Association of polo-like kinase with alpha-, beta- and gamma-tubulins in a stable complex.
The polo-like kinase (Plk) family has been shown to have an important role in the regulation of the cell-division cycle, especially in organization of the spindle structure, in species from fungi to humans. Recent reports have demonstrated that in mammalian cells Plk is associated with components of the anaphase-promoting complex and a peptidyl-prolyl isomerase, Pin1. To characterize a putative Plk-containing complex, we fractionated mitotic cell lysates on a gel-filtration column. The Plk complex was eluted from the column at molecular sizes ranging from 669 to 2500 kDa in the presence of detergent and high concentrations of salt. Specific associations of Plk with alpha-, beta- and gamma-tubulins in both interphase and mitotic cells were shown by reciprocal immunoprecipitations and immunoblottings and were independent of the microtubule polymerization state, whereas binding assays in vitro indicated that Plk interacts with alpha- and beta-tubulins directly. In addition, mitotic Plk was able to phosphorylate associated tubulins in vitro. Finally, we show that the kinase domain of the Plk molecule is both required and sufficient for its binding to tubulins in vivo. The specific interaction between Plk and tubulins might provide a molecular basis for the physiological functions of Plk in regulating the cell cycle, particularly in establishing the normal bipolar spindle.
Feng
Y
Y
Biological Mechanisms Section, Laboratory of Leukocyte Biology, NCI-FCRDC, Frederick, MD 21702, USA.
Hodge
D R
DR
Palmieri
G
G
Chase
D L
DL
Longo
D L
DL
Ferris
D K
DK
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D017931:DNA Primers; D007399:Interphase; D051379:Mice; D008938:Mitosis; D010766:Phosphorylation; D011233:Precipitin Tests; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D013912:Threonine / Q000378:metabolism; D014404:Tubulin / Q000302:isolation & purification / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D014404:Tubulin; D013912:Threonine; D012694:Serine; D017346:Protein Serine-Threonine Kinases