title: Stabilization of the prostate-specific tumor suppressor NKX3.1 by the oncogenic protein kinase Pim-1 in prostate cancer cells.
abstract: Loss of NKX3.1 is an early and consistent event in prostate cancer and is associated with increased proliferation of prostate epithelial cells and poor prognosis. NKX3.1 stability is regulated post-translationally through phosphorylation at multiple sites by several protein kinases. Here, we report the paradoxical stabilization of the prostate-specific tumor suppressor NKX3.1 by the oncogenic protein kinase Pim-1 in prostate cancer cells. Pharmacologic Pim-1 inhibition using the small molecule inhibitor CX-6258 decreased steady state levels and half-life of NKX3.1 protein but mRNA was not affected. This effect was reversed by inhibition of the 26S-proteasome, demonstrating that Pim-1 protects NKX3.1 from proteasome-mediated degradation. Mass spectrometric analyses revealed Thr89, Ser185, Ser186, Ser195, and Ser196 as Pim-1 phospho-acceptor sites on NKX3.1. Through mutational analysis, we determined that NKX3.1 phosphorylation at Ser185, Ser186, and within the N-terminal PEST domain is essential for Pim-1-mediated stabilization. Further, we also identified Lys182 as a critical residue for NKX3.1 stabilization by Pim-1. Pim-1-mediated NKX3.1 stabilization may be important in maintaining normal cellular homeostasis in normal prostate epithelial cells, and may maintain basal NKX3.1 protein levels in prostate cancer cells.
authors.affiliation: Department of Biological Sciences, University of Maryland Baltimore County, Baltimore, MD, USA.
authors.forename: Achuth
authors.initials: A
authors.lastname: Padmanabhan
authors.forename: Eliza B
authors.initials: EB
authors.lastname: Gosc
authors.forename: Charles J
authors.initials: CJ
authors.lastname: Bieberich
mesh_terms: D001381:Azepines / Q000494:pharmacology; D045744:Cell Line, Tumor; D004252:DNA Mutational Analysis; D006207:Half-Life; D018398:Homeodomain Proteins / Q000378:metabolism*; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D008239:Lysine / Q000378:metabolism; D008297:Male; D008954:Models, Biological; D050505:Mutant Proteins / Q000378:metabolism; D009928:Organ Specificity / Q000187:drug effects; D010766:Phosphorylation / Q000187:drug effects; D011467:Prostate / Q000201:enzymology* / Q000473:pathology*; D011471:Prostatic Neoplasms / Q000201:enzymology* / Q000473:pathology*; D014176:Protein Biosynthesis / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D055550:Protein Stability / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D051573:Proto-Oncogene Proteins c-pim-1 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013379:Substrate Specificity / Q000187:drug effects; D014157:Transcription Factors / Q000378:metabolism*; D025521:Tumor Suppressor Proteins / Q000378:metabolism*
chemical_list: D001381:Azepines; C000607921:CX-6258; D018398:Homeodomain Proteins; D007211:Indoles; D050505:Mutant Proteins; C491178:NKX3-1 protein, human; D047428:Protein Kinase Inhibitors; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C496634:PIM1 protein, human; D051573:Proto-Oncogene Proteins c-pim-1; D008239:Lysine