Role of MEK/ERK signalling in protein synthesis and cardiac hypertrophy

Abstract

The Gq protein-coupled receptor agonist phenylephrine (PE) induces cardiac hypertrophy and stimulates protein synthesis in adult rat ventricular cardiomyocytes (ARVC). Here we show that the adenoviral expression of MAP kinase phosphatase 3 (Adv-MKP3) inhibited the induction of ERK1/2 phosphorylation by PE and completely ablated the activation of protein synthesis. Moreover, MKP3 expression also inhibited the PE-induced phosphorylation of two downstream targets of the mammalian target of rapamycin (mTOR): eIF4E binding protein 1 (4E-BP1) and p70 ribosomal S6 kinase 1 (p70S6K1). This clearly implicates ERK1/2 signalling as an essential component in the control of regulatory translation factors and the activation of protein synthesis by PE. The tuberous sclerosis complex (TSC) is a tumour suppressor complex composed of two proteins, termed hamartin (TSC1) and tuberin (TSC2). TSC1/2, which lies upstream of mTOR and negatively regulates its activity, has been implicated in the control of cell size. Consequently, TSC1/2 are key candidates for involvement in the induction of protein synthesis by hypertrophic agents such as PE; therefore the effects of PE treatment of ARVC on TSC2 protein were also analysed. PE induced phosphorylation of TSC2 and was blocked by inhibition of MEK. Phosphorylation was detected on Ser/Thr residues that encompass the RXRXX(S/T) motif, which may be a target for p90RSK. These data strongly suggest a link between the MAPK pathway and TSC1/2. Previously published data shows that MEK inhibition completely ablated PE-induced protein synthesis; whereas rapamycin pre-treatment gave only a partial inhibition. This suggests that an element of PE-activated protein synthesis is MEK-dependent but independent of mTOR. A potential aspect of this is eukaryotic initiation factor 4E (eIF4E), which promotes 5′-cap-mediated translation. eIF4E is phosphorylated by MAP kinase integrating kinases 1/2 (Mnk1/2), which are downstream targets of ERK1/2. Pre-treatment of ARVC with the Mnk-inhibitor CGP57380 partially attenuated the activation of protein synthesis by PE, but in combination with rapamycin almost completely blocked PE-induced protein synthesis. CGP57380 pre-treatment also reduced the level of eIF4E phosphorylation. These data suggest that the activation of protein synthesis by PE may involve two outputs that depend upon MEK/ERK signalling i.e. a rapamycin-sensitive mTOR pathway (p70S6K1/2 and 4E-BP1), through TSC1/2, and an Mnk-dependent step, possibly involving phosphorylation of eIF4E.