Regulation of STRA13 by the von Hippel-Lindau Tumor Suppressor Protein, Hypoxia, and the UBC9/Ubiquitin Proteasome Degradation Pathway

Abstract

In this study, we focus on different modes of regulation of STRA13, a human ortholog of the mouse basic helix-loop-helix transcriptional factor, previously identified by us as a new von Hippel-Lindau tumor suppressor gene (VHL) target. The gene was overexpressed in VHL-deficient cell lines and tumors, specifically clear cell renal carcinomas and hemangioblastomas. Introduction of wild type VHL transgene into clear cell renal carcinoma restored low level expression of STRA13. Overexpression was also detected in many common malignancies with an intact VHL gene, suggesting the existence of another, VHL-independent pathway of STRA13 regulation. Similar to many other von HippelLindau tumor-suppressor protein (pVHL) targets, the expression of STRA13 on the mRNA level was hypoxiasensitive, indicating oxygen-dependent regulation of the gene, presumably through the pVHL/hypoxia-inducible factor 1 (HIF-1) pathway. The yeast two-hybrid screening revealed interaction of the STRA13 protein with the human ubiquitin-conjugating enzyme (UBC9) protein, the specificity of which was confirmed in mammalian cells. By adding the proteasome inhibitor acetylleucinyl-leucinyl-norleucinal, we demonstrated that the 26 S proteasome pathway regulates the stability of pSTRA13. Co-expression of STRA13 and UBC9 led to an increase of the pSTRA13 ubiquitination and subsequent degradation. These data established that UBC9/STRA13 association in cells is of physiological importance, presenting direct proof of UBC9 involvement in the ubiquitindependent degradation of pSTRA13. Hypoxia treatment of mammalian cells transiently expressing STRA13 protein showed that stability of pSTRA13 is not affected by hypoxia or VHL. Thus, STRA13, a new pVHL target, is regulated in cells on multiple levels. We propose that STRA13 may play a critical role in carcinogenesis, since it is a potent transcriptional regulator, abundant in a variety of common tumors.Functional inactivation of the von Hippel-Lindau tumor-suppressor protein (pVHL) 1 causes a hereditary cancer syndrome characterized by the predisposition to develop tumors of kidney, central nervous system, retina, pancreas, and adrenal gland. Recent studies highlighted pVHL as a key regulator of cellular responses to oxygen deprivation. pVHL regulates activity of the hypoxia-inducible factor 1 (HIF-1), consisting of an HIF-1␣/HIF-1␤ heterodimer. Under normoxic conditions, the ␣ subunit interacts with the pVHL-elongin B-elongin C complex and is rapidly degraded (1). Hypoxia or pVHL deficiency stabilizes HIF-1␣, which results in activation of a set of genes including vascular endothelial growth factor (VEGF) (2, 3), glucose transporter GLUT-1 (3), glycolytic enzymes (reviewed in Ref. 4), transforming growth factor-␣ (5), and transforming growth factor-␤1 (6). We have recently expanded the list of hypoxia-sensitive pVHL targets. Using the RNA differential display technique, we identified and characterized two novel hypoxia-responsive VHL targets, carbonic anhydrases 9 and 12 (7). Here we describe a third gene down...

Mutations of the von Hippel–Lindau gene confer increased susceptibility to natural killer cells of clear-cell renal cell carcinoma

“…Our results are in accordance with previous reports showing that HIF-1a stabilization in VHL-deficient cell lines and clear-cell RCC results in the repression of endogenous signal transducer of activation of transcription (STAT)1 mediated by stimulated with retinoic acid (STRA)13 (Ivanova et al, 2001;Ivanov et al, 2007). STAT-1 regulates the constitutive expression of LMP2, a protein implicated in HLA-I antigen presentation (ChatterjeeKishore et al, 2000) likely responsible for the modulation of the HLA-I expression.…”

Kinetic analysis of genomewide gene expression reveals molecule circuitries that control T cell activation and Th1/2 differentiation

“…HIF-1 ␣ is induced (38) on T cell activation. Consistent with this pattern, its downstream target gene Stra13 (39,40) is also induced on T cell activation. Although no differences were observed in the expression of HIF-1␣ between Th1͞2 culture conditions, the downstream gene Stra13 was higher in Th1 than Th2.…”

DEC1 is a downstream target of TGF-β with sequence-specific transcriptional repressor activities

“…DEC1 was also shown to be induced by hypoxia (35,36), cAMP (37), and serum starvation (38), and was included in a list of 26 genes induced by TGF-␤ treatment of human mammary epithelial cells (39). DEC1 can act as a transcriptional repressor through interaction with the histone deacetylase HDAC1 or with the basal transcription factor TFIIB and can repress c-myc transcription and its own transcription through an autoregulatory loop (38).…”