Ankyrin-B protein in heart failure: identification of a new component of metazoan cardioprotection.
ABSTRACT Ankyrins (ankyrin-R, -B, and -G) are adapter proteins linked with defects in metazoan physiology. Ankyrin-B (encoded by ANK2) loss-of-function mutations are directly associated with human cardiovascular phenotypes including sinus node disease, atrial fibrillation, ventricular tachycardia, and sudden cardiac death. Despite the link between ankyrin-B dysfunction and monogenic disease, there are no data linking ankyrin-B regulation with common forms of human heart failure. Here, we report that ankyrin-B levels are altered in both ischemic and non-ischemic human heart failure. Mechanistically, we demonstrate that cardiac ankyrin-B levels are tightly regulated downstream of reactive oxygen species, intracellular calcium, and the calcium-dependent protease calpain, all hallmarks of human myocardial injury and heart failure. Surprisingly, β(II)-spectrin, previously thought to mediate ankyrin-dependent modulation in the nervous system and heart, is not coordinately regulated with ankyrin-B or its downstream partners. Finally, our data implicate ankyrin-B expression as required for vertebrate myocardial protection as hearts deficient in ankyrin-B show increased cardiac damage and impaired function relative to wild-type mouse hearts following ischemia reperfusion. In summary, our findings provide the data of ankyrin-B regulation in human heart failure, provide insight into candidate pathways for ankyrin-B regulation in acquired human cardiovascular disease, and surprisingly, implicate ankyrin-B as a molecular component for cardioprotection following ischemia.
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ABSTRACT: Spatial and timely variations in QT interval, even within its normal range, may underlie susceptibility to cardiac arrhythmias and sudden cardiac death. Given its important role in cardiac electrophysiology, we hypothesized that common genetic variation in ankyrin-B gene (ANK2) might modify QT interval length. The study population consisted of 1188 participants of the World Health Organizational Multinational Monitoring of Trends and Determinants in Cardiovascular Disease (WHO MONICA) general population survey Cooperative Health Research in the Region of Augsburg (KORA S3). Corrected QT interval was calculated using population specific linear regression formulas. A total of 22 single-nucleotide polymorphisms in the genomic region of ANK2 gene were genotyped using TaqMan technology. In a replication study, 6 single nucleotide polymorphisms were genotyped in 3890 individuals from a second population study (KORA S4). The rare variant of the single-nucleotide polymorphism rs6850768 (allele frequency, 0.28) significantly influenced duration of the QT interval, both in KORA S3 and KORA S4 populations. In homozygotes, the shortening of the QT interval was 3.79 ms (95% CI, 1.48 to 5.58; P=0.001 and P=0.0008 for log-additive and dominant model, respectively) in KORA S3 and 2.94 ms (95% CI, 1.11 to 4.77; P=0.001 and P=0.006 for log-additive and dominant genetic model, respectively) in KORA S4. A common 2-locus haplotype (rs11098171-rs6850768; population frequency, 28%) was associated with a QT interval difference of 2.85 ms (permutation; P=0.006) in KORA S3 and 1.23 ms (permutation; P=0.009) in KORA S4. Reverse transcription-polymerase chain reaction expression analysis of the human ANK2 5' genomic region in the human left ventricular tissue revealed 2 previously unidentified ANK2 5' exons in the proximity of the identified variants. Common genetic variants juxtaposed with novel exons in the distant 5' genomic region of ANK2 influence the QT interval length in the general population. These findings support the role of ankyrin-B in normal cardiac electric activity.Circulation Cardiovascular Genetics 12/2008; 1(2):93-9. · 6.73 Impact Factor
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ABSTRACT: Mice with genetic inhibition (AC3-I) of the multifunctional Ca(2+)/calmodulin dependent protein kinase II (CaMKII) have improved cardiomyocyte survival after ischemia. Some K(+) currents are up-regulated in AC3-I hearts, but it is unknown if CaMKII inhibition increases the ATP sensitive K(+) current (I(KATP)) that underlies ischemic preconditioning (IP) and confers resistance to ischemia. We hypothesized increased I(KATP) was part of the mechanism for improved ventricular myocyte survival during ischemia in AC3-I mice. AC3-I hearts were protected against global ischemia due to enhanced IP compared to wild type (WT) and transgenic control (AC3-C) hearts. IKATP was significantly increased, while the negative regulatory dose-dependence of ATP was unchanged in AC3-I compared to WT and AC3-C ventricular myocytes, suggesting that CaMKII inhibition increased the number of functional I(KATP) channels available for IP. We measured increased sarcolemmal Kir6.2, a pore-forming I(KATP) subunit, but not a change in total Kir6.2 in cell lysates or single channel I(KATP) opening probability from AC3-I compared to WT and AC3-C ventricles, showing CaMKII inhibition increased sarcolemmal I(KATP) channel expression. There were no differences in mRNA for genes encoding I(KATP) channel subunits in AC3-I, WT and AC3-C ventricles. The I(KATP) opener pinacidil (100 microM) reduced MI area in WT to match AC3-I hearts, while the I(KATP) antagonist HMR1098 (30 microM) increased MI area to an equivalent level in all groups, indicating that increased I(KATP) and augmented IP are important for reduced ischemic cell death in AC3-I hearts. Our study results show CaMKII inhibition enhances beneficial effects of IP by increasing I(KATP).Channels (Austin, Tex.) 01/2007; 1(5):387-94. · 1.91 Impact Factor
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ABSTRACT: Ankyrins contain significant amino acid identity and are co-expressed in many cell types yet maintain unique functions in vivo. Recent studies have identified the highly divergent C-terminal domain in ankyrin-B as the key domain for driving ankyrin-B-specific functions in cardiomyocytes. Here we identify an intramolecular interaction between the C-terminal domain and the membrane-binding domain of ankyrin-B using pure proteins in solution and the yeast two-hybrid assay. Through extensive deletion and alanine-scanning mutagenesis we have mapped key residues for interaction in both domains. Amino acids (1597)EED(1599) located in the ankyrin-B C-terminal domain and amino acids Arg(37)/Arg(40) located in ANK repeat 1 are necessary for inter-domain interactions in yeast two-hybrid assays. Furthermore, conversion of amino acids EED(1597) to AAA(1597) leads to a loss of function in the localization of inositol 1,4,5-trisphosphate receptors in ankyrin-B mutant cardiomyocytes. Physical properties of the ankyrin-B C-terminal domain determined by circular dichroism spectroscopy and hydrodynamic parameters reveal it is unstructured and highly extended in solution. Similar structural studies performed on full-length 220-kDa ankyrin-B harboring alanine substitutions, (1597)AAA(1599), reveal a more extended conformation compared with wild-type ankyrin-B. Taken together these results suggest a model of an extended and unstructured C-terminal domain folding back to bind and potentially regulate the membrane-binding domain of ankyrin-B.Journal of Biological Chemistry 04/2006; 281(9):5741-9. · 4.65 Impact Factor