Journal Article

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

Lorenza Tacchini, Paola Dansi, Emanuela Matteucci and Maria Alfonsina Desiderio

in Carcinogenesis

Volume 22, issue 9, pages 1363-1371
Published in print September 2001 | ISSN: 0143-3334
Published online September 2001 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/22.9.1363
Hepatocyte growth factor signalling stimulates hypoxia inducible factor-1 (HIF-1) activity in HepG2 hepatoma cells

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Hepatocyte growth factor (HGF), a multifunctional cytokine of mesenchymal origin, activates the DNA binding of hypoxia inducible factor-1 (HIF-1) in the HepG2 cell line: the activated complex contained the inducible α subunit. An increased expression of HIF-1α (mRNA and nuclear protein levels) was observed. To investigate the molecular basis of the HIF-1 response under this non-hypoxic condition, we evaluated first the expression of putative target genes. We found a time-dependent increase in steady-state mRNA levels of heme oxygenase and urokinase plasminogen activator at 4 h, followed by that of urokinase receptor at 10 h. The enhanced expression of these genes might confer the invasive phenotype, since HGF is a proliferative and scatter 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 stabilization of the cytosolic α-subunit protein level is not sufficient to generate a functional form: also under these conditions nuclear protein level of HIF-1α did not increase; (ii) N-acetylcysteine, a free radical scavenger, strongly decreased HIF-1 activation suggesting a role of reactive oxygen species in this process; (iii) the thiol reducing agent dithiothreitol was ineffective. Third, consistent with these data, N-acetylcysteine reduced the stimulatory effect of HGF on stress kinase activities, while p42/44 mitogen activated kinase (MAPK) was unmodified, suggesting an involvement of 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 p42/44 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.

Keywords: GSH, reduced glutathione; HGF, hepatocyte growth factor; HIF-1, hypoxia-inducible factor; Hox, heme oxygenase; HRE, hypoxia responsive element; JNK/SAPK, c-Jun-N-terminal kinase/stress activated protein kinase; MAPK, mitogen activated protein kinase; Oct-1, octamer-1; PI3K, phosphatidylinositol 3-kinase; rhHGF, recombinant human HGF; ROS, reactive oxygen species; uPA, urokinase plasminogen activator; uPAR, urokinase receptor.

Journal Article.  8041 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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