Specific regulation of CENP-E and kinetochores during meiosis I/meiosis IItransition in pig oocytes

To understand the mechanisms which regulate meiosis-specific cell cycle and chromosome distribution in mammalian oocytes, the level and the localizati on of CENP-E and the kinetochore number and direction on a half bivalent we re examined during pig oocyte maturation. CENP-E is a kinetochore motor pro tein whose intracellular level and localization are strictly regulated in t he somatic cell cycle. The localizations of CENP-E on meiotic chromosomes f rom diakinesis stage to anaphase I and at the spindle midzone at telophase I were shown by immunofluorescent confocal microscopy to be similar to thos e in somatic cells of pig and other species. Further, ultrastructural analy sis revealed the presence of CENP-E on fibrous corona and outer plate of ki netochores of the meiotic chromosomes. However, unlike mitosis, CENP-E stai ning was continuously detected either at the spindle midzone or on the kine tochores of segregated chromosomes during the first polar body emission. Co nsistent with this, immunoblot analysis revealed that CENP-E level remained high during meiosis l/meiosis II (MI/MII) transition and that some of CENP -E survived through the transition even in cycloheximide-treated oocytes in which cyclin B1 was completely degraded. Furthermore, examinations of CENP -E signals in confocal microscopy and kinetochores in electron microscopy i n MI and MH oocytes provide the cytological evidence in mammalian oocytes w hich suggests that each sister chromatid in a pair has its own kinetochore which localizes side-by-side so that two sister chromatids on a half bivale nt are oriented toward and connected to the same pole in M1. (C) 2000 Wiley -Liss, Inc.