A HIF-1alpha-dependent autocrine feedback loop promotes survival of serum-deprived prostate cancer cells.
BACKGROUND
We previously reported that normoxic, serum-deprived prostate cancer (PCa) cells upregulate hypoxia-inducible factor-1alpha (HIF-1alpha) protein, which promotes survival during serum deprivation via insulin-like growth factor-2 (IGF-2) upregulation. This study investigated the molecular mechanism of autocrine regulation of HIF-1alpha, IGF-2 and cell survival in serum-deprived PC-3 and LNCaP PCa cells.


METHODS
Cell viability was assessed by trypan blue assay. PI3K activity was inhibited with LY294002, and PTEN overexpression. mRNA was assessed by RT-PCR, and IGF-2 protein by ELISA. Activated insulin-like growth factor-I receptor (IGF-IR) was detected by probing immunoprecipitated IGF-IR for phospho-tyrosine. IGF-IR activity was inhibited with IGF-2 neutralizing antibody and IGF-IR-specific siRNA. HIF-1alpha, phospho-Akt, total-Akt and IGF-IR protein was assessed by immunoblots. HIF-1alpha was suppressed with siRNA.


RESULTS
We detected a time-dependent increase in Akt activation during serum deprivation, and inhibition of Akt activation attenuated the serum deprivation-mediated increase in HIF-1alpha and cell survival. Importantly, IGF-2 secretion significantly increased during serum deprivation, and was accompanied by increased activation of its receptor, IGF-IR. Additionally, inhibition of IGF-2 activity markedly attenuated the serum deprivation-mediated increase in IGF-IR and Akt activation, HIF-1alpha expression, and also its own transcription, suggesting autocrine regulation of HIF-1alpha expression via IGF-2. Cross-talk between IGF-2/ IGF-IR system and PI3K-Akt pathway was further demonstrated by findings wherein IGF-IR suppression inhibited Akt activation, and IGF-IR activation was inhibited following PI3K inhibition. Furthermore, HIF-1alpha suppression attenuated the serum deprivation-mediated increase in Akt and IGF-IR activation.


CONCLUSIONS
Collectively, our study demonstrates existence of a pro-survival HIF-1alpha-dependent autocrine feedback loop in normoxic, serum-deprived PCa cells.
Thomas
Rusha
R
Department of Molecular Biology and Immunology, University of North Texas Health Science Center, Fort Worth, Texas 76107, USA.
Kim
Myoung H
MH
D019898:Autocrine Communication / Q000502:physiology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000502:physiology; D016895:Culture Media, Serum-Free / Q000494:pharmacology*; D025461:Feedback, Physiological / Q000502:physiology; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics / Q000378:metabolism*; D007335:Insulin-Like Growth Factor II / Q000378:metabolism; D008297:Male; D051059:PTEN Phosphohydrolase / Q000235:genetics; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011471:Prostatic Neoplasms / Q000378:metabolism* / Q000473:pathology*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D017526:Receptor, IGF Type 1 / Q000378:metabolism; D014162:Transfection; D015854:Up-Regulation / Q000502:physiology
D016895:Culture Media, Serum-Free; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007335:Insulin-Like Growth Factor II; D019869:Phosphatidylinositol 3-Kinases; D017526:Receptor, IGF Type 1; D051057:Proto-Oncogene Proteins c-akt; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human