Regulation of rDNA Transcription During Endothelin-1Induced Hypertrophy of Neonatal Cardiomyocytes: Hyperphosphorylation of Upstream Binding Factor, an rDNA Transcription Factor

Abstract

Treatment of cultured neonatal cardiomyocytes with endothelin-1 and phorbol 12-myristate 13-acetate (PMA) results in cardiomyocyte hypertrophy. However, the signal transduction pathways involved in this process are poorly understood. Because increased ribosome biogenesis is a requisite for hypertrophy, we sought to (1) confirm the hypothesis that these two hypertrophic agents did indeed induce rRNA synthesis and (2) examine the mechanism through which this induction was accomplished. In this study, hypertrophy of contraction-arrested neonatal cardiomyocytes induced by treatment with either endothelin-1 or PMA was associated with increased rDNA transcription. Western blots demonstrated that the enhanced rates of rDNA transcription were not mediated by increased amounts of either RNA polymerase I or upstream binding factor (UBF), an rDNA transcription factor. However, immunoprecipitation of [32P]orthophosphate-labeled UBF from hypertrophying neonatal cardiomyocytes suggested that the increased rate of rDNA transcription may be due to the hyperphosphorylation of UBF, which would increase the activity of UBF. The increase in UBF phosphorylation occurred within 3 to 6 hours after exposure to either agent, was maximal at 12 hours, and was sustained for at least the first 24 hours of exposure. Phosphoamino acid analysis of UBF immunoprecipitated from control and treated cardiomyocytes demonstrated that UBF was phosphorylated exclusively on serine residues. Our previous studies have shown that the cellular UBF content increased in adrenergic- and contraction-induced models of cardiac hypertrophy. This study with endothelin-1 and PMA demonstrates that the modulation of UBF phosphorylation is an additional pathway by which ribosome biogenesis may be regulated in neonatal cardiomyocytes. These results support the hypothesis that UBF is an important regulatory factor during the initiation and maintenance of the accelerated rate of rDNA transcription observed during neonatal cardiomyocyte hypertrophy mediated by both phorbol esters and endothelin-1.