Met acts through Abl to regulate p53 transcriptional outcomes and cell survival in the developing liver.

BACKGROUND & AIMS: Genetic studies indicate that distinct signaling modulators are each necessary but not individually sufficient for embryonic hepatocyte survival in vivo. Nevertheless, how signaling players are interconnected into functional circuits and how they coordinate the balance of cell survival and death in developing livers are still major unresolved issues. In the present study, we examined the modulation of the p53 pathway by HGF/Met in embryonic livers. METHODS: We combined pharmacological and genetic approaches to biochemically and functionally evaluate p53 pathway modulation in primary embryonic hepatocytes and in developing livers. RT-PCR arrays were applied to investigate the selectivity of p53 transcriptional response triggered by Met. RESULTS: Met recruits p53 to regulate the liver developmental program, by qualitatively modulating its transcriptional properties: turning on the Mdm2 survival gene, while keeping death and cell-cycle arrest genes Pmaip1 and p21 silent. We investigated the mechanism leading to p53 regulation by Met and found that Abl and p38MAPK are required for p53 phosphorylation on S(389), Mdm2 upregulation, and hepatocyte survival. Alteration of this signaling mechanism switches p53 properties, leading to p53-dependent cell death in embryonic livers. RT-PCR array studies affirmed the ability of the Met-Abl-p53 axis to modulate the expression of distinct genes that can be regulated by p53. CONCLUSIONS: A signaling circuit involving Abl and p38MAPK is required downstream of Met for the survival of embryonic hepatocytes, via qualitative regulation of the p53 transcriptional response, by switching its proapoptotic into survival properties.

DOCA-induced phosphorylation of glycogen synthase kinase 3beta.

Mineralocorticoid excess leads to cardiac fibrosis, a leading cause of morbidity and mortality. Cardiac hypertrophy and fibrosis are inhibited by the glycogen synthase kinase GSK3 which itself is a target of protein kinase B (PKB) and the serum and glucocorticoid inducible kinase SGK1. Phosphorylation of GSK3 by PKB or SGK1 inhibits GSK3 activity and should thus favour the development of cardiac hypertrophy and fibrosis. As SGK1 is transcriptionally upregulated by mineralocorticoids and has been recently shown to play an important role in the pathogenesis of mineralocorticoid-induced cardiac fibrosis, the present study explored whether mineralocorticoid excess had any effect on the phosphorylation status of the a and beta isoforms of GSK3. Western blotting using an antibody specific for the PKB/SGK1 consensus phosphorylation site in GSK3a/beta (serine 21 and 9 respectively) revealed an increase in GSK3a/beta phosphorylation in human embryonic kidney 293 (HEK293) cells overexpressing wild type SGK1, constitutively active SGK1, but not catalytically inactive SGK1. The effect of SGK1 was mimicked by PKB and SGK3. Furthermore, DOCA/high salt treatment of wild type mice induced a robust increase in cardiac GSK3beta phosphorylation and, to a much lesser extent, GSK3a phosphorylation. However, under this treatment GSK3beta phosphorylation was apparent even in mice lacking functional SGK1, indicating that the phosphorylation of GSK3beta was not exclusively mediated by this kinase. Despite similar cardiac GSK3beta phosphorylation cardiac fibrosis following DOCA/high salt treatment was significantly blunted in SGK1 knockout mice. In conclusion, mineralocorticoid excess leads to phosphorylation and thus inactivation of GSK3beta, an effect not only due to upregulation of SGK1 but as well due to activation of additional kinases. The inactivation of GSK3 may play a permissive role in the stimulation of cardiac fibrosis but may by itself not be sufficient to trigger cardiac fibrosis.