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Article

Antibody Microarray Analyses of Signal Transduction Protein Expression and Phosphorylation during Porcine Oocyte Maturation

Supporting Info

Steven Pelech*^†^§, Lucie Jelinkova^#, Andrej Susor^#, Hong Zhang^†, Xiaoqing Shi^§, Antonin Pavlok^#, Michal Kubelka^# and Hana Kovarova^#

Kinexus Bioinformatics Corporation, Suite 1, 8755 Ash Street, Vancouver, BC, Canada V6P 6T3, Department of Medicine, University of British Columbia, Vancouver, BC, Canada, and Department of Reproductive and Developmental Biology, Institute of Animal Physiology and Genetics, Rumburska 89, Libechov, Czech Republic

J. Proteome Res., 2008, 7 (7), pp 2860–2871

DOI: 10.1021/pr800082a

Publication Date (Web): May 17, 2008

* To whom correspondence should be addressed. E-mail, spelech@kinexus.ca.; phone, 604-323-2547 , extension, 10; fax, 604-323-2548.

†

Kinexus Bioinformatics Corporation.

Department of Medicine, University of British Columbia.

Department of Reproductive and Developmental Biology, Institute of Animal Physiology and Genetics.

Abstract

Kinex antibody microarray analyses was used to investigate the regulation of 188 protein kinases, 24 protein phosphatases, and 170 other regulatory proteins during meiotic maturation of immature germinal vesicle (GV+) pig oocytes to maturing oocytes that had completed meiosis I (MI), and fully mature oocytes arrested at metaphase of meiosis II (MII). Increases in apparent protein levels of protein kinases accounted for most of the detected changes during the GV to MI transition, whereas reduced protein kinase levels and increased protein phosphorylation characterized the MI to MII transition. During the MI to MII period, many of the MI-associated increased levels of the proteins and phosphosites were completely or partially reversed. The regulation of these proteins were also examined in parallel during the meiotic maturation of bovine, frog, and sea star oocytes with the Kinex antibody microarray. Western blotting analyses confirmed altered expression levels of Bub1A, IRAK4, MST2, PP4C, and Rsk2, and the phosphorylation site changes in the kinases Erk5 (T218 + Y220), FAK (S722), GSK3-beta (Y216), MEK1 (S217 + S221) and PKR1 (T451), and nucleophosmin/B23 (S4) during pig oocyte maturation.

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