PACSIN proteins bind tubulin and promote microtubule assembly.
PACSINs are intracellular adapter proteins involved in vesicle transport, membrane dynamics and actin reorganisation. In this study, we report a novel role for PACSIN proteins as components of the centrosome involved in microtubule dynamics. Glutathione S-transferase (GST)-tagged PACSIN proteins interacted with protein complexes containing alpha- and gamma-tubulin in brain homogenate. Analysis of cell lysates showed that all three endogenous PACSINs co-immunoprecipitated dynamin, alpha-tubulin and gamma-tubulin. Furthermore, PACSINs bound only to unpolymerised tubulin, not to microtubules purified from brain. In agreement, the cellular localisation of endogenous PACSIN 2 was not affected by the microtubule depolymerising reagent nocodazole. By light microscopy, endogenous PACSIN 2 localised next to gamma-tubulin at purified centrosomes from NIH 3T3 cells. Finally, reduction of PACSIN 2 protein levels with small-interfering RNA (siRNA) resulted in impaired microtubule nucleation from centrosomes, whereas microtubule centrosome splitting was not affected, suggesting a role for PACSIN 2 in the regulation of tubulin polymerisation. These findings suggest a novel function for PACSIN proteins in dynamic microtubuli nucleation.
Grimm-Günter
Eva-Maria S
EM
Center for Biochemistry, Medical Faculty, University of Cologne, Joseph-Stelzmann-Str. 52, D-50931 Cologne, Germany.
Milbrandt
Mark
M
Merkl
Barbara
B
Paulsson
Mats
M
Plomann
Markus
M
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001921:Brain / Q000378:metabolism; D018385:Centrosome / Q000378:metabolism*; D003598:Cytoskeletal Proteins; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D008870:Microtubules / Q000378:metabolism*; D041681:NIH 3T3 Cells; D009479:Neuropeptides / Q000235:genetics / Q000378:metabolism*; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D014404:Tubulin / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D003598:Cytoskeletal Proteins; D047908:Intracellular Signaling Peptides and Proteins; D009479:Neuropeptides; C120714:PACSIN2 protein, human; C114726:Pacsin1 protein, mouse; C485644:Pacsin2 protein, mouse; C486019:Pacsin3 protein, mouse; D010750:Phosphoproteins; D020033:Protein Isoforms; D011506:Proteins; D034741:RNA, Small Interfering; D014404:Tubulin