A peptide from a ras effector-domain blocks ras-dependent cardiac hypertrophy in myocytes.

PNC-2 is a peptide corresponding to an effector domain (residues 96-110) of ras-p21 that strongly and specifically blocks mitogenic signal transduction by oncogenic but not activated, normally-expressed wild-type ras-p21 protein. Since myocardial hypertrophy can be induced both by oncogenic and overexpressed wild-type ras-p21, we investigated whether PNC-2 can block norepinephrine (NE)-induced, ras-dependent myocardial hypertrophy in cardiac myocytes. Since PNC-2 blocks oncogenic ras-p21-induced activation of JNK and ERK, we further determined whether this peptide blocks activation of these kinases in NE-treated myocytes. Using cultured neonatal rat ventricular myocytes (NRVM), we found that NE alone significantly increased NRVM surface area, (3)H-leucine uptake, protein/DNA ratio, and atrial nartiuretic factor (ANF) mRNA levels in these cells. However, pretreatment of the NRVM with PNC-2 linked on its carboxyl terminal end to a transmembrane-penetrating leader sequence (PNC-2-leader) resulted in strong inhibition of NE-mediated cell growth and (3)H-leucine uptake and in significantly lower protein/DNA ratios. Induction of ANF mRNA levels was likewise inhibited by PNC-2-leader. In contrast, no inhibition of any of these NE-induced events was observed with a negative control peptide, X13-leader. Western blot analysis showed that JNK and ERK1/2 activity, but not p38 activity, was increased in NRVM within 5 min of exposure to NE (2 microM). Pretreatment with PNC-2-leader decreased ERK1/2 and JNK activity to basal levels. We conclude that a synthetic peptide designed to block oncogenic ras can also counter the effects of NE-induced hypertrophy associated with overexpression of ras p21 by blocking JNK/ jun and ERK activation. PNC-2 may provide a prototype for novel therapy in cardiac conditions associated with activation of NE.