Contrasting effects on HIF-1 alpha regulation by disease-causing pVHL mutations correlate with patterns of tumourigenesis in von Hippel-Lindau disease

The von Hippel-Lindau tumour suppressor gene product (pVHL) associates with the elongin B and C and Cul2 proteins to form a ubiquitin-ligase complex ( VCBC), To date, the only VCBC substrates identified are the hypoxia-inducib le factor alpha subunits (HIF-1 alpha and HIF-2 alpha), However, pVHL is th ought to have multiple functions and the significance of HIF-1 alpha and HI F-2 alpha regulation for tumour suppressor activity has not been defined. V HL disease is characterized by distinct clinical subtypes, Thus haemangiobl astomas (HABs) and renal cell carcinoma (RCC) but not phaeochromocytoma (PH E) occur in type 1 VHL disease. Type 2 subtypes are characterized by PHE su sceptibility but differ with respect to additional tumours (type 2A, PHE+HA B but not RCC; type 2B, PHE+ HAB+RCC; type 2C, PHE only). We investigated i n detail the effect of 13 naturally occurring VHL mutations (11 missense), representing each phenotypic subclass, on HIF-alpha subunit regulation. Con sistent effects on pVHL function were observed for all mutations within eac h subclass. Mutations associated with the PHE-only phenotype (type 2C) prom oted HIF-alpha ubiquitylation in vitro and demonstrated wildtype binding pa tterns with pVHL interacting proteins, suggesting that loss of other pVHL f unctions are necessary for PHE susceptibility. Mutations causing HAB suscep tibility (types 1, 2A and 28) demonstrated variable effects on HIF-a subuni t and elongin binding, but all resulted in defective HIF-alpha( regulation and loss of p220 (fibronectin) binding. All RCC-associated mutations caused complete HIF-alpha dysregulation and loss of p220 (fibronectin) binding. O ur findings are consistent with impaired ability to degrade HIF-alpha subun it being required for HAB development and RCC susceptibility.