1994 Fiscal Year Final Research Report Summary
Structural and Functional Diversity of Eukaryotic Vacuolar ATPases : Identification of Superfamily in Mammalian Cells and Studies on the Physiological Roles of Vacuolar ATPases by Functional and Genetical Disruption
| Project/Area Number |
05671824
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Biological pharmacy
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| Research Institution | Okayama University |
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Principal Investigator |
NOUMI Takato Okayama University, Faculty of Engineering Department of Biotechnology Associate Professor, 工学部, 助教授 (90189374)
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| Project Period (FY) |
1993 – 1994
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| Keywords | Vacuolar ATPase / H^+_-ATPase / H^+_-pump / Gene structure / Pappiloma virus / Lysosomal pH |
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| Research Abstract |
(1) Transformation of Rat Fibroblasts by E5 Onco-protein from Bovine Pappiloma Virus
The gene for E5 onco-protein was cloned under the control of SV40 promoter from bovine pappiloma virus genome. The expression plasmid was transfected into the rat fibroblasts and stable transformants were isolated. The oncogenic transformation was stimulated by PDGF or co-transfection with PDGF-R and proteolipid cDNAs. The intra-lysosomal pH was revealed to be elevated in these transformants. Currently the molecular mechanism for cell transformation by E5 through association with PDGF-R and proteolipid is under the investigation.
(2) Molecular Cloning and Genomic Structures of Genes for Mouse Proteolipid
The genomic library from mouse was screened and genes encoding proteolipid were isolated. The genome structure including 3 exons were determined by DNA sequenceing. Two possible pseudo-genes were also isolated and their genome structures were determined. Expression of proteolipid gene in ES cells were confirmed by Northern blot analysis. The targeting vector for disruption of the proteolipid gene was constructed and transfected into ES cells by electroporation. Currently homologous recombinants are being screened.
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