• Search Research Projects
  • Search Researchers
  • How to Use
  1. Back to project page

1998 Fiscal Year Final Research Report Summary

AUTOREHLATION OF TRANSCRIPTION OF G-PROTEIN MESANGIAL GELLS

Research Project

Project/Area Number 09671185
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field Kidney internal medicine
Research InstitutionTOKYO WOMEN'S MEDICAL UNIVERSITY

Principal Investigator

KAWASHIMA Akira  TOKYO WOMEN'S MEDICAL UNIVERSITY,SCHOOL OF MEDICINE, INSTRUCTOR, 医学部, 助手 (20224769)

Co-Investigator(Kenkyū-buntansha) UCHIDA Keiko  TOKYO WOMEN'S MEDICAL UNIVERSITY,SCHOOL OF MEDICINE, INSTRUCTOR, 医学部, 助手 (60246478)
NITTA Kosaku  TOKYO WOMEN'S MEDICAL UNIVERSITY,SCHOOL OF MEDICINE, INSTRUCTOR, 医学部, 助手 (50241071)
Project Period (FY) 1997 – 1998
KeywordsAUTOREGULATION / TRANSCRIPTION / G-PROTEIN / LLC-PK1 CELL / Sp1
Research Abstract

A previous report demonstrated that Gαi-2 is regulated by steroid hormone. A mutation of the Gαi-2 gene that decreases GTPase activity produces the oncogene gip2. Autoregulation of this gene should be critical to prevent overgrowth of the cells. We detected Gai-2 protein in glomerular cells by immunostaining and immunoblotting. Then, we tried to transfect Gαi-2 gene into mesangial cells using calcium phosphate precipitation method or lipofection method. However, it was insufficient for the following experiments. Thus, we used LLC-PK1 cells in place of mesangial cells. In the cells transfected with 5'-flanking sequences fused to firefly luciferase cDNA reporter gene, we demonstrated a 24 % transcriptional repression of Gαi-2 gene after overexpressing Gαi-2 protein and a 88 % repression of gip2 protein when gip2 protein is overexpressed. Deletion analysis of the reporter gene indicated that the site of repression in gip2 transfected cells occurred in the region of the Gαi-2 gene promoter. Utilizing the 27-bp DNA segment as a probe in mobility shift assay, we identified a complex with decreased binding in gip2 induced cells. This DNA segment containing tow GC boxes was recognized specifically by a transcription factor Sp1. Antibody to Sp1 added to the mobility shift assay supershifted the complex. These studies demonstrate that overexpression of Gαi-2 protein represses Gαi-2 gene transcription by a reduction of transcription factor Sp1, and suggest the possibility of an autoregulation mechanism of Gαi-2 transcription.

URL: 

Published: 2001-10-23  

Information User Guide FAQ News Terms of Use Attribution of KAKENHI

Powered by NII kakenhi