemoglobin clearance, and tissue repair profiles, along with activation of distinct complement pathways. DISCUSSION: Our findings demonstrate the existence of distinct populations of pulmonary interstitial macrophages corresponding to acute and prolonged hypoxia exposure, pivotal in regulating the inflammatory and remodeling phases of PH pathogenesis. This understanding offers potential avenues for targeted interventions, tailored to specific populations and distinct phases of the disease. Moreover, further identification of triggers for pro-remodeling IMs holds promise in unveiling novel therapeutic strategies for pulmonary hypertension

Resultant variants and dbNSFP scores after bioinformatic filtering in AD-FDC1.

<p>Shown are the six genes containing exome-detected variants present in all three subjects tested. The Genebank NM number, chromosome and nucleotide position are shown along with the predicted consequence of each variant. Only the TNNT2 variant segregates with the disease phenotype in AD-FDC1. SIFT, Polyphen2, LRT, and MutationTaster scores derived from dbNSFP are presented. SIFT scores less than 0.05 are predicted to be damaging, otherwise they are predicted to be tolerated. Polyphen2_HDIV_scores range from 0 to 1. Scores in the range of 0.957 to 1 are predicted to be possibly damaging and those in the range of 0.453 to 0.956 are predicted to be benign. Polyphen2_HVAR_scores range from 0 to 1. Scores in the range of 0.909 to1 are predicted to be possibly damaging and scores in the range of 0.447 to 0.908 are predicted to be benign. Lower LRT p-values correspond to predictions that are more damaging. A MutationTaster value close to 1 indicates a high ‘security’ of the prediction.</p

Bioinformatics filtering algorithm.

<p>Shown are bioinformatics analysis tools used and individual and mean variant numbers at each step of filtering with resulting decrease in variant numbers at each decision step.</p

Pedigrees of families (A) AD-FDC1 and (B) AD-FDC27.

<p>The family structures of both TNNT2 mutation families are shown. The proband is indicated by an arrow. Males and females are depicted as squares and circles, respectively. Affected individuals are identified by shading. Presence or absence of the Arg173Trp variant confirmed by Sanger sequencing is indicated by plus (‘+’) and minus (‘−’) signs, respectively. Paired numbers beneath individuals represent the numbered haplotypes according to (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0078104#pone-0078104-t004" target="_blank">Table 4</a>); double-numbers represent recombinant haplotypes further detailed in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0078104#pone.0078104.s003" target="_blank">Table S3</a>. Individuals studied by exome sequencing are indicated by present of black underline beneath haplotype numbering.</p

Hap Map project haplotypes and population frequencies of Utah residents with northern and western European ancestry (CEU).

<p>The TNNT2, Arg173Trp variant (Chr1∶201,332,477) was present on separate haplotype, segregating with haplotypes #2 and #1 in families AD-FDC1 and AD-FDC27, respectively.</p

Venn diagram reflecting variant overlap between and among patients.

<p>A Venn diagram depicts the number of variants after bioinformatics filtering to identify nonsynonymous, nonsense and splice site variants that were located in conserved regions (by ANNOVAR), novel (absent in 1000 Genomes and NHLBI Exome Sequencing Project datasets), and were predicted to be damaging by in silico analyses. The numbers and overlap regions in the Venn diagram show the variants unique to or shared among three individuals (IV:4, IV:7, and IV:14) from family AD-FDC1.</p

Visualization of NGS alignment and chromatogram from Sanger sequencing confirming the <i>TNNT2 Arg173Trp</i> variant.

<p>The alignment and Sanger sequencing profiles of the TNNT2 R173W variant are shown. A) C>T variant alignment reads of Arg173Trp variant B) (inset) Chromatogram of C>T variant of Arg173Trp variant from Sanger sequencing; arrow depicts the c.517T C>T (chr1∶201,332,477) position.</p