Cellular proteins that bind the von Hippel-Lindau disease gene product: mapping of binding domains and the effect of missense mutations.
The von Hippel-Lindau disease (VHL) gene is a novel tumor suppressor gene that plays a role in the pathogenesis of renal cell carcinomas and hemangioblastomas of the central nervous system. To begin an evaluation of the biological functions of the VHL gene product (pVHL), we prepared bacterial fusion protein between glutathione S-transferase and wild-type or mutant pVHLs. The fusion proteins were used to identify cellular proteins that bind to pVHL in vitro. Monkey kidney cells transfected with wild-type or mutant VHL cDNAs were used to identify cellular proteins that bind to pVHL in vivo. Wild-type pVHL consistently bound two cellular proteins with apparent molecular masses of 10 and 14 kilodaltons that were designated p10 and p14, respectively. Mapping studies with a panel of VHL deletion mutant proteins demonstrated that p10 and p14 bound to a 32-amino acid peptide located in the carboxy terminal portion of pVHL. Missense mutation located within this 32-amino acid peptide abrogated the ability of the VHL protein to bind p10 and p14. Of 67 VHL families with identified germline mutations, 42 families had mutations predicted to affect the p10/p14-binding region. Maintenance of the integrity of the p10/p14-binding region appears to be essential for cellular growth regulation by pVHL.
Kishida
T
T
Laboratory of Immunobiology, National Cancer Institute-Frederick Cancer Research Facility and Development Center, Maryland 21702-1201, USA.
Stackhouse
T M
TM
Chen
F
F
Lerman
M I
MI
Zbar
B
B
D000818:Animals; D001665:Binding Sites; D002522:Chlorocebus aethiops; D016147:Genes, Tumor Suppressor; D006801:Humans; D066298:In Vitro Techniques; D008025:Ligases; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed; D009363:Neoplasm Proteins / Q000737:chemistry / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D017354:Point Mutation; D011485:Protein Binding; D011993:Recombinant Fusion Proteins; D013329:Structure-Activity Relationship; D014407:Tumor Cells, Cultured; D025521:Tumor Suppressor Proteins; D044767:Ubiquitin-Protein Ligases; D051794:Von Hippel-Lindau Tumor Suppressor Protein
D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D025521:Tumor Suppressor Proteins; D044767:Ubiquitin-Protein Ligases; D051794:Von Hippel-Lindau Tumor Suppressor Protein; D008025:Ligases; C497446:VHL protein, human