LIM kinase activity is required for microtubule organising centre positioning in mouse oocyte meiosis
Xin Li A , Yubo Zhu B , Yan Cao A , Qian Wang A , Juan Du A , Jianhui Tian C , Yuanjing Liang A D and Wei Ma A D
+ Author Affiliations
- Author Affiliations
A Department of Histology and Embryology, School of Basic Medical Sciences, Capital Medical University, Beijing, 100069, China.
B College of Animal Science and Veterinary Medicine, Shenyang Agricultural University, Shenyang, 110886, China.
C Key Laboratory of Animal Genetics and Breeding of the Ministry of Agriculture, College of Animal Sciences and Technology, China Agricultural University, Beijing, 100094, China.
D Corresponding authors. Emails: mawei1026@ccmu.edu.cn; y3liang@126.com
*These authors contributed equally to this work.
Reproduction, Fertility and Development 29(4) 791-804 https://doi.org/10.1071/RD15406
Submitted: 8 October 2015 Accepted: 27 November 2015 Published: 6 January 2016
Abstract
LIM kinase 1 (LIMK1) activity is essential for cell migration and cell cycle progression. Little is known about LIMK1 expression and function in mammalian oocytes. In the present study we assessed LIMK1 protein expression, subcellular distribution and function during mouse oocyte meiosis. Western blot analysis revealed high and stable expression of LIMK1 from the germinal vesicle (GV) to MII stage. In contrast, activated LIMK1 (i.e. LIMK1 phosphorylated at threonine 508 (pLIMK1Thr508)) was only detected after GV breakdown, with levels increasing gradually to peak at MI and MII. Immunofluorescence showed pLIMK1Thr508 was colocalised with the microtubule organising centre (MTOC) components pericentrin and γ-tubulin at the spindle poles. A direct interaction between γ-tubulin and pLIMK1Thr508 was confirmed by co-immunoprecipitation. LIMK inhibition with 1 μM BMS3 damaged MTOC protein localisation to spindle poles, undermined the formation and positioning of functional MTOC and thus disrupted spindle formation and chromosome alignment. These effects were phenocopied by microinjection of LIMK1 antibody into mouse oocytes. In summary, the data demonstrate that LIMK activity is essential for MTOC organisation and distribution and so bipolar spindle formation and maintenance in mouse oocytes.
Additional keywords: bipolar establishment, BMS3, spindle formation.
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