The Effects of Prenylation Deficiency and Statin Treatment in Cardiomyocytes

Abstract

Isoprenylation is an important lipid modification for the function and localisation of CAAX-proteins. The CAAX-protein protein X is involved in actin reorganisation, agonist-induced increased cell size and change in protein expression and is believed to be a key mediator in cardiomyopathies, such as cardiac hypertrophy. It is widely believed that Statins reduce isoprenylation of proteins in the Rho-family, including protein X, and thereby reduce their activity, leading to a cardioprotective effect. However, unpublished studies performed in the group of Martin Bergo have indicated that prenylation deficiency increases levels of the active form of protein X and that inhibition of prenylation leads to cardiomyopathy. It was therefore hypothesised that Statins increase protein X activity which leads to reduced cardiomyocyte function. In this study, increased levels of protein X was evaluated by inducing prenylation deficiency in cardiomyocytes using Simvastatin, Rosuvastatin, Atorvastatin and GGTi. GTP-assay followed by western blot was conducted to see if GTP-bound protein levels were increased. The impact of increased activity in cardiomyocytes was evaluated by investigating levels of downstream targets by performing western blot and RT-qPCR. Functional assays in the form of staining of the actin cytoskeleton was also conducted. It was verified that active levels of the protein were increased in prenylation deficient HL-1 cardiomyocytes treated with Simvastatin, Rosuvastatin and GGTi. Downstream protein expression indicated that the protein of interest is signalling properly downstream, i.e. is activated and functioning. Abnormal behaviour in the cells treated with Statins was rescued by the addition of GGPP indicating that geranylgeranylation deficiency was the cause.