XB-ART-10876Methods Mol Med. June 1, 2000; 11 (6): 2047-56.
Cytoplasmic dynein-mediated assembly of pericentrin and gamma tubulin onto centrosomes.
Centrosome assembly is important for mitotic spindle formation and if defective may contribute to genomic instability in cancer. Here we show that in somatic cells centrosome assembly of two proteins involved in microtubule nucleation, pericentrin and gamma tubulin, is inhibited in the absence of microtubules. A more potent inhibitory effect on centrosome assembly of these proteins is observed after specific disruption of the microtubule motor cytoplasmic dynein by microinjection of dynein antibodies or by overexpression of the dynamitin subunit of the dynein binding complex dynactin. Consistent with these observations is the ability of pericentrin to cosediment with taxol-stabilized microtubules in a dynein- and dynactin-dependent manner. Centrosomes in cells with reduced levels of pericentrin and gamma tubulin have a diminished capacity to nucleate microtubules. In living cells expressing a green fluorescent protein-pericentrin fusion protein, green fluorescent protein particles containing endogenous pericentrin and gamma tubulin move along microtubules at speeds of dynein and dock at centrosomes. In Xenopus extracts where gamma tubulin assembly onto centrioles can occur without microtubules, we find that assembly is enhanced in the presence of microtubules and inhibited by dynein antibodies. From these studies we conclude that pericentrin and gamma tubulin are novel dynein cargoes that can be transported to centrosomes on microtubules and whose assembly contributes to microtubule nucleation.
PubMed ID: 10848628
PMC ID: PMC14902
Article link: Methods Mol Med.
Grant support: GM-51994 NIGMS NIH HHS , R01 GM051994 NIGMS NIH HHS , R56 GM051994 NIGMS NIH HHS , GM-51994 NIGMS NIH HHS
Genes referenced: dctn2 dnai1 dync1li1 pcnt tubg1