The von Hippel-Lindau tumor suppressor protein influences microtubule dynamics at the cell periphery.
The von Hippel-Lindau (VHL) protein protects microtubules (MTs) from destabilization by nocodazole treatment. Based on this fixed-cell assay with static end points, VHL has been reported to directly stabilize the MT cytoskeleton. To investigate the dynamic changes in MTs induced by VHL in living cells, we measured the influence of VHL on tubulin turnover using fluorescence recovery after photobleaching (FRAP). To this end, we engineered VHL-deficient renal cell carcinoma cells to constitutively incorporate fluorescently labeled tubulin and to inducibly express VHL. Induction of VHL in these cells resulted in a decrease of tubulin turnover as measured by FRAP at the cell periphery, while minimally influencing MT dynamics around the centrosome. Our data indicates that VHL changes the behavior of MTs dependent on their subcellular localization implying a role for VHL in cellular processes such as migration, polarization, and cell-cell interactions. Here we propose a complementary method to directly measure VHL-induced subcellular changes in microtubule dynamics, which may serve as a tool to study the effect of MT binding proteins such as VHL.
Lolkema
Martijn P
MP
Department of Medical Oncology, University Medical Center Utrecht, Utrecht, The Netherlands.
Mehra
Niven
N
Jorna
Anita S
AS
van Beest
Moniek
M
Giles
Rachel H
RH
Voest
Emile E
EE
D002292:Carcinoma, Renal Cell / Q000473:pathology; D045744:Cell Line, Tumor; D018385:Centrosome; D003599:Cytoskeleton / Q000378:metabolism*; D005091:Exons; D005786:Gene Expression Regulation; D006801:Humans; D007700:Kinetics; D008856:Microscopy, Fluorescence / Q000379:methods; D008870:Microtubules / Q000378:metabolism*; D038761:Photobleaching; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000502:physiology*; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000502:physiology*; D051794:Von Hippel-Lindau Tumor Suppressor Protein
D025521:Tumor Suppressor Proteins; D044767:Ubiquitin-Protein Ligases; D051794:Von Hippel-Lindau Tumor Suppressor Protein; C497446:VHL protein, human