mt(1) melatonin receptor stimulation effects on microtubule-associated proteins

Melatonin, N-acetyl-5-methoxytryptamine, is the primary hormone secreted from the pineal gland. It is involved with several biological functions such as circadian and seasonal rhythms and reproduction. However, its cellular mechanisms are unknown. Witt-Enderby et al (2000) previously found that when pharmacological concentrations of melatonin are applied to Chinese hamster ovary (CHO) cells transformed to express the human mt1 melatonin receptor (mt1-CHO), neurite-like outgrowths extend from the cells in a bipolar morphology. In neuronal cells, microtubules are the main structural component involved in elongating and supporting growing neurites. In this study, fluorescence microscopy showed that the melatonin-induced outgrowths contain tubulin bundles, resulting from the induced rearrangement of the microtubule cytoskeleton. Through Western blot analysis, specific "neuronal" microtubule associated proteins (MAP) were not present in the non-neuronal mt1-CHO cell line. "Neuronal" MAPs were found only in the two neuronal cell lines tested, N1E-115 and BE(2)C. However, melatonin had no effect on "neuronal" MAP protein levels. In addition, the microtubule motor protein, cytoplasmic dynein, which is involved in tubulin transport, was found to be sensitive to melatonin in the mt1-CHO cell line. Further, Western blot analysis of cytoplasmic dynein protein levels with inhibitors of transcription and translation suggested that the protein was relatively stable and stored mRNA may be present in the mt1-CHO cells. In addition, the cytoplasmic dynein levels present in the cell before melatonin treatments were not sufficient for outgrowth formation. Cytoplasmic dynein was thought to have a major role in the production of the melatonin-induced neurite-like outgrowths.