Regulation of intracellular MEK1/2 translocation in mouse oocytes: cytoplasmic dynein/dynactin-mediated poleward transport and cyclin B degradation-dependent release from spindle poles.
We recently reported that MEK1/2 plays an important role in microtubule organization and spindle pole tethering in mouse oocytes, but how the intracellular transport of this protein is regulated remains unknown. In the present study, we investigated the mechanisms of poleward MEK1/2 transport during the prometaphase I/metaphase I transition and MEK1/2 release from the spindle poles during the metaphase I/anaphase I transition in mouse oocytes. Firstly, we found that p-MEK1/2 was colocalized with dynactin at the spindle poles. Inhibition of the cytoplasmic dynein/dynactin complex by antibody microinjection blocked polar accumulation of p-MEK1/2 and caused obvious spindle abnormalities. Moreover, coimmunoprecipitation of p-MEK1/2 and dynein or dynactin from mouse oocyte extracts confirmed their association at metaphase I. Secondly, disruption of microtubules by nocodazole resulted in the failure of poleward p-MEK1/2 transport. Whereas, when the nocodazole-treated oocytes were recovered in fresh culture medium, the spindle reformed and p-MEK1/2 relocalized to the spindle poles. Finally, we examined the mechanism of p-MEK1/2 release from the spindle poles. In control oocytes, polar p-MEK1/2 was gradually released during metaphase I/anaphase I transition. By contrast, in the presence of nondegradable cyclin B (Delta90), p-MEK1/2 still remained at the spindle poles at anaphase I. Our results indicate that poleward MEK1/2 transport is a cytoplasmic dynein/dynactin-mediated and spindle microtubule-dependent intracellular movement, and that its subsequent anaphase release from spindle poles is dependent on cyclin B degradation.
Xiong
Bo
B
State Key Laboratory of Reproductive Biology, Institute of Zoology and Graduate School, Chinese Academy of Sciences, Beijing, PR China.
Yu
Ling-Zhu
LZ
Wang
Qiang
Q
Ai
Jun-Shu
JS
Yin
Shen
S
Liu
Jing-He
JH
OuYang
Ying-Chun
YC
Hou
Yi
Y
Chen
Da-Yuan
DY
Zou
Hui
H
Sun
Qing-Yuan
QY
D000818:Animals; D019926:Cyclin B / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism*; D000072159:Dynactin Complex; D004398:Dyneins / Q000378:metabolism; D015151:Immunoblotting; D047468:Immunoprecipitation; D048369:MAP Kinase Kinase 1 / Q000378:metabolism*; D048370:MAP Kinase Kinase 2 / Q000378:metabolism*; D008677:Metaphase / Q000502:physiology*; D051379:Mice; D008856:Microscopy, Fluorescence; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D015739:Nocodazole / Q000633:toxicity; D009865:Oocytes / Q000166:cytology / Q000502:physiology; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D008941:Spindle Apparatus / Q000187:drug effects / Q000378:metabolism*; D050257:Tubulin Modulators / Q000633:toxicity
D019926:Cyclin B; D000072159:Dynactin Complex; D008869:Microtubule-Associated Proteins; D050257:Tubulin Modulators; C482128:MAP2K2 protein, human; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; D004398:Dyneins; D015739:Nocodazole