1992 Fiscal Year Final Research Report Summary
Study on role of protein phosphorylation in sialagogue-stimulated parotid gland.
| Project/Area Number |
02454427
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
Functional basic dentistry
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| Research Institution | The University of Tokushima |
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Principal Investigator |
INOUE Hideo Tokushima Univ., School of Dentistry, Professor, 歯学部, 教授 (30028732)
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| Co-Investigator(Kenkyū-buntansha) |
TSURUMI Chizuko Tokushima Univ., School of Dentistry, Research Associate, 歯学部, 助手 (60236958)
ASHIDA Yoshiyuki Tokushima Univ., School of Dentistry, Research Associate, 歯学部, 助手 (40212500)
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| Project Period (FY) |
1990 – 1992
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| Keywords | sialagogue / protein phosphorylation / parotid / ornithine decarboxylase / amylase secretion / proto-oncogene / mRNA / herbimycin A |
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| Research Abstract |
Changes in protein phosphorylation and mRNA levels of ornithine decarboxylase (ODC) and proto-oncogenes in sialagogue-stimulated parotid gland were investigated in rat parotid explants cultured on siliconized lens paper floating on serum-free 199 medium.
The results were summarized as follows;
1) Isoproterenol (IPR) enhanced the phosphorylation of serine/threonine residues of four proteins with apparent molecular weights of 17K, 20K, 31K and 32K and carbachol (CC) stimulated the phosphorylation of31K and 32K proteins. These protein phosphorylations preceded the increase of ODC activity. IPR-dependent ODC induction and phosphorylation of the proteins were selectively suppressed by monensin but not by polymyxin B, whereas CC-dependent ODC induction and protein phosphorylation were inhibited by polymyxin B but not by monensin. Neither monensin nor polymyxin B suppressed IPR- or CC-stimulated amylase secretion.
2) Northern/dot and Western blot analyses revealed that the sialagogues (IPR, CC,
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methoxamine) elevated the steady-state levels of ODC mRNA and ODC protein to maxima at 2 h and 6 h, respectively, after stimulation. The increases were roughly proportional to those in ODC activity, suggesting that sialagogue-dependent enzyme induction is regulated at the transcriptional level. The mRNAs of four of nine proto-oncogenes examined showed sialagogue-dependent increases to maxima at 30 min (c-fos) or 60 min (c-jun, c-myc and c-src) after stimulation. These increases were all transient, with the levels returning to the unstimulated levels within 60 min.
3) Tyrosine kinase inhibitors such as herbimycin A, genistein and methyl 2,5-dihydroxycinnamate inhibited a sialagogue-stimulated ODC induction but enhanced a sialagogue-induced amylase secretion. A tyrosine phosphatase inhibitor, Na_3 VO_4, increased ODC activity and this increase was suppressed by herbimycin A.
These results suggest that serine/threonine and tyrosine phosphorylations of proteins participate in a sialagogue-stimulated ODC induction, and that protein tyrosine phosphorylation shows an inhibitory effect on amylase secretion. Less
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