Regulation of Ribosomal DNA Transcription during Contraction-induced Hypertrophy of Neonatal Cardiomyocytes

Abstract

Cardiac hypertrophy requires protein accumulation. This results largely from an increased capacity for protein synthesis, which in turn is the result of an elevated rate of ribosome biogenesis. The process of ribosome formation is regulated at the level of transcription of the ribosomal RNA genes. In this study, we examined the amounts and activities of various components of the ribosomal DNA transcription apparatus in contraction-arrested neonatal cardiomyocytes and in spontaneously contracting cardiomyocytes that hypertrophy. Nuclear run-on assays demonstrated that spontaneously contracting cardiomyocytes supported a 2-fold increased rate of ribosomal DNA transcription. However, enzymatic assay of total solubilized RNA polymerase I and Western blots demonstrated that contraction-induced increases in ribosomal RNA synthesis were not accompanied by increased activity or amounts of RNA polymerase I. In contrast, accelerated ribosome biogenesis was accompanied by an increased amount of the ribosomal DNA transcription factor, UBF. Immunoprecipitation of [32P]orthophosphate-labeled UBF from hypertrophying, neonatal cardiomyocytes indicated that the accumulated UBF protein was phosphorylated and, thus, in the active form. UBF mRNA levels began to increase within 3-6 h of the initiation of contraction and preceded the elevation in rDNA transcription. Nuclear run-on assays demonstrated increased rates of transcription of the UBF gene. Transfection of chimeric reporter constructs containing deletions of the 5′-flanking region of the UBF gene revealed the presence of contraction response elements between −1189 and −665 relative to the putative start of transcription. These results are consistent with the hypothesis that UBF is an important factor in the regulation of rDNA transcription during contraction-mediated neonatal cardiomyocyte hypertrophy.