Hepatocyte growth factor signalling stimulates hypoxia inducible factor-1 (HIF-1) activity in HepG2 hepatoma cells

Hepatocyte growth factor (HGF), a multifunctional cytokine of mesenchymal o rigin, activates the DNA binding of hypoxia inducible factor-1 (HIF-1) in t he HepG2 cell line: the activated complex contained the inducible a subunit . An increased expression of HIF-1 alpha (mRNA and nuclear protein levels) was observed. To investigate the molecular basis of the HIF-1 response unde r this non-hypoxic condition, we evaluated first the expression of putative target genes. We found a time-dependent increase in steady-state mRNA leve ls of heme oxygenase and urokinase plasminogen activator at 4 h, followed b y that of urokinase receptor at 10 h. The enhanced expression of these gene s might confer the invasive phenotype, since HGF is a proliferative and sca tter factor. Second, we examined some aspects of HIF-1 activity regulation in HGF-treated cells,with the following findings: (i) the activation of HIF -1 DNA binding was prevented by proteasome blockade, probably because stabi lization of the cytosolic oc-subunit protein level is not sufficient to gen erate a functional form: also under these conditions nuclear protein level of HIF-1 alpha did not increase; (ii) N-acetyleysteine, a free radical scav enger, strongly decreased HIF-1 activation suggesting a role of reactive ox ygen species in this process; (iii) the thiol reducing agent dithiothreitol was ineffective. Third, consistent with these data, N-acetylcysteine reduc ed the stimulatory effect of HGF on stress kinase activities, while p42/44 mitogen activated kinase (MAPK) was unmodified, suggesting an involvement o f c-jun-N-terminal kinase (JNK) and p38 MAPK in HIF-1 activation. Finally, LY 294002 induced the blockade of phosphatidylinositol 3-kinase (PI3K), one of the principal transducers of HGF/Met receptor signalling, prevented the enhancement of HIF-1 DNA binding and JNK activity, but the inhibition of p 42/44 MAPK phosphorylation with PD 98059 was ineffective. In conclusion, we suggest that HGF triggers a signal transduction cascade involving PI3K and ultimately activates HIF-1.