1994 Fiscal Year Final Research Report Summary
The Effect of Ca^<2+> on the assembly and transport of Na, K-ATPase
| Project/Area Number |
05670059
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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 |
General physiology
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| Research Institution | Kansai Medical University |
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Principal Investigator |
OMORI Koichiro Kansai Med.Univ., Dpt.of Physiol., Assistant Prof., 医学部, 助教授 (80094465)
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| Co-Investigator(Kenkyū-buntansha) |
YAMAMOTO Akitsugu Kansai Med.Univ., Dpt.of Physiol., Lecturer, 医学部, 講師 (30174775)
MASAKI Ryuichi Kansai Med.Univ., Dpt.of Physiol., Lecturer, 医学部, 講師 (70140283)
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| Project Period (FY) |
1993 – 1994
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| Keywords | Na, K-ATPase / Thapsigargin / BAPTA-AM / A23187 / Retinal pigment epithelial cell / Muscle spindle / Polycystic kidney / Thyroid hormone |
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| Research Abstract |
Na, K-ATPase consists of the alpha- and beta-subunits. The alpha-subunit contains the binding site for ouabain and plays as the catalytic subunit, whereas the beta-subunit is a glycosylated membrane protein and its function is not yet completely known. Both subunits are synthesized concurrently and assembled on the ER immediately after polypeptide synthesis. Recently, several studies have suggested the functional roles of the beta-subunit in the maturation of the alpha-subunit on the ER and/or the transport of this subunit to the plasma membrane. On the bases of these findings, we tried to elucidate the mechanisms of the assembly of both subunits and the transport of the enzyme.
1. We investigated the effects of Ca^<2+> on the assembly and transport of the enzyme using a HeLa cell clone permanently expressing N-terminal half of the beta-subunit (beta_N) which is confined to the ER in the cells. Thapsigargin, BAPTA-AM and A23187 treatments or low Ca^<2+> medium treatment could not move the beta_N from the ER to the plasma membrane. Interestingly, A23187 treatment caused the change of oligosaccharide processing in the beta-subunit, converting the molecular weight of the subunit from 5.5 K to 4.5 K.In addition, we detected beta・beta_N complex in A23187 treated cells, immediately after polypeptide synthesis. Elucidation of the functional significance of these findings is now in progress.
2. We analyzed the distribution of the enzyme in developing retinal pigment epithelial cells, muscle spindles and polycystic kidney, immunohistochemically.
3. We investigated the effects of T_3 on the gene expression and enzyme activity of Na, K-ATPase in rat liver. T_3 treatment increased the level of mRNA encoding alpha-and beta-subunits 3-4-fold, and both the amount and activity of the enzyme 2-fold.
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