Investigating the Spindle Assembly Checkpoint Regulation in C. elegans Spermatogenesis

• Main Authors: Shang-Yang Chen, 陳尚暘
• Other Authors: 吳瑞菁
• Format: Others
• Language: en_US
• Published: 2017
• Online Access: http://ndltd.ncl.edu.tw/handle/73405458770817628880

碩士 === 國立臺灣大學 === 醫學檢驗暨生物技術學研究所 === 105 === Cell division is important in all multicellular organisms for vital processes including growth, regeneration and reproduction. During cell cycle, the transition from metaphase to anaphase strictly controlled by Spindle Assembly Checkpoint (SAC). Until all chromosomes are properly attached by spindle microtubules, SAC prevents chromosome separation by inhibiting APC/C, an E3 ubiquitin ligase essential for releasing sister chromatid cohesion. In male meiosis, however, two consecutive chromosome separation events occur after one round of chromosome duplication. Thus far, it is unclear if both male meiotic divisions subjected to the control of canonical SAC signaling. To investigate if SAC signals function during male meiotic divisions, we developed an algorithm that allows automatic quantification of the levels of chromosome-associated proteins through the divisions in time-lapse recordings. We found that outer kinetochore SAC signaling protein BUB-1 released from chromosomes during first chromosome segregation event and recruited back to chromosome before second chromosome segregation takes place, indicating proper kinetochore structure and platform for SAC disassembled and re-assembled between two divisions. Contrarily, securin, the direct target of APC/C activity, though is degraded when first chromosome separation is initiated, fails to be recruited to chromosome during second division. These results indicate that the second male meiotic division does not required APC/C and the proteasome-dependent protein degradation. To test this, we examined the progression of division in primary and secondary spermatocytes treated with proteasome inhibitor MG132. As expected, primary spermatocytes treated with MG132 were stalled at metaphase I. Interestingly, secondary spermatocytes treated with MG132 were able to complete chromosome segregation and division. These results suggest the second male meiotic division might regulated differently compared to the first meiosis. Taken together, we hypothesize that the APC/C-proteasome system, which is crucial for canonical SAC signaling, does not participate in second chromosome segregation.