Genome-wide co-expression analysis in multiple tissues

Expression quantitative trait loci (eQTLs) represent genetic control points of gene expression, and can be categorized as cis- and trans-acting, reflecting local and distant regulation of gene expression respectively. Although there is evidence of co-regulation within clusters of trans-eQTLs, the extent of co-expression patterns and their relationship with the genotypes at eQTLs are not fully understood. We have mapped thousands of cis- and trans-eQTLs in four tissues (fat, kidney, adrenal and left ventricle) in a large panel of rat recombinant inbred (RI) strains. Here we investigate the genome-wide correlation structure in expression levels of eQTL transcripts and underlying genotypes to elucidate the nature of co-regulation within cis- and trans-eQTL datasets. Across the four tissues, we consistently found statistically significant correlations of cis-regulated gene expression to be rare (<0.9% of all pairs tested). Most (>80%) of the observed significant correlations of cis-regulated gene expression are explained by correlation of the underlying genotypes. In comparison, co-expression of trans-regulated gene expression is more common, with significant correlation ranging from 2.9%-14.9% of all pairs of trans-eQTL transcripts. We observed a total of 81 trans-eQTL clusters (hot-spots), defined as consisting of > or =10 eQTLs linked to a common region, with very high levels of correlation between trans-regulated transcripts (77.2-90.2%). Moreover, functional analysis of large trans-eQTL clusters (> or =30 eQTLs) revealed significant functional enrichment among genes comprising 80% of the large clusters. The results of this genome-wide co-expression study show the effects of the eQTL genotypes on the observed patterns of correlation, and suggest that functional relatedness between genes underlying trans-eQTLs is reflected in the degree of co-expression observed in trans-eQTL clusters. Our results demonstrate the power of an integrative, systematic approach to the analysis of a large gene expression dataset to uncover underlying structure, and inform future eQTL studies

Legal Acts Aimed at the Termination of an Employment Relationship

This diploma thesis is dealing with legal acts aimed at the termination of an employment relationship. Its aim is to characterize each individual legal act aimed at the termination of an employment relationship and analysing the legislation behind them, but also to elaborate on individual conditions for their valid realization not only as amended by law, but also while using the wide judicature, which this area provides. The thesis consists of an introduction, ten chapters, which are, except from the seventh and eight chapter, further internally divided, and the conclusion. In the introduction, the choice of the given theme is explained as well as the desired achievement of this diploma thesis. It also contains a brief introduction of the thesis' structure. The first chapter consists of a description of the term labour law, its functions and position in the legal system, especially regarding its relation to civil law. Furthermore, the definitions of some fundamental terms related to labour law such as an employee, an employer and dependent work can be found in this chapter as well. The second chapter shortly describes employment relations, an employment relationship, its termination in general, but also regarding other ways of terminating an employment relationship not dependant on human will. In...Tato diplomová práce se zabývá právními jednáními směřujícími ke skončení pracovního poměru. Jejím cílem je charakterizovat jednotlivá právní jednání směřující ke skončení pracovního poměru, analyzovat jejich právní úpravu a rozvést jednotlivé podmínky pro jejich platné uskutečnění, a to nejen za užití znění zákona, ale také bohaté judikatury, kterou tato oblast poskytuje. Práce se skládá z úvodu, deseti kapitol, které jsou, vyjma sedmé a osmé kapitoly, dále vnitřně členěny, a závěru. V úvodu se nachází odůvodnění volby daného tématu, vymezení cíle této diplomové práce a dále je ve zkratce představena struktura práce. V první kapitole je vymezen pojem pracovní právo a jeho funkce. Dále je rozebráno postavení pracovního práva v systému práva, zejména s ohledem na vztah k občanskému právu. V této kapitole jsou také definovány základní pojmy týkající se pracovního poměru jako jsou zaměstnanec, zaměstnavatel a závislá práce. Druhá kapitola ve zkratce popisuje pracovněprávní vztahy, pracovní poměr a jeho skončení obecně, ale i s ohledem na způsoby skončení pracovního poměru, které nejsou závislé na lidské vůli. Ve třetí kapitole je krátce věnována pozornost právnímu jednání, jeho pro pracovní právo nejdůležitějšímu dělení a také jeho náležitostem. Těžištěm této práce jsou čtvrtá až sedmá kapitola, ve...Department of Labor Law and Social Security LawKatedra pracovního práva a práva sociálního zabezpečeníFaculty of LawPrávnická fakult

Health Quality Control of Laboratory Anjmals 0f the Czechoslovak Academy of Sciences (CSAV)

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Cap analysis of gene expression reveals alternative promoter usage in a rat model of hypertension

The role of alternative promoter usage in tissue-specific gene expression has been well established, however, its role in complex diseases is poorly understood. We performed cap analysis of gene expression (CAGE) sequencing from the left ventricle (LV) of a rat model of hypertension, the spontaneously hypertensive rat (SHR), and a normotensive strain, Brown Norway (BN) to understand the role of alternative promoter usage in complex disease. We identified 26,560 CAGE-defined transcription start sites (TSS) in the rat LV, including 1,970 novel cardiac TSSs. We identified 28 genes with alternative promoter usage between SHR and BN, which could lead to protein isoforms differing at the amino terminus between two strains and 475 promoter switching events altering the length of the 5’ UTR. We found that the shift in Insr promoter usage was significantly associated with insulin levels and blood pressure within a panel of HXB/BXH recombinant inbred rat strains, suggesting that hyperinsulinemia due to insulin resistance might lead to hypertension in SHR. Our study provides a preliminary evidence of alternative promoter usage in complex diseases

Transgenic overexpression of glutathione S-transferase μ-type 1 reduces hypertension and oxidative stress in the stroke-prone spontaneously hypertensive rat

Background: Combined congenic breeding and microarray gene expression profiling previously identified glutathione S-transferase m-type 1 (Gstm1) as a positional and functional candidate gene for blood pressure (BP) regulation in the stroke-prone spontaneously hypertensive (SHRSP) rat. Renal Gstm1 expression in SHRSP rats is significantly reduced when compared with normotensive Wistar Kyoto (WKY) rats. As Gstm1 plays an important role in the secondary defence against oxidative stress, significantly lower expression levels may be functionally relevant in the development of hypertension. The aim of this study was to investigate the role of Gstm1 in BP regulation and oxidative stress by transgenic overexpression of the Gstm1 gene. Method: Two independent Gstm1 transgenic SHRSP lines were generated by microinjecting SHRSP embryos with a linear construct controlled by the EF-1a promoter encoding WKY Gstm1 cDNA [SHRSP-Tg(Gstm1)1WKY and SHRSPTg(Gstm1)2WKY]. Results: Transgenic rats exhibit significantly reduced BP and pulse pressure when compared with SHRSP [systolic: SHRSP 205.2 3.7 mmHg vs. SHRSP-Tg(Gstm1)1WKY 175.5 1.6 mmHg and SHRSP-Tg(Gstm1)2WKY 172 3.2 mmHg, P< 0.001; pulse pressure: SHRSP 58.4 0.73 mmHg vs. SHRSP-Tg(Gstm1)1WKY 52.7 0.19 mmHg and SHRSP-Tg(Gstm1)2WKY 40.75 0.53 mmHg, P< 0.001]. Total renal and aortic Gstm1 expression in transgenic animals was significantly increased compared with SHRSP [renal relative quantification (RQ): SHRSP-Tg(Gstm1)1WKY 1.95 vs. SHRSP 1.0, P< 0.01; aorta RQ: SHRSP-Tg(Gstm1)1WKY 2.8 vs. SHRSP 1.0, P< 0.05]. Renal lipid peroxidation (malondialdehyde: protein) and oxidized : reduced glutathione ratio levels were significantly reduced in both transgenic lines when compared with SHRSP [malondialdehyde: SHRSP 0.04 0.009mmol/l vs. SHRSP-Tg(Gstm1)1WKY 0.024 0.002mmol/l and SHRSPTg(Gstm1)2WKY 0.021 0.002mmol/l; (oxidized : reduced glutathione ratio): SHRSP 5.19 2.26mmol/l vs. SHRSPTg(Gstm1)1WKY 0.17 0.111mmol/l and SHRSPTg(Gstm1)2WKY 0.471 0.223mmol/l]. Transgenic SHRSP rats containing the WKY Gstm1 gene demonstrate significantly lower BP, reduced oxidative stress and improved levels of renal Gstm1 expression. Conclusion: These data support the hypothesis that reduced renal Gstm1 plays a role in the development of hypertension

New insights into the genetic control of gene expression using a Bayesian multi-tissue approach.

The majority of expression quantitative trait locus (eQTL) studies have been carried out in single tissues or cell types, using methods that ignore information shared across tissues. Although global analysis of RNA expression in multiple tissues is now feasible, few integrated statistical frameworks for joint analysis of gene expression across tissues combined with simultaneous analysis of multiple genetic variants have been developed to date. Here, we propose Sparse Bayesian Regression models for mapping eQTLs within individual tissues and simultaneously across tissues. Testing these on a set of 2,000 genes in four tissues, we demonstrate that our methods are more powerful than traditional approaches in revealing the true complexity of the eQTL landscape at the systems-level. Highlighting the power of our method, we identified a two-eQTL model (cis/trans) for the Hopx gene that was experimentally validated and was not detected by conventional approaches. We showed common genetic regulation of gene expression across four tissues for ∼27% of transcripts, providing >5 fold increase in eQTLs detection when compared with single tissue analyses at 5% FDR level. These findings provide a new opportunity to uncover complex genetic regulatory mechanisms controlling global gene expression while the generality of our modelling approach makes it adaptable to other model systems and humans, with broad application to analysis of multiple intermediate and whole-body phenotypes

Contribution of Autosomal Loci and the Y Chromosome to the Stress Response in Rats

Stress is a critical contributor to cardiovascular diseases through its impact on blood pressure variability and cardiac function. Familial clustering of reactivity to stress has been demonstrated in human subjects, and some rodent models of hypertension are hyperresponsive to stress. Therefore, the present study was designed to uncover the genetic determinants of the stress response. We performed a total genome linkage search to identify the loci of the body temperature response to immobilization stress in a set of recombinant inbred strains (RIS) originating from reciprocal crosses of spontaneously hypertensive rats (SHR) with a normotensive Brown Norway Lx strain. Two quantitative trait loci (QTLs) were revealed on chromosomes (Chrs) 10 and 12 (logarithm of odds scores, 2.2 and 1.3, respectively). The effects of these QTLs were enhanced by a high sodium diet (logarithm of odds scores, 4.0 and 3.3 for Chrs 10 and 12, respectively), which is suggestive of a salt-sensitive component for the phenotype, Congenics for Chr 10 confirmed both the QTL and the salt effect in RIS. Negatively associated loci were also identified on Chrs 8 and 11. Interaction between the loci of Chrs 10 and 12 was demonstrated, with the rat strains bearing SHR alleles at both loci having the highest thermal response to stress. Furthermore, the Y Chr of SHR origin enhanced the response to immobilization stress, as demonstrated in 2 independent models, RIS and Y Chr consomics. However, its full effect requires autosomes of the SHR strain. These findings provide the first evidence for the genetic determination of reactivity to stress with interactions between autosomal loci and between the Y and autosomal Chrs that contribute to the explanation of the 46% of variance in the stress response

Towards precision medicine for hypertension: a review of genomic, epigenomic, and microbiomic effects on blood pressure in experimental rat models and humans

Compelling evidence for the inherited nature of essential hypertension has led to extensive research in rats and humans. Rats have served as the primary model for research on the genetics of hypertension resulting in identification of genomic regions that are causally associated with hypertension. In more recent times, genome-wide studies in humans have also begun to improve our understanding of the inheritance of polygenic forms of hypertension. Based on the chronological progression of research into the genetics of hypertension as the "structural backbone," this review catalogs and discusses the rat and human genetic elements mapped and implicated in blood pressure regulation. Furthermore, the knowledge gained from these genetic studies that provide evidence to suggest that much of the genetic influence on hypertension residing within noncoding elements of our DNA and operating through pervasive epistasis or gene-gene interactions is highlighted. Lastly, perspectives on current thinking that the more complex "triad" of the genome, epigenome, and the microbiome operating to influence the inheritance of hypertension, is documented. Overall, the collective knowledge gained from rats and humans is disappointing in the sense that major hypertension-causing genes as targets for clinical management of essential hypertension may not be a clinical reality. On the other hand, the realization that the polygenic nature of hypertension prevents any single locus from being a relevant clinical target for all humans directs future studies on the genetics of hypertension towards an individualized genomic approach

Targeting of the Plzf gene in the rat by transcription activator-like effector nuclease results in caudal regression syndrome in spontaneously hypertensive rats

Recently, it has been found that spontaneous mutation Lx (polydactyly-luxate syndrome) in the rat is determined by deletion of a conserved intronic sequence of the Plzf (Promyelocytic leukemia zinc finger protein) gene. In addition, Plzf is a prominent candidate gene for quantitative trait loci (QTLs) associated with cardiac hypertrophy and fibrosis in the spontaneously hypertensive rat (SHR). In the current study, we tested the effects of Plzf gene targeting in the SHR using TALENs (transcription activator-like effector nucleases). SHR ova were microinjected with constructs pTAL438/439 coding for a sequence-specific endonuclease that binds to target sequence in the first coding exon of the Plzf gene. Out of 43 animals born after microinjection, we detected a single male founder. Sequence analysis revealed a deletion of G that resulted in frame shift mutation starting in codon 31 and causing a premature stop codon at position of amino acid 58. The Plzftm1Ipcv allele is semi-lethal since approximately 95% of newborn homozygous animals died perinatally. All homozygous animals exhibited manifestations of a caudal regression syndrome including tail anomalies and serious size reduction and deformities of long bones, and oligo- or polydactyly on the hindlimbs. The heterozygous animals only exhibited the tail anomalies. Impaired development of the urinary tract was also revealed: one homozygous and one heterozygous rat exhibited a vesico-ureteric reflux with enormous dilatation of ureters and renal pelvis. In the homozygote, this was combined with a hypoplastic kidney. These results provide evidence for the important role of Plzf gene during development of the caudal part of a body-column vertebrae, hindlimbs and urinary system in the rat

Translational regulation shapes the molecular landscape of complex disease phenotypes.

The extent of translational control of gene expression in mammalian tissues remains largely unknown. Here we perform genome-wide RNA sequencing and ribosome profiling in heart and liver tissues to investigate strain-specific translational regulation in the spontaneously hypertensive rat (SHR/Ola). For the most part, transcriptional variation is equally apparent at the translational level and there is limited evidence of translational buffering. Remarkably, we observe hundreds of strain-specific differences in translation, almost doubling the number of differentially expressed genes. The integration of genetic, transcriptional and translational data sets reveals distinct signatures in 3'UTR variation, RNA-binding protein motifs and miRNA expression associated with translational regulation of gene expression. We show that a large number of genes associated with heart and liver traits in human genome-wide association studies are primarily translationally regulated. Capturing interindividual differences in the translated genome will lead to new insights into the genes and regulatory pathways underlying disease phenotypes