Summary of Research Project Results Under the JSPS FY2000
"Research for the future Program"



1.Research Institution The University of Tokyo
 
2.Research Area Life Sciences
 
3.Research Field Cellular Signaling
 
4.Term of Project FY1996〜FY2000
 
5.Project Number 96L00308
 
6.Title of Project Molecular interaction of inositolphospholipids with signal proteins

7.Projetct Leader
Name Institution,Department Title of Position
Tadaomi, Takenawa The University of Tokyo, The Institute of Medical Science Professor

8.Core Members

Names Institution,Department Title of Position
Kiyoko, Fukami The University of Tokyo, The Institute of Medical Science Lecture

9.Cooperating Researchers

Names Institution,Department Title of Position
Yasuhiro, Mochizuki The University of Tokyo, The Institute of Medical Science Post Doctor

10.Summary of Research Results

To elucidate the new functions of PI(4,5)P2, we surveyed the new PI(4,5)P2 binding domains. It is already well known that PH domains act as PI(4,5)P2 binding domains, which are contained in a variety of proteins. Here, we found that ENTH domain contained in epsin bound to PI(4,5)P2. The ENTH domain also bound to PI(3,4,5)P3 but not other inositolphosphatides. We further investigated the 3 dimentional structure of the ENTH domain by NMR. The ENTH domain consists of 6 a-helix. In the binding to PI(4,5)P2, N-terminal unstructural region and the loop between helix 3 and 4 play important roles. Substitution of basic amino acids including those areas to alanine completely diminished the binding activity.
Epsin was known to be essential for the vesicle formation induced by clathrin complexes. To demonstrate PI(4,5)P2 binding is crucial for the vesicle formation, we studied the effect on the internalization of EGF in PI(4,5)P2-bining lost mutant expressing cells. That mutant inhibited the internalization strongly. All these data show that Epsin ENTH domain is PI(4,5)P2 binding domain and its binding is essential for endocytosis.

11.Key Words

(1)PI(4,5)P2 binding、(2)Endocytosis、(3)Epsin
(4)ENTH domain、(5)Domain structure

12.References

[Reference Articles]
Author Title of Article
Park, S Phosphatidylinositol 4-phosphate 5-kinase type I is regulated through phosphorylation response by extracellular stimuli
Journal Volume Year Pages Concerned
J. Biol. Chem. 274 2001 4781-4787

Author Title of Article
Itoh, T. The Epsin N-terminal homology(ENTH) domain:a Phosphatidylinositol 4, 5-bisphosphate binding domain esential for endocytosis
Journal Volume Year Pages Concerned
Science 291 2001 1047-1051

Author Title of Article
Zhang, Y. Phosphatidylinositol 4-phosphate 5-kinase its3 and calcineurin ppb1 coordinately regulate cytokinesis in fission yeast.
Journal Volume Year Pages Concerned
J. Biol. Chem. 275 2000 35600-35606

Author Title of Article
Ijyuuin, T. Identification and characterization of a novel inositol polyphosphate 5-phosphatase.
Journal Volume Year Pages Concerned
J. Biol. Chem. 275 2000 10870-10875

Author Title of Article
Fukami, K. Growth factor-induced promoter activation of murine phospholipase C delta4 gene.
Journal Volume Year Pages Concerned
Eur J Biochem. 267 2000 28-36


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