Search for the biological significance of the interaction between Plk1 and RACK1

• Main Authors: Siao-Mei Chen, 陳筱梅
• Other Authors: Chin Li
• Format: Others
• Language: zh-TW
• Published: 2007
• Online Access: http://ndltd.ncl.edu.tw/handle/57429165865266668534

碩士 === 國立中正大學 === 分子生物研究所 === 95 === Polo-like kinase 1 (Plk1) is a serine-threonine protein kinase functioning in the regulation of multiple aspects of mitosis. Using recombinant linker-polo box domain protein in an in vitro pull-down assay followed by mass spectrometry to search for Plk1 interacting partners, we identified receptor for activated C kinase 1 (RACK1) as a putative interacting protein. Therefore, we wish to confirm and to understand the biological significance of the interaction between Plk1 and RACK1. RACK1 was originally identified as an anchoring protein for activated kinase C (PKC). By virtue of its ability to coordinate the interaction of key signaling molecules, RACK1 is thought to play a central role in critical biological responses, including signal transduction and translation. Using the cell extracts prepared from various stages of cell cycle, we determined that the linker-PBD interacts with RACK1 in the extracts at G1/S junction and at S phase. However, recombinant RACK1 protein failed to interact with the Plk1 proteins in vitro. These conflicted results could be due to that posttranslational modification of RACK1 is required for stable complex formation with Plk1, or the RACK1-Plk1 interaction could be bridged by another unknown protein. Since, treaty the cell extracts with λ-PPase before pull down assay, showed no effect to the interaction between RACK1 and Plk1, it is likely that on unknown cellular protein is required to bridge the interaction between Plk1 and RACK1. Since RACK1 is an integrated component of ribosomes, we thus first examined whether Plk1 is involved in translation regulation. The results of the preliminary experiments employing coupled in vitro transcription-translation assay showed that addition of recombinant Plk1 in the reaction led to inhibition of translation in a Plk1 kinase activity independent manner. The biological significance of our current findings is under further investigation.