We herein report the subcellular distribution of methionine transfer RNA (tRNA^Met), which has two forms of initiator tRNA^Met (Meti) and elongator tRNA^Met (Mete). Meti induces translation initiation, and Mete is responsible for the elongation of the peptide chain. A recent report showed that the formation of tRNA^Met granules occurs following heat shock stress. We analyzed the subcellular distribution change in tRNA^Met using ionizing irradiation. The images acquired using confocal microscopy showed the presence of Mete granules—but not Meti granules—in the cytoplasm. Furthermore, while the Mete granules co-localized with T-cell intracellular antigen-1, they did not co-localize to typical stress granules nor processing body specific markers.

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Aurora B kinase activity is known to be required for chromosome alignment, segregation and cytokinesis. Its function in vivo has been characterized with small-molecule inhibitors, such as ZM447439, Hesperadin and VX-680. Since the inhibition of Aurora B kinase often results in a premature exit of mitosis without chromosome alignment, the events after chromosome segregation have yet to be fully analyzed. To monitor the typical events of mitosis and cytokinesis in the living state, we constructed two fluorescent human melanoma MDA-MB-435S cell lines, expressing mPlum-histoneH3/EGFP-survivin or mPlum-histone-H3/mOrange-actin. We examined the effect of the Aurora B kinase inhibitor AZD1152-HQPA on the entire process from the initiation of mitosis to the completion of cytokinesis. We extensively captured live cell images using a multi-point time lapse imaging system equipped with a PlanApo 60x objective. Although most of the mitotic cells showed premature mitotic exit, some populations of the cells proceeded to anaphase but failed to complete cytokinesis, resulting in binucleated cells. In these cells, we noticed that a chromosome bridge had been formed at the cleavage site during chromosome segregation and that the bridge was maintained after the ingression of the cleavage furrow, known as an “anaphase bridge” in mis-segregating cells. The cytokinesis of these cells was ultimately interrupted with the cleavage furrow regression. A multi-point time lapse imaging analysis of a nonsynchronous, randomly growing cell culture in a higher magnification would be useful for characterizing the real-time effects of other Aurora kinase-inhibitors in living cells.

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