p90 Ribosomal S6 kinases play a significant role in early gene regulation in the cardiomyocyte response to Gq-protein-coupled receptor stimuli, endothelin-1 and α1-adrenergic receptor agonists

Abstract

ERK1/2 (extracellular-signal-regulated kinase 1/2) and their substrates RSKs (p90 ribosomal S6 kinases) phosphorylate different transcription factors, contributing differentially to transcriptomic profiles. In cardiomyocytes ERK1/2 are required for >70% of the transcriptomic response to endothelin-1. In the present study we investigated the role of RSKs in the transcriptomic responses to the Gq-protein-coupled receptor agonists endothelin-1, phenylephrine (a generic α1-adrenergic receptor agonist) and A61603 (α1A-adrenergic receptor selective). Phospho-ERK1/2 and phospho-RSKs appeared in cardiomyocyte nuclei within 2–3 min of stimulation (endothelin-1>A61603≈phenylephrine). All agonists increased nuclear RSK2, but only endothelin-1 increased the nuclear RSK1 content. PD184352 (inhibits ERK1/2 activation) and BI-D1870 (inhibits RSKs) were used to dissect the contribution of RSKs to the endothelin-1-responsive transcriptome. Of the 213 RNAs up-regulated after 1 h, 51% required RSKs for their up-regulation, whereas 29% required ERK1/2 but not RSKs. The transcriptomic response to phenylephrine overlapped with, but was not identical with, endothelin-1. As with endothelin-1, PD184352 inhibited the up-regulation of most phenylephrine-responsive transcripts, but the greater variation in the effects of BI-D1870 suggests that differential RSK signalling influences global gene expression. A61603 induced similar changes in RNA expression in cardiomyocytes as phenylephrine, indicating that the signal was mediated largely through α1A-adrenergic receptors. A61603 also increased expression of immediate early genes in perfused adult rat hearts and, as in cardiomyocytes, up-regulation of the majority of genes was inhibited by PD184352. PD184352 or BI-D1870 prevented the increased surface area induced by endothelin-1 in cardiomyocytes. Thus RSKs play a significant role in regulating cardiomyocyte gene expression and hypertrophy in response to Gq-protein-coupled receptor stimulation.

Endothelin-1 promotes hypertrophic remodelling of cardiac myocytes by activating sustained signalling and transcription downstream of endothelin type A receptors

“…As previously described, a low level of phosphorylated active ERK1/2 (pERK1/2) was detected under baseline conditions [18]. Consistent with previous reports, ET-1 application resulted in a rapid and substantial increase in pERK1/2 levels by 5 min [18], that remained elevated before returning to baseline after the 1 h time point (Fig. 7A).…”

“…The temporal profile of ERK1/2 activation following ET-1 stimulation of NRVMs described here and previously did not support this notion [15], [16], [18]. Consistent with previous studies, ERK1/2 phosphorylation is rapidly increased within minutes and then diminishes to baseline 1 h post-agonist stimulation [15].…”

Modelling Negative Feedback Networks for Activating Transcription Factor 3 Predicts a Dominant Role for miRNAs in Immediate Early Gene Regulation

“…Although BI-D1870 alone increase ERK1/2 phosphorylation in cardiomyocytes, this over a similar time and to a lesser degree than that induced by ET-1 and it does not affect the activation of ERK1/2 by ET-1 [9]. It is therefore unlikely that the enhancement of Atf3 mRNA expression induced by ET-1 seen in the presence of BI-D1870 is due to its effects on phosphorylation of ERK1/2 and suggests that a negative feedback system downstream of RSKs (or other kinases that may be inhibited by BI-D1870) moderates Atf3 mRNA expression.…”

Phenotypic Screening with Human iPS Cell–Derived Cardiomyocytes: HTS-Compatible Assays for Interrogating Cardiac Hypertrophy

“…Moreover, these results are quite comparable with other data from primary cell models for ET-1-induced hypertrophy. [22][23][24][25][26] To further establish the use of iPS cell-derived cardiomyocytes as a disease model for cardiac hypertrophy, we more rigorously explored additional markers as defined previously by other cell models. We compared induced (i.e., 10 nM ET-1) versus uninduced (i.e., no or 0 mM ET-1) samples via analysis by qPCR and DNA microarray.…”