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A Novel Method for Analyzing Mitochondrial Movement: Inhibition by Paclitaxel in a Pheochromocytoma Cell Model

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Abstract

A method was developed to assess mitochondrial movement in the living cell that is dependent, in part, on microtubule and/or associating protein interactions. The leader sequence from cytochrome-c was used to drive DsRed2 fluorescent proteins to accumulate in the mitochondria, thus enabling to follow mitochondrial (cytochrome-c’s) movement. For calculating the percentage of mitochondrial movement, an image-processing program was used (ImageJ). Paclitaxel, an antitumor agent, is a potent microtubule-stabilizing agent that increases the stability of tubulin polymers, inhibiting mitosis and mitochondrial activity in dividing cells. Here, we tested whether paclitaxel inhibits mitochondrial movement in pheochromocytoma cells (a neuronal model, when tested in a differentiated state). While a 2-day exposure to paclitaxel resulted in cellular toxicity (measured as inhibition of mitochondrial activity), 2–3 h exposure to paclitaxel were sufficient to inhibit mitochondrial movement as assessed in 10–20-s imaging sessions in living cells. Mitotracker deep-red staining validated the staining obtained with DsRed2-cytochrome-c and identified intact mitochondria. Results showed a significant paclitaxel dose-dependent inhibition of mitochondrial movement. This new method should enable further assessment of microtubule-interacting drugs and other cytoskeletal components for their potential influence of mitochondrial movement as a test for activity and side effects.

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Acknowledgements

This work is the fulfillment of the M.Sc. thesis requirements of Mr. Tal Shprung. We thank Dr. Yevgeny Berdichevsky for his help with the cloning techniques, Dr. Leonid Mittelman for his help with the imaging equipment and Drs. Alistair Stewart, Sharon Furman-Assaf, and Bruce Morimoto for critical reading of the manuscript and excellent suggestions. Support was provided by the BSF, ISF, and Allon Therapeutics Inc. Professor Gozes is the incumbent of the Lily and Avraham Gildor Chair for the Investigation of Growth Factors, and the Director of the Adams Super Center for Brain Studies, the Levie–Edersheim–Gitter Institute for Functional Brain Imaging and the Dr. Diana and Zelman Elton (Elbaum) Laboratory for Molecular Neuroendocrinology. Professor Illana Gozes serves as the Chief Scientific Officer of Allon Therapeutics Inc.

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Authors and Affiliations

  1. Elton Laboratory for Molecular Neuroendocrinology, Department of Human Molecular Genetics and Biochemistry, Sackler Medical School, Tel Aviv University, Tel Aviv, 69978, Israel

    Tal Shprung & Illana Gozes

  2. The Lily and Avraham Gildor Chair for the Investigation of Growth Factors, The Adams Super Center for Brain Studies, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, 69978, Israel

    Illana Gozes

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  1. Tal Shprung
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Correspondence to Illana Gozes.

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Shprung, T., Gozes, I. A Novel Method for Analyzing Mitochondrial Movement: Inhibition by Paclitaxel in a Pheochromocytoma Cell Model. J Mol Neurosci 37, 254–262 (2009). https://doi.org/10.1007/s12031-008-9129-8

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  • DOI: https://doi.org/10.1007/s12031-008-9129-8

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