Genetic variants associated with risk of atrial fibrillation regulate expression of PITX2, CAV1, MYOZ1, C9orf3 and FANCC.

Genome-wide association studies (GWAS) have identified genetic variants in a number of chromosomal regions that are associated with atrial fibrillation (AF). The mechanisms underlying these associations are unknown, but are likely to involve effects of the risk haplotypes on expression of neighbouring genes. To investigate the association between genetic variants at AF-associated loci and expression of nearby candidate genes in human atrial tissue and peripheral blood. Right atrial appendage (RAA) samples were collected from 122 patients undergoing cardiac surgery, of these, 12 patients also had left atrial appendage samples taken. 22 patients had a history of AF. Peripheral blood samples were collected from 405 patients undergoing diagnostic cardiac catheterisation. In order to tag genetic variation at each of nine loci, a total of 367 single nucleotide polymorphisms (SNPs) were genotyped using the Sequenom platform. Total expression of 16 candidate genes in the nine AF-associated regions was measured by quantitative PCR. The relative expression of each allele of the candidate genes was measured on the Sequenom platform using one or more transcribed SNPs to distinguish between alleles in heterozygotes. We tested association between the SNPs of interest and gene expression using total gene expression (integrating cis and trans acting sources of variation), and allelic expression ratios (specific for cis acting influences), in atrial tissue and peripheral blood. We adjusted for multiple comparisons using a Bonferroni approach. In subsidiary analyses, we compared the expression of candidate genes between patients with and without a history of AF. Total expression of 15 transcripts of 14 genes and allelic expression ratio of 14 transcripts of 14 genes in genomic regions associated with AF were measured in right atrial appendage tissue. 8 of these transcripts were also expressed in peripheral blood. Risk alleles at AF-associated SNPs were associated in cis with an increased expression of PITX2a (2.01-fold, p=6.5×10(-4)); and with decreased expression of MYOZ1 (0.39 fold; p=5.5×10(-15)), CAV1 (0.89 fold; p=5.9×10(-8)), C9orf3 (0.91 fold; 1.5×10(-5)), and FANCC (0.94-fold; p=8.9×10(-8)) in right atrial appendage. Of these five genes, only CAV1 was expressed in peripheral blood; association between the same AF risk alleles and lower expression of CAV1 was confirmed (0.91 fold decrease; p=4.2×10(-5)). A history of AF was also associated with a decrease in expression of CAV1 in both right and left atria (0.84 and 0.85 fold, respectively; p=0.03), congruent with the magnitude of the effect of the risk SNP on expression, and independent of genotype. The analyses in peripheral blood showed association between AF risk SNPs and decreased expression of KCNN3 (0.85-fold; p=2.1×10(-4)); and increased expression of SYNE2 (1.12-fold; p=7.5×10(-24)); however, these associations were not detectable in atrial tissue. We identified novel cis-acting associations in atrial tissue between AF risk SNPs and increased expression of PITX2a/b; and decreased expression of CAV1 (an association also seen in peripheral blood), C9orf3 and FANCC. We also confirmed a previously described association between AF risk variants and MYOZ1 expression. Analyses of peripheral blood illustrated tissue-specificity of cardiac eQTLs and highlight the need for larger-scale genome-wide eQTL studies in cardiac tissue. Our results suggest novel aetiological roles for genes in four AF-associated genomic regions.

The landscape of DNA repeat elements in human heart failure.

BACKGROUND: The epigenomes of healthy and diseased human hearts were recently examined by genome-wide DNA methylation profiling. Repetitive elements, heavily methylated in post-natal tissue, have variable methylation profiles in cancer but methylation of repetitive elements in the heart has never been examined. RESULTS: We analyzed repetitive elements from all repeat families in human myocardial samples, and found that satellite repeat elements were significantly hypomethylated in end-stage cardiomyopathic hearts relative to healthy normal controls. Satellite repeat elements are almost always centromeric or juxtacentromeric, and their overexpression correlates with disease aggressiveness in cancer. Similarly, we found that hypomethylation of satellite repeat elements correlated with up to 27-fold upregulation of the corresponding transcripts in end-stage cardiomyopathic hearts. No other repeat family exhibited differential methylation between healthy and cardiomyopathic hearts, with the exception of the Alu element SINE1/7SL, for which a modestly consistent trend of increased methylation was observed. CONCLUSIONS: Satellite repeat element transcripts, a form of non-coding RNA, have putative functions in maintaining genomic stability and chromosomal integrity. Further studies will be needed to establish the functional significance of these non-coding RNAs in the context of heart failure.

MICA polymorphism: biology and importance in immunity and disease.

The human major histocompatibility complex class I chain-related gene A (MICA) is one of the genes in the HLA class I region of chromosome 6. Unlike HLA classical class I gene products, MICA does not present any antigen but acts as a ligand for several immune cells including natural killer (NK) cells bearing NKG2D receptors. MICA is the member of the non-classical class I family that displays the greatest degree of polymorphism. MICA alleles can be divided into two large groups with the polymorphisms found in alpha3 domains. This division could be explained by a possible polyphyletic origin that is in line with recent findings from evolutionary, population and functional studies of this gene. MICA polymorphisms are associated with a number of diseases related to NK activity, such as viral infection, cancer and allograft rejection or graft-versus-host disease (GVHD). The mechanisms underlying these associations include NK cell-mediated cytotoxicity and MICA shedding to produce immunosuppressive soluble MICA particles. The MICA-induced humoral response has attracted interest recently because of its possible role in graft rejection in solid organ transplantation. Here, we discuss the genetics and biology of the MICA gene and its products, and their importance in disease.

PKB/Akt activation inhibits p53-mediated HIF1A degradation that is independent of MDM2.

Cross-talk between the two transcription factors, p53 and hypoxia inducible factor 1alpha (HIF1A), is important in different pathophysiological conditions (Hammond and Giaccia, 2006, Clin Cancer Res 12:5007-5009) such as in the transition from myocardial hypertrophy to cardiac dilatation and heart failure. In that context, p53 induces HIF1A degradation which in turn provokes the transition from compensatory hypertrophy to myocardial thinning and chamber dilatation (Sano et al., 2007, Nature 446:444-448). In order to investigate the mechanism of p53-induced HIF1A degradation, we used the established in vitro model of deferroxamine (DFX)-induced HIF1A accumulation in H9c2 cardiac cells (Sano et al., 2007, Nature 446:444-448). Here, we report that opposite to HIF1A accumulation following exposure to DFX, prolonged DFX-induced p53 activation and HIF1A protein decrease, without any change in Hif1a mRNA. HIF1A protein decrease accompanied upregulated HIF1A ubiquitination. MDM2, an ubiquitin E3 ligase target gene of p53, was upregulated following prolonged DFX, but using p53/Mdm2 double-null mouse embryonic fibroblasts, we found that p53 upregulated HIF1A ubiquitination and degradation independently of MDM2. Moreover, with prolonged DFX treatment, an enhanced interaction between MDM2 and HIF1A was lacking. Instead, phospho-Akt(ser473) was decreased during the phase coinciding with HIF1A degradation, and inhibition of PKB/Akt phosphorylation using PI3K inhibitor (LY294002) upregulated HIF1A ubiquitination. In summary, we propose that p53-induced HIF1A degradation is not exclusively MDM2-mediated, but reversible by PKB/Akt phosphorylation.

Possible polyphyletic origin of major histocompatibility complex class I chain-related gene A (MICA) alleles.

Phylogenetic relationships among 23 nonhuman primate (NHP) major histocompatibility complex class I chain-related gene (MIC) sequences, 54 confirmed human MICA alleles, and 16 human MICE alleles were constructed with methods of sequence analysis. Topology of the phylogenetic tree showed separation between NHP MICs and human MICs. For human MICs, the topology indicated monophyly for the MICB alleles, while MICA alleles were separated into two lineages, LI and LII. Of these, LI MICA alleles shared a common ancestry with gorilla (Ggo) MIC. One conservative amino acid difference and two nonconservative amino acid differences in the alpha3 domain were found between the MICA lineages. The nonconservative amino acid differences might imply structural and functional differences. Transmembrane (TM) trinucleotide-repeat variants were found to be specific to the MICA lineages such as A4, A9, and A10 to LI and A5 to LII. Variants such as A5.1 and A6 were commonly found in both MICA lineages. Based on these analyses, we postulate a polyphyletic origin for MICA alleles and their division into two lineages, LI and LII. As such, there would be 30 alleles in LI and 24 alleles in LII, thereby reducing the current level of polymorphism that exists, based on a presumed monophyletic origin. The lower degree of polymorphism in MICA would then be in line with the rest of the human major histocompatibility complex nonclassical class I genes.