I. ABSTRACT Heart disease remains the leading cause of death in the United States. Despite what we know about the risk factors associated with heart disease, the molecular mechanisms are still largely unknown. The healthy adult heart is unique from other tissues in that the rate of protein synthesis is dramatically lower than in most tissues and lower than the developing heart. However, during cardiac hypertrophy, translation rates increase. This suggests a tissue-specific mechanism for regulating translation rates in the mammalian heart. Our lab has identified a mechanism by which total protein synthesis in cardiomyocytes is decreased during development through shortening of poly(A) tails, leading to a decrease in polysome formation through the closed-loop model of translation. This regulation is reversed during both physiologic and pathologic hypertrophy when the translation needs of cardiomyocytes are increased. Also, we have discovered that the nuclear poly(A) binding protein (PABPN1) is post-transcriptionally silenced in mammalian adult cardiac and skeletal muscle but it becomes re-expressed in pathologic cardiac hypertrophy. PABPN1 is a regulator of alternative polyadenylation (APA) and poly(A) tail length, both of which can influence the translation of transcripts. Our central hypothesis is that PABPN1 is dynamically regulated in cardiac myocytes to tune translation rates and suite growth needs through a polyadenylation dependent mechanism. The objective of this proposal is to elucidate the exact function(s) of PABPN1 in cardiac development and growth and identify how PABPN1 is regulated during these conditions.
Aims 1 and 2 will use conditional PABPN1-knockout and overexpressing mice to determine the physiologic roles of PABPN1 in cardiac development and hypertrophy while defining the molecular basis of PABPN1 activity and its role in determining cardiac-specific gene expression programs.
In Aim 3, we will use super-resolution microscopy, CRISPR-Cas9 mediated genome editing, and RNA antisense-oligo pulldown approaches to identify the regulatory mechanism(s) and factors that post-transcriptionally silence PABPN1 during cardiac development.
The proposed studies will provide a deep understanding of the genetic programs that regulate normal cardiac growth during development; and reactivate fetal gene expression in adult heart under disease conditions. This project will also dissect the functional role of a nuclear poly (A) binding protein in stimulation of cardiac hypertrophy, a condition that often accompanies the response to myocardial infarction and heart failure. Completion of the proposed aims will provide unprecedented insight into how the cardiac transcriptome is shaped into the cardiac translatome and potentially uncover new regulatory pathways to target with therapeutics.
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