Inactivation of VHL by tumorigenic mutations that disrupt dynamic coupling of the pVHL.hypoxia-inducible transcription factor-1alpha complex.
The von Hippel-Lindau (VHL) gene product, pVHL, targets the alpha subunit of the hypoxia-inducible transcription factor (HIF-alpha) for ubiquitin-dependent degradation. This tumor suppressor function is mediated by the alpha- and beta-domains responsible for assembling the pVHL E3 ubiquitin ligase complex and for recognizing the prolyl-hydroxylated HIF-alpha, respectively. The molecular basis for a large number of tumor-derived mutations can be attributed to alterations that directly compromise the ability of pVHL to assemble the E3 or to contact the substrate. Here we describe a new mechanism of oncogenic inactivation by VHL missense mutations that lie in the L1 and L7 linker regions distal to the HIF-alpha-binding pocket. Employing molecular dynamics simulations, we show that the tumorigenic L1 loop mutation of Ser(65) to Leu, deficient in promoting the degradation of HIF-alpha, disrupts the coordination of internal motions of the pVHL.HIF-1alpha complex. Furthermore, we demonstrate that in addition to S65L, five other tumor-derived VHL mutations located within the L1 loop are each defective in mediating proteolysis of HIF-2alpha. Moreover, dynamic organization of pVHL.HIF-1alpha recognition is focally centered on Gln(145) within the L7 loop, and its tumorigenic mutant Q145H abolishes almost all of the correlated dynamic motions. Intriguingly, Q145H, whereas defective in targeting cellular HIF-alpha for degradation, had an attenuated hydroxylation dependence in binding to HIF-1alpha in vitro. Taken together, our results suggest that specific association between pVHL and the hydroxylated HIF-alpha requires both the L1 and L7 loops to coordinate dynamic coupling among distant pVHL regions, whose mutational disruption inactivates VHL and is hence responsible for tumorigenesis.
Miller
Felicia
F
Department of Oncological Sciences, The Mount Sinai School of Medicine, New York, New York 10029-6574, USA.
Kentsis
Alex
A
Osman
Roman
R
Pan
Zhen-Qiang
ZQ
D000199:Actins / Q000737:chemistry; D064173:Anaphase-Promoting Complex-Cyclosome; D000911:Antibodies, Monoclonal / Q000737:chemistry; D051792:Basic Helix-Loop-Helix Proteins; D002460:Cell Line; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D006900:Hydroxylation; D000860:Hypoxia; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007930:Leucine / Q000737:chemistry; D008239:Lysine / Q000737:chemistry; D008958:Models, Molecular; D009154:Mutation; D020125:Mutation, Missense; D009369:Neoplasms / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D011392:Proline / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012694:Serine / Q000737:chemistry; D013997:Time Factors; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D025521:Tumor Suppressor Proteins / Q000737:chemistry / Q000235:genetics* / Q000502:physiology*; D025801:Ubiquitin / Q000737:chemistry / Q000378:metabolism; D043743:Ubiquitin-Protein Ligase Complexes / Q000737:chemistry; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry / Q000235:genetics* / Q000502:physiology*; D051794:Von Hippel-Lindau Tumor Suppressor Protein; D000099162:Endothelial PAS Domain-Containing Protein 1
D000199:Actins; D064173:Anaphase-Promoting Complex-Cyclosome; D000911:Antibodies, Monoclonal; D051792:Basic Helix-Loop-Helix Proteins; D005982:Glutathione Transferase; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007930:Leucine; D008239:Lysine; D010455:Peptides; D011392:Proline; D012694:Serine; D015534:Trans-Activators; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D025801:Ubiquitin; D043743:Ubiquitin-Protein Ligase Complexes; D044767:Ubiquitin-Protein Ligases; D051794:Von Hippel-Lindau Tumor Suppressor Protein; D000099162:Endothelial PAS Domain-Containing Protein 1; C497442:HIF1A protein, human; C497446:VHL protein, human