Date on Master's Thesis/Doctoral Dissertation

8-2016

Document Type

Doctoral Dissertation

Degree Name

Ph. D.

Department

Physiology and Biophysics

Degree Program

Physiology and Biophysics, PhD

Committee Chair

Jones, Steven

Committee Co-Chair (if applicable)

Joshua, Irving

Committee Member

Joshua, Irving

Committee Member

Schuschke, Dale

Committee Member

Maldonado, Claudio

Committee Member

Hill, Bradford

Author's Keywords

Myocardial Infarction; OGT; OGA; Heart failure; Diabetes

Abstract

Global augmentation of protein O-GlcNAcylation occurs in response to a myriad of stressors and confers a survival advantage at the cellular level. This protective phenomenon has been demonstrated to mediate cardioprotection through various in vitro and in vivo studies during ischemia-reperfusion, myocardial infarction, and oxidative stress; however, relatively little is known of the regulation of protein O-GlcNAcylation. Protein O-GlcNAcylation is regulated by two antagonistic enzymes, namely, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). Ablation of cardiomyocyte OGT, the enzyme that catalyzes the addition of O-GlcNAc to proteins, exacerbates cardiac dysfunction during infarct-induced heart failure (HF). However, little is known of the enzyme mediating the removal of the O-GlcNAc modification, OGA, in the context of HF. The present study focused on this limitation in the field. We characterized the temporal expression of OGA following myocardial infarction (MI) and found that OGA expression is decreased and remains suppressed for 4 wk post MI. Conversely, OGT expression is augmented early, but normalizes by 4 wk post MI. Despite the normalization of OGT expression, O-GlcNAcylation remains elevated, which may be due to chronic OGA suppression. Furthermore, we observed upregulation of miRNA-539 in HF. In vitro studies confirmed induction of miRNA-539 negatively regulated OGA expression. These data indicate that suppression of OGA could be mediated by miRNA-539. Next, we developed a genetic model of cardiomyocyte specific OGA ablation to test whether ablation of OGA would augment O-GlcNAcylation and attenuate HF. Our model successfully suppressed OGA expression, augmented cardiac protein O-GlcNAcylation, and did not induce cardiac dysfunction; however, genetic ablation of OGA prior to coronary ligation hastened cardiac dysfunction within 1 wk compared to wild-type mice. Hearts from OGA KO mice were more dilated and less efficient, which suggests rejection of our hypothesis. These data indicate that OGA expression may be proadaptive during HF. Because O-GlcNAcylation of mitochondrial complexes has been implicated to depress mitochondrial respiration we hypothesized that augmented O-GlcNAcylation may mediate mitochondrial dysfunction and may help explain the exacerbation in cardiac dysfunction we observed after 1 wk of HF. We virally augmented either OGT or OGA in cardiomyocytes to alter overall protein O-GlcNAcylation. Neither overexpression of OGA nor OGT mediated mitochondrial dysfunction. Though induction of O-GlcNAcylation through hyperglycemia did suppress mitochondrial reserve capacity. This depression in mitochondrial function was recapitulated with an osmotic control. We concluded that modulation of O-GlcNAc alone did not cause mitochondrial dysfunction. These data indicate that suppression of OGA occurs during HF and may be mediated by posttranscriptional regulation by miR-539. In addition, ablation of OGA expression can hasten HF. Furthermore, the exacerbation in cardiac dysfunction is not likely due to O-GlcNAc-mediated mitochondrial dysfunction. These data indicate that chronic augmentation of O-GlcNAcylation may be detrimental in HF. More specifically they indicate that dynamic cycling of O-GlcNAcylation may be more beneficial in HF than permanently driving O-GlcNAc levels in one direction.

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