Regulation of translocation of parathyroid hormone-related protein at the ER membrane

2002 Fiscal Year Final Research Report Summary

• Project/Area Number: 13671934
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Functional basic dentistry
• Research Institution: NIIGATA UNIVERSITY
• Principal Investigator: AMAYA Yoshihiro Niigata University, Graduate School of Medical and Dental Sciences, Associate Professor, 大学院・医歯学総合研究科, 助教授 (50193032)
• Co-Investigator(Kenkyū-buntansha): MIURA Satoshi Yokohama City University, Faculty of Medicine, Associate Proffesor, 医学部, 助教授 (60157427) ; ODA Kimimitsu Niigata University, Graduate School of Medical and Dental, Sciences Professor, 大学院・医歯学総合研究科, 教授 (10122681)
• Project Period (FY): 2001 – 2002
• Keywords: protein translocation / endoplasmic reticulum / protein secretion / nucleus / parathyroid hormone-related protein

Research Abstract

Parathyroid hormone related protein (PTHrP) is known for its dual localization, that is, secretion out of the cell through the secretory pathway and transport to the nucleus and/or the nucleolus under lain conditions. In this project, we have attempted to reveal a possibility that the dual localization of the protein is regulated at the ER membrane during its translocation across the membrane. Two important conclusions can be drawn from our results
1. Although the ER targeting of the signal sequence of PTHrP dependent on the signal recognition particle (SRP) as usual secretory proteins, its post targeting reactions are quite different from usual one. The protein translocation promoted by the signal sequence of PTHrP is not cotranslational mode but posttranslational mode
2. Protein sequences that regulate the ER translocation are found in mid-region (35-87) and C-terminal region (107-141) of PTHrP. These sequences strongly inhibit the ER translocation of the protein at least in vitro reco nstitution system. Moreover, these sequences dissociate protease resistant ribosome membrane junction that is required for usual cotranslational translocation
These features of the precursor form of PTHrP would be advantageous for cytoplasmic molecules such as a receptor for protein nuclear localization, a lanase(s) and a component(s) of cytoskelton, which may involved in regulation of its localization
To identify molecules that involved in the regulation of translocation and the nuclear/nucleolar function of PTHrP, we established a method of purification of recombinant PTHrP proteins that are over produced in Escherichia coli cells under a native condition Several candidates that interact with ~ were found by pulldown assay of cytosolic and nucleoplasmic extinct with the recombinant proteins. We are going to identify these molecules and reveal their functions. Interaction of a part of these molecules are required a domain of PTHrP that is essential for interaction with the receptor for protein nuclear localization