Quantitative image analysis identifies pVHL as a key regulator of microtubule dynamic instability.
Von Hippel-Lindau (VHL) tumor suppressor gene mutations predispose carriers to kidney cancer. The protein pVHL has been shown to interact with microtubules (MTs), which is critical to cilia maintenance and mitotic spindle orientation. However, the function for pVHL in the regulation of MT dynamics is unknown. We tracked MT growth via the plus end marker EB3 (end-binding protein 3)-GFP and inferred additional parameters of MT dynamics indirectly by spatiotemporal grouping of growth tracks from live cell imaging. Our data establish pVHL as a near-optimal MT-stabilizing protein: it attenuates tubulin turnover, both during MT growth and shrinkage, inhibits catastrophe, and enhances rescue frequencies. These functions are mediated, in part, by inhibition of tubulin guanosine triphosphatase activity in vitro and at MT plus ends and along the MT lattice in vivo. Mutants connected to the VHL cancer syndrome are differentially compromised in these activities. Thus, single cell-level analysis of pVHL MT regulatory function allows new predictions for genotype to phenotype associations that deviate from the coarser clinically defined mutant classifications.
Thoma
Claudio R
CR
Institute of Cell Biology, Swiss Federal Institute of Technology (ETH) Zurich, 8093 Zurich, Switzerland.
Matov
Alexandre
A
Gutbrodt
Katrin L
KL
Hoerner
Christian R
CR
Smole
Zlatko
Z
Krek
Wilhelm
W
Danuser
Gaudenz
G
D000595:Amino Acid Sequence; D016000:Cluster Analysis; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis / Q000187:drug effects; D021621:Imaging, Three-Dimensional / Q000379:methods*; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D008870:Microtubules / Q000187:drug effects / Q000378:metabolism*; D008969:Molecular Sequence Data; D015739:Nocodazole / Q000494:pharmacology; D010641:Phenotype; D017354:Point Mutation / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014404:Tubulin / Q000378:metabolism; D051794:Von Hippel-Lindau Tumor Suppressor Protein / Q000737:chemistry / Q000378:metabolism*; D006623:von Hippel-Lindau Disease / Q000235:genetics
C403664:MAPRE3 protein, human; D008869:Microtubule-Associated Proteins; D011993:Recombinant Fusion Proteins; D014404:Tubulin; D049452:Green Fluorescent Proteins; D006160:Guanosine Triphosphate; D051794:Von Hippel-Lindau Tumor Suppressor Protein; D015739:Nocodazole