HIF‐1α: A key survival factor for serum‐deprived prostate cancer cells

Abstract

Hypoxia-inducible factor-1 alpha (HIF-1 alpha) is commonly overexpressed in prostate cancer (PCa) cells. As PCa cells are known to survive serum deprivation, we investigated the effect of prolonged serum deprivation on HIF-1 alpha expression, and the function of HIF-1 alpha in regulating the survival of normoxic serum-deprived PCa cells. HIF-1 alpha protein was assessed by immunoblots. Cell viability and proliferation were assessed by trypan blue assay and flow cytometric analysis. Transcriptional activity was assessed by luciferase reporter assay and RT-PCR. HIF-1 alpha expression was suppressed with siRNA. Activities of HIF-1 alpha-target genes were inhibited with neutralizing antibody. Prolonged serum deprivation is a potent inducer of HIF-1 alpha in PC-3 and LNCaP PCa cells, despite normal oxygen conditions. In contrast, cells grown in the presence of serum did not show HIF-1 alpha protein accumulation. Moreover, HIF-1 alpha protein increase during serum deprivation correlated with increased cell survival, while suppression of HIF-1 alpha expression significantly decreased PCa cell viability. Our results further demonstrate that HIF-1 alpha protein increase is due to increased HIF-1 alpha protein synthesis. First, there was no significant increase in HIF-1 alpha mRNA. Secondly, cycloheximide, a protein synthesis inhibitor, prevented HIF-1 alpha protein increase in serum-deprived PCa cells. Moreover, the expression of HIF-1 alpha-target genes, VEGF and IGF-2, was concomitantly increased in serum-deprived PCa cells, while suppression of HIF-1 alpha expression significantly inhibited their induction. Furthermore, inhibition of IGF-2 activity resulted in a significant decline in PCa cell survival. PCa cells counteract the stress of prolonged serum deprivation by upregulating HIF-1 alpha protein which increases IGF-2 expression to promote cell survival.