PKM2 and HIF-1α regulation in prostate cancer cell lines

Abstract

Prostate cancer (PCA) is one of the most common cancer types in men, with cancer progression being linked to hypoxia and the induction of hypoxia-inducible factor (HIF).We investigated the expression of pyruvate kinase M2 (PKM2), its regulation by HIF isoforms 1α and 2α, and its role in HIF stabilization. We additionally examined cell survival in the prostate cancer cell lines PC3 and LNCaP under severe hypoxic (0.1% O2) and normoxic (20% O2) conditions. qRT-PCR showed higher up-regulation of PKM2 mRNA expression in LNCaP cells than in PC3 cells, while western blotting showed that PKM2 protein levels were up-regulated only in LNCaP cells. Inhibition of HIF-1α and HIF-2α by small interfering RNA (si-RNA) demonstrated HIF-1α dependent up-regulation of PKM2 at the mRNA and protein levels in LNCaP cells. PKM2 inhibition by si-RNA significantly decreased hypoxia-response element (HRE) activation in a gene reporter assay and down-regulated HIF-1α target vascular endothelial growth factor (VEGF) mRNA expression in PC3 cells, whereas HIF-1α protein levels were not significantly reduced. Additionally, PKM2 inhibition significantly reduced clonogenic survival in both cell lines in a colony formation assay. Prolyl hydroxylase 3 (PHD3) mRNA expression was up-regulated in both cell lines. It has been shown that PKM2 expression is regulated by HIF-1α and that PKM2 favors HIF-1α transactivation under mild (1% O2) but not severe (0.1% O2) hypoxic conditions, and some of our findings are consistent with these previous results. However, this mechanism was not fully observed in our studied cell lines, as PKM2 regulation and HIF-1α stabilization at the transactivation level occurred under severe hypoxic conditions. This discrepancy suggests that tumor tissue origin and cell type influence this model. Our findings expand the current knowledge of the mechanisms of PCA regulation, and would be important in developing novel therapeutic strategies.

Pyruvate Kinase M2 and Cancer: The Role of PKM2 in Promoting Tumorigenesis

“…They showed that that PKM2 mRNA was expressed at a higher level in the PC3 cell line than in LNCaP cells under normoxic conditions. Severe hypoxia significantly increased PKM2 mRNA expression in both cell lines (41). This suggested that, under hypoxic conditions, the expression of PKM2 is further promoted by HIF-1α activation.…”

“…Hasan et al (41) demonstrated the expression of PKM2 in normoxic (20% O 2 ) and hypoxic (0.1% O 2 ) conditions in two prostate cancer cell lines, PC3 and LNCaP. They showed that that PKM2 mRNA was expressed at a higher level in the PC3 cell line than in LNCaP cells under normoxic conditions.…”

Shmt2: A Stat3 Signaling New Player in Prostate Cancer Energy Metabolism

“…Also PKM2 shows a different expression level in the two cell lines, more abundant in DU145 cells when compared to LNCaP ones (Figure 4B,C). This difference may be correlated to the higher level of HIF-1α in DU145 cells and to the potential cooperative role of the two proteins in controlling cell metabolism [34,35]. Western blotting analysis, carried out on total and nuclear extracts, highlights a different localization of PKM2 between the two cell lines; in fact, it is mainly localized in the cytosol of LNCaP cells, whereas it is primarily present in the nuclear compartment in DU145 cells (Figure 4D).…”

TERF1 downregulation promotes the migration and invasion of the PC3 prostate cancer cell line as a target of miR‑155

“…Hypoxia inducible factor 1α (HIF-1α) induces the expression of VEGF, which has been reported to promote neovascularization in PCa (41). This pathological process has been suggested to promote the proliferation and metastasis of PCa targeted by miRNAs (42). Previous studies have illustrated that the overexpression of HIF-1α increased the risk of CRPC and the metastasis of PCa.…”