Role of mRNA-binding protein tristetraprolin in cardiac mRNA regulation and the development of heart failure

Heart failure (HF) is a major health epidemic in developed countries, however, its underlying pathology is not well characterized. Tristetraprolin (TTP) is a tandem zinc finger protein that binds to AU-rich elements (ARE) in the 3’-untranslated region (UTR) of target mRNA molecules, and induces their degradation. Global TTP knockout

(KO) mice display systemic inflammation, since TNFα mRNA is normally degraded by TTP, and deletion of TTP leads to elevated TNFα levels. Thus, very few studies have assessed the role of TTP in metabolism despite its original discovery as an insulin-inducible gene, and genetic studies linking TTP to metabolic syndrome. We are

addressing this fundamental gap in knowledge, and our strong preliminary data suggest critical activities by TTP in cardiac metabolism and the development of HF. Specifically, we have shown that TTP inhibits fatty acid (FA) and branched-chain amino acid (BCAA) metabolism (independent of its effects on inflammation), and reduces

the mRNA levels of key proteins in these processes, i.e., peroxisome proliferator-activated receptor (PPAR)-α and branched-chain α–ketoacid acid dehydrogenase complex (BCKDC)-E2 subunits. The central hypothesis of this proposal is that TTP inhibits cardiac FA and BCAA metabolism by binding to and degrading

PPARα and BCKDC-E2 mRNAs, and that TTP exacerbates the development of HF by impairing FA and BCAA metabolism. In Aim 1, we will assess whether TTP inhibits cardiac FA metabolism by binding to PPARα mRNA and promoting its degradation. We will assess whether TTP binds to PPARα mRNA by performing RNA

co-immunopreciptation (co-IP) and deletion studies of PPARα 3’-UTR AREs. We will also measure FA uptake and metabolism in the hearts from cardiac-specific TTP KO (csTTP-KO) mice, and will determine whether these changes are through PPARα using TTP/PPARα double KO mice. In Aim 2, we will determine whether TTP

decreases BCAA catabolism through binding and degradation of BCKDC-E2 mRNA. We will first determine whether TTP binds BCKDC-E2 mRNA by performing RNA co-IP and deletion studies on BCKDC-E2 3’-UTR AREs. We will also measure BCAA levels and BCKDC activity in heart tissue from csTTP-KO mice. To demonstrate whether the reduction in BCAA catabolism with TTP KO is through BCKDC-E2, we will perform

similar studies with knockdown of TTP and BCKDC-E2. In Aim 3, we will determine whether TTP has detrimental effects on the heart under stress conditions, and whether this depends upon impaired FA and BCAA metabolism. We will subject csTTP-KO mice to pressure overload and ischemia, then assess their cardiac function and

metabolism. To determine the role of PPARα and BCKDC in this process, we will use csTTP/PPARα double KO mice and will cross csTTP-KO with protein phosphatase 2Cm KO mice (which have reduced BCKDC activity), and assess cardiac response to stress. We will also show our studies to develop novel drugs that target TTP

without inducing inflammation, providing potential clinical implications for Aim 3. These studies will improve our understanding of cardiac metabolism, and may lead to new avenues for treatment of HF.