Hypoxia on the Expression of Hepatoma Upregulated Protein in Prostate Cancer Cells

Abstract

Hepatoma upregulated protein (HURP) is a multifunctional protein with clinical promise. This protein has been demonstrated to be a predictive marker for the outcome in high-risk prostate cancer (PCa) patients, besides being a resistance factor in PCa. Although changes in oxygen tension (pO2) are associated with PCa aggressiveness, the role of hypoxia in the regulation of tumor progression genes such as HURP has not yet been described. We hypothesized that pO2 alteration is involved in the regulation of HURP expression in PCa cells. In the present study, PCa cells were incubated at 2% O2 (hypoxia) and 20% O2 (normoxia) conditions. Hypoxia reduced cell growth rate of PCa cells, when compared to the growth rate of cells cultured under normoxia (p < 0.05). The decrease in cell viability was accompanied by fivefold (p < 0.05) elevated rate of vascular endothelial growth factor (VEGF) release. The expression of VEGF and the hypoxia-inducible metabolic enzyme carbonic anhydrase 9 were elevated maximally nearly 61-fold and 200-fold, respectively (p < 0.05). Noted in two cell lines (LNCaP and C4-2B) and independent of the oxygen levels, HURP expression assessed at both mRNA and protein levels was reduced. However, the decrease was more pronounced in cells cultured under hypoxia (p < 0.05). Interestingly, the analysis of patients’ specimens by Western blot revealed a marked increase of HURP protein (fivefold), when compared to control (cystoprostatectomy) tissue (p < 0.05). Immunohistochemistry analysis showed an increase in the immunostaining intensity of HURP and the hypoxia-sensitive molecules, hypoxia-inducible factor 1-alpha (HIF-1α), VEGF, and heat-shock protein 60 (HSP60) in association with tumor grade. The data also suggested a redistribution of subcellular localization for HURP and HIF-1α from the nucleus to the cytoplasmic compartment in relation to increasing tumor grade. Analysis of HURP Promoter for HIF-1-binding sites revealed presence of four putative HIF binding sites on the promoter of DLGAP5/HURP gene in the non-translated region upstream from the start codon, suggesting association between HIF-1α and the regulation of HURP protein. Taken together, our findings suggest a modulatory role of hypoxia on the expression of HURP. Additionally our results provide basis for utilization of tumor-associated molecules as predictors of aggressive PCa.

Elevated mRNA expression levels of DLGAP5 are associated with poor prognosis in breast cancer

“…DLGAP5, a microtubule-associated protein, is phosphorylated by Aurora kinase A (AURKA) (43). In addition, DLGAP5 upregulation is associated with poor prognosis of patients with colorectal cancer (44), lung cancer (10), bladder cancer (11) and prostate cancer (12), indicating that DLGAP5 plays an important role in cancer prognosis. Branchi et al (44) reported that DLGAP5 was associated with the nodal status, and high DLGAP5 expression levels were associated with a less favorable overall survival rate in distinct molecular colorectal cancer subtypes.…”

“…Furthermore, targeting mitosis-associated genes, including DLGAP5, TPX2 and RAN, has been reported to induce apoptosis of the BRG1 mutant NSCLC cells, both in vitro and in vivo (47). Espinoza et al (12) demonstrated that DLGAP5 was a predictive biomarker for the prognosis of high-risk prostate cancer in addition to functioning as a resistance factor. Furthermore, hypoxia-inducible factor 1-alpha (HIF-1α) binding sites were revealed on the promoter of DLGAP5, indicating that hypoxia plays a modulatory role on DLGAP5 expression (11).…”

“…In cytological experiments, the knockdown of DLGAP5 caused inhibition of proliferation, migration and invasion of NSCLC cells (10). Previous studies have also shown that the expression levels of DLGAP5 are upregulated in bladder (11), prostate (12) and liver cancer (13), as well as leukemia (14) were associated with poor prognosis. In the present study, DLGAP5 was identified to be a hub gene in GEO and Gene Expression-Based Outcome for BC Online (GOBO) databases.…”

Pan-cancer Analysis Combining with Experiments Indicates the Sensitivity of Renal Carcinogenesis to DLGAP5 Expression

“…Second, apart from promoting cancer cells proliferation, what other functions does it have? Indeed, based on bioinformatics analysis, it has been found that DLGAP5 involved in the occurrence, development and prognosis of malignant tumors [18,24], and may serve as potential molecular biomarkers for a set of carcinomas, including glioblastoma [24], prostate cancer [16,17,25]. Interestingly, DLGAP5 protein has been found to be one of top ten oxygen-sensitive proteins in prostate cancer [16,26], and is proposed as an independent prognostic biomarker in this tumor [16].…”

“…Interestingly, DLGAP5 protein has been found to be one of top ten oxygen-sensitive proteins in prostate cancer [16,26], and is proposed as an independent prognostic biomarker in this tumor [16]. By using the published date, it has been found that several putative HIF binding sites located at promoter region of DLGAP5, most of which are highly methylated region [16,25]. In addition, the inhibitory effect of DLGAP5 knockout on prostate cancer is related to androgen receptor [17].…”