Invited Symposium: Molecular and Cellular Analysis of Dopamine and Serotonin Transporters
Ratovitski, T (Department of Molecular Genetics, Weizmann Institute of Science, Israel)
Tauber, M (Department of Molecular Genetics, Weizmann Institute of Science, Israel)
Tafet, G (Department of Molecular Genetics, Weizmann Institute of Science, Israel)
Pantaleoni, G (Department of Molecular Genetics, Weizmann Institute of Science, Israel)
Simantov, R (Department of Molecular Genetics, Weizmann Institute of Science, Israel)
Antibodies for serotonin transporter (SERT) were used to assess expression of SERT in neuronal and non-neuronal cells, and to investigate trancriptional and post-tranlational regulations of this gene. Immunoblotting of NMB and COS-7 cells transfected with pCMV-rSERT revealed two immunoreactive bands, 83±3 and 110±3 kDa, that were larger than the two bands of human placental JAR cells (75±3 and 95±3 kDa), expressing the transporter constitutively. The inhibitor of N-linked glycosylation tunicamycin decreased by 40% the size of SERT in both NMB- and COS-7-transfected cells, and in JAR cells. It is well known that a short treatment with TPA decreases serotonin uptake. Whether TPA also has a long-term effect on serotonin transport, possibly at the transcriptional level, was studied. TPA treatment for 24-72 hr increased 3H-5HT uptake in JAR cells in a time- and dose-dependent way, and Lineweaver-Burk analysis indicated an increase of 2-3 folds in Vmax, with no effect on Km. Phorbol analogues that do not activate protein kinase C did not increase 3H-5HT uptake. As expected, 30 min treatment with TPA had no effect on SERT protein level, whereas 48 hr treatment increased SERT immunoreactivity. The cyclic-AMP phosphodiesterase inhibitor IBMX, like cholera toxin, increased 3H-5HT uptake. Interestingly, the effect of long-term co-treatment with cholera toxin and TPA was not additive. RT-PCR analysis showed that the glucocorticoid hormone dexamethasone inhibited the long-term effect of TPA on SERT mRMA level. Whether transcription factors such as AP-1 proteins participate in modulating expression of SERT gene is discussed.
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|Dus, L; Ratovitski, T; Tauber, M; Tafet, G; Pantaleoni, G; Simantov, R; (1998). Transcriptional and Post-translational Regulation of Serotonin Transporters Expressed Constitutively or in Transfected Cells. Presented at INABIS '98 - 5th Internet World Congress on Biomedical Sciences at McMaster University, Canada, Dec 7-16th. Invited Symposium. Available at URL http://www.mcmaster.ca/inabis98/simantov/dus0133/index.html|
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