REGULATION OF PEROXISOME PROLIFERATOR-ACTIVATED RECEPTOR ALPHA ACTIVITY VIA CROSS-TALK WITH GLYCOGEN SYNTHASE KINASE 3
Open Access
- Author:
- Burns, Katherine Anne
- Graduate Program:
- Pathobiology
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- March 08, 2007
- Committee Members:
- John Patrick Vanden Heuvel, Committee Chair/Co-Chair
Pamela Hankey Giblin, Committee Member
Donna Hope Korzick, Committee Member
Gary H Perdew, Committee Member
Jeffrey Maurice Peters, Committee Member - Keywords:
- fasting
phosphorylation
GSK3
PPARalpha - Abstract:
- Peroxisome proliferator-activated receptor alpha (PPAR alpha) is a nuclear receptor involved in the maintenance of fatty acid homeostasis and is important for mediating the adaptive response to fasting. PPAR alpha is phosphorylated by multiple kinases which differentially regulate its activity. Computational analysis of PPAR alpha revealed 5 potential glycogen synthase kinase 3 (GSK3) phosphorylation sites. GSK3 is a key enzyme involved in glycogen metabolism, and is important for signaling by insulin, growth factors, and nutrients. It was hypothesized that PPAR alpha is a substrate for GSK3 phosphorylation and this association is involved in fasting mediated homeostasis. PPAR alpha is phosphorylated by GSK3 beta, primarily at serine-73 in the A/B domain of rat PPAR alpha. This residue is conserved across PPAR alpha species including humans. The regulation of PPAR alpha target gene expression was affected by manipulating GSK3 activity via chemical inhibitors and specific RNA inhibitors (RNAis). We also demonstrated an increased turnover and ubiquitination of PPAR alpha with the over-expression of GSK3 beta, presumably the result of increased PPAR alpha phosphorylation. In addition to GSK3 affecting PPAR alpha activity, it has been noted that ligands for this nuclear receptor, the peroxisome proliferators, influence GSK3 alpha and GSK3 beta expression and activity. The adaptive response in the liver to fasting involves both PPAR alpha and GSK3. Thus, mice subjected to fasting represents an ideal, physiologically-relevant model system to explore PPAR alpha and GSK3 activity and their confluence. Fatty acid metabolism genes, such as Acyl CoA Oxidase (ACO), the bifunctional enzyme (BIEN), and 3-ketoacyl-CoA thiolase (thiolase) were increased during fasting in wild-type (PPAR alpha +/+) but not in PPAR alpha knockout (PPAR alpha -/-) mice. GSK3 alpha was increased at the mRNA and protein level, while GSK3 beta was decreased at the phosphospecific level by fasting in wild-type mice. Two potential mechanistic links between PPAR alpha activation upon fasting and GSK3 expression and activity were examined, Tribbles 3 (TRB3) and Peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1 alpha). TRB3 is a fasting inducible inhibitor of the serine-threonine kinase AKT/PKB and PGC-1 alpha is a ligand inducible coactivator for PPAR alpha involved in the control of cellular energy metabolic pathways. In contrast to previously published results, TRB3 was not differentially regulated by fasting in wild-type or PPAR alpha -/- mice at the mRNA or protein level. However, PGC-1 alpha mRNA levels were increased upon fasting, and unexpectedly, the levels were increased in a PPAR alpha-dependent manner. This establishes a novel regulation of PGC-1 alpha that is dependent on the expression of PPAR alpha during fasting. This thesis has demonstrated that PPAR alpha is a substrate for GSK3 beta phosphorylation that causes increased ubiquitination and turnover of PPAR alpha, and that PPAR alpha, GSK3 alpha/beta, and PGC-1 alpha are coordinately regulated during a 24 hr fast in the murine model. These studies reveal how these pathways converge and are not only important for understanding what occurs during fasting, but may also be used to further study other metabolic states, namely insulin resistance. Clearer understanding of the interconnectedness of PPAR alpha, GSK3 alpha/beta, and PGC-1 alpha can ultimately aid in how metabolic syndrome and other related diseases are approached and treated.