The mammalian gamma-tubulin complex contains homologues of the yeast spindle pole body components spc97p and spc98p.
gamma-Tubulin is a universal component of microtubule organizing centers where it is believed to play an important role in the nucleation of microtubule polymerization. gamma-Tubulin also exists as part of a cytoplasmic complex whose size and complexity varies in different organisms. To investigate the composition of the cytoplasmic gamma-tubulin complex in mammalian cells, cell lines stably expressing epitope-tagged versions of human gamma-tubulin were made. The epitope-tagged gamma-tubulins expressed in these cells localize to the centrosome and are incorporated into the cytoplasmic gamma-tubulin complex. Immunoprecipitation of this complex identifies at least seven proteins, with calculated molecular weights of 48, 71, 76, 100, 101, 128, and 211 kD. We have identified the 100- and 101-kD components of the gamma-tubulin complex as homologues of the yeast spindle pole body proteins Spc97p and Spc98p, and named the corresponding human proteins hGCP2 and hGCP3. Sequence analysis revealed that these proteins are not only related to their respective homologues, but are also related to each other. GCP2 and GCP3 colocalize with gamma-tubulin at the centrosome, cosediment with gamma-tubulin in sucrose gradients, and coimmunoprecipitate with gamma-tubulin, indicating that they are part of the gamma-tubulin complex. The conservation of a complex involving gamma-tubulin, GCP2, and GCP3 from yeast to mammals suggests that structurally diverse microtubule organizing centers such as the yeast spindle pole body and the animal centrosome share a common molecular mechanism for microtubule nucleation.
Murphy
S M
SM
Department of Biological Sciences, Stanford University, Stanford, California.
Urbani
L
L
Stearns
T
T
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002460:Cell Line; D002461:Cell Line, Transformed; D018385:Centrosome / Q000378:metabolism; D003001:Cloning, Molecular; D006224:Cricetinae; D018076:DNA, Complementary; D006801:Humans; D051379:Mice; D008869:Microtubule-Associated Proteins / Q000032:analysis* / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D008941:Spindle Apparatus / Q000737:chemistry*; D018411:Spodoptera; D014404:Tubulin / Q000737:chemistry*; D014981:Xenopus
D018076:DNA, Complementary; D008869:Microtubule-Associated Proteins; C112305:TUBGCP2 protein, human; C112306:TUBGCP3 protein, human; D014404:Tubulin