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VHL Type 2B gene mutation moderates HIF dosage in vitro and in vivo.

Lee CM, Hickey MM, Sanford CA, McGuire CG, Cowey CL, Simon MC, Rathmell WK - Oncogene (2009)

Bottom Line: Von Hippel-Lindau (VHL) disease is caused by germline mutations in the VHL tumor suppressor gene, with Type 2B missense VHL mutations predisposing to renal cell carcinoma, hemangioblastoma and pheochromocytoma.Type 2B mutant pVHL is predicted to be defective in hypoxia inducible factor (HIF)-alpha regulation.Our experiments support a model in which the representative Type 2B R167Q mutant pVhl produces a unique profile of HIF dysregulation, thereby promoting tissue-specific effects on cell growth, development and tumor predisposition.

View Article: PubMed Central - PubMed

Affiliation: Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599-7295, USA.

ABSTRACT
Von Hippel-Lindau (VHL) disease is caused by germline mutations in the VHL tumor suppressor gene, with Type 2B missense VHL mutations predisposing to renal cell carcinoma, hemangioblastoma and pheochromocytoma. Type 2B mutant pVHL is predicted to be defective in hypoxia inducible factor (HIF)-alpha regulation. Murine embryonic stem (ES) cells in which the endogenous wild-type Vhl gene was replaced with the representative Type 2B VHL hotspot mutation R167Q (Vhl(2B/2B)) displayed preserved physiological regulation of both HIF factors with slightly greater normoxic dysregulation of HIF-2alpha. Differentiated Vhl(2B/2B)-derived teratomas overexpressed joint HIF targets Vegf and EglN3 but not the HIF-1alpha-specific target Pfk1. Vhl(2B/2B) teratomas additionally displayed a growth advantage over Vhl(-/-)-derived teratomas, suggestive of a tight connection between perturbations in the degree and ratio of HIF-1alpha and HIF-2alpha stabilization and cell growth. Vhl(2B/2B) mice displayed mid-gestational embryonic lethality, whereas adult Vhl(2B/+) mice exhibited susceptibility to carcinogen-promoted renal neoplasia compared with wild-type littermates at 12 months. Our experiments support a model in which the representative Type 2B R167Q mutant pVhl produces a unique profile of HIF dysregulation, thereby promoting tissue-specific effects on cell growth, development and tumor predisposition.

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Exploration of hyper-angiogenic phenotype of homozygous Type 2B Vhl teratomas. Quantitative RT-PCR for transcriptional activation of angiogenesis-related genes Cdh5, Eng, Kdr, Nrp1, and Vegfc in a representative set of teratomas relative to J1. Cycle thresholds were corrected with 18S ribosomal RNA. Error bars indicate standard deviation. *p<0.05. **p<0.001.
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Figure 4: Exploration of hyper-angiogenic phenotype of homozygous Type 2B Vhl teratomas. Quantitative RT-PCR for transcriptional activation of angiogenesis-related genes Cdh5, Eng, Kdr, Nrp1, and Vegfc in a representative set of teratomas relative to J1. Cycle thresholds were corrected with 18S ribosomal RNA. Error bars indicate standard deviation. *p<0.05. **p<0.001.

Mentions: To explore the differences in vascular proclivities between Vhl-/- and Vhl2B/2B teratomas, we performed a screen for candidate angiogenesis-related genes (Supplemental Table 1 and Figure 4). Four novel angiogenesis-related candidate genes, in addition to Vegfa, were uncovered in the candidate screen and validated by qRT-PCR with independent primers : vascular endothelial (VE)-cadherin (Cdh5), the TgfβR endothelial co-receptor endoglin (Eng), Vegf receptor 2 (Kdr), and the Vegfr2 co-receptor neuropilin-1 (Nrp1). All four are considered direct HIF targets (Brusselmans et al., 2005; Elvert et al., 2003; Le Bras et al., 2007; Sanchez-Elsner et al., 2002). While both the Vhl-/- and Vhl2B/2B teratomas significantly over-expressed Cdh5, Eng, and Kdr relative to the J1 teratoma, the Vhl2B/2B teratoma additionally significantly (p<0.001) over-expressed Nrp1 relative to both J1 and Vhl-/- (Figure 4).


VHL Type 2B gene mutation moderates HIF dosage in vitro and in vivo.

Lee CM, Hickey MM, Sanford CA, McGuire CG, Cowey CL, Simon MC, Rathmell WK - Oncogene (2009)

Exploration of hyper-angiogenic phenotype of homozygous Type 2B Vhl teratomas. Quantitative RT-PCR for transcriptional activation of angiogenesis-related genes Cdh5, Eng, Kdr, Nrp1, and Vegfc in a representative set of teratomas relative to J1. Cycle thresholds were corrected with 18S ribosomal RNA. Error bars indicate standard deviation. *p<0.05. **p<0.001.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2667565&req=5

Figure 4: Exploration of hyper-angiogenic phenotype of homozygous Type 2B Vhl teratomas. Quantitative RT-PCR for transcriptional activation of angiogenesis-related genes Cdh5, Eng, Kdr, Nrp1, and Vegfc in a representative set of teratomas relative to J1. Cycle thresholds were corrected with 18S ribosomal RNA. Error bars indicate standard deviation. *p<0.05. **p<0.001.
Mentions: To explore the differences in vascular proclivities between Vhl-/- and Vhl2B/2B teratomas, we performed a screen for candidate angiogenesis-related genes (Supplemental Table 1 and Figure 4). Four novel angiogenesis-related candidate genes, in addition to Vegfa, were uncovered in the candidate screen and validated by qRT-PCR with independent primers : vascular endothelial (VE)-cadherin (Cdh5), the TgfβR endothelial co-receptor endoglin (Eng), Vegf receptor 2 (Kdr), and the Vegfr2 co-receptor neuropilin-1 (Nrp1). All four are considered direct HIF targets (Brusselmans et al., 2005; Elvert et al., 2003; Le Bras et al., 2007; Sanchez-Elsner et al., 2002). While both the Vhl-/- and Vhl2B/2B teratomas significantly over-expressed Cdh5, Eng, and Kdr relative to the J1 teratoma, the Vhl2B/2B teratoma additionally significantly (p<0.001) over-expressed Nrp1 relative to both J1 and Vhl-/- (Figure 4).

Bottom Line: Von Hippel-Lindau (VHL) disease is caused by germline mutations in the VHL tumor suppressor gene, with Type 2B missense VHL mutations predisposing to renal cell carcinoma, hemangioblastoma and pheochromocytoma.Type 2B mutant pVHL is predicted to be defective in hypoxia inducible factor (HIF)-alpha regulation.Our experiments support a model in which the representative Type 2B R167Q mutant pVhl produces a unique profile of HIF dysregulation, thereby promoting tissue-specific effects on cell growth, development and tumor predisposition.

View Article: PubMed Central - PubMed

Affiliation: Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599-7295, USA.

ABSTRACT
Von Hippel-Lindau (VHL) disease is caused by germline mutations in the VHL tumor suppressor gene, with Type 2B missense VHL mutations predisposing to renal cell carcinoma, hemangioblastoma and pheochromocytoma. Type 2B mutant pVHL is predicted to be defective in hypoxia inducible factor (HIF)-alpha regulation. Murine embryonic stem (ES) cells in which the endogenous wild-type Vhl gene was replaced with the representative Type 2B VHL hotspot mutation R167Q (Vhl(2B/2B)) displayed preserved physiological regulation of both HIF factors with slightly greater normoxic dysregulation of HIF-2alpha. Differentiated Vhl(2B/2B)-derived teratomas overexpressed joint HIF targets Vegf and EglN3 but not the HIF-1alpha-specific target Pfk1. Vhl(2B/2B) teratomas additionally displayed a growth advantage over Vhl(-/-)-derived teratomas, suggestive of a tight connection between perturbations in the degree and ratio of HIF-1alpha and HIF-2alpha stabilization and cell growth. Vhl(2B/2B) mice displayed mid-gestational embryonic lethality, whereas adult Vhl(2B/+) mice exhibited susceptibility to carcinogen-promoted renal neoplasia compared with wild-type littermates at 12 months. Our experiments support a model in which the representative Type 2B R167Q mutant pVhl produces a unique profile of HIF dysregulation, thereby promoting tissue-specific effects on cell growth, development and tumor predisposition.

Show MeSH
Related in: MedlinePlus