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BRCA1 regulates microtubule dynamics and taxane-induced apoptotic cell signaling

Oncogene volume 33, pages 1418–1428 (2014)Cite this article

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Abstract

The taxanes are effective microtubule-stabilizing chemotherapy drugs used in the treatment of various solid tumors. However, the emergence of drug resistance hampers their clinical efficacy. The molecular basis of clinical taxane resistance remains poorly understood. Breast cancer 1, early onset gene, BRCA1, is a tumor-suppressor gene, whose expression has been correlated with taxane sensitivity in many solid tumors including non-small cell lung cancer. However, the molecular mechanism underlying the relationship between BRCA1 (B1) expression and taxane activity remains unclear. To this end, we created a stable B1 knockdown A549 cell line (B1-KD) to investigate B1’s role in microtubule biology and response to taxane treatment. We show that B1-KD rendered A549 cells resistant to paclitaxel (PTX), phenocopying clinical studies showing that low B1 expression correlated with taxane resistance. As previously reported, we show that loss of B1 enhanced centrosomal γ-tubulin localization and microtubule nucleation. Interestingly, we found that the B1-KD cells exhibited increased microtubule dynamics as compared with parental A549 cells, as assessed by live-cell confocal microscopy using enhanced green fluorescent protein-tagged α-tubulin or EB1 protein. In addition, we showed that loss of B1 impairs the ability of PTX to induce microtubule polymerization using immunofluorescence microscopy and a cell-based tubulin polymerization assay. Furthermore, B1-KD cells exhibited significantly lower intracellular binding of a fluorescently labeled PTX to microtubules. Recent studies have shown that PTX-stabilized microtubules serves as a scaffold for pro-caspase-8 binding and induction of apoptosis downstream of induced-proximity activation of caspase-8. Here we show that loss of B1 reduces the association of pro-caspase-8 with microtubules and subsequently leads to impaired PTX-induced activation of apoptosis. Taken together, our data show that B1 regulates indirectly endogenous microtubule dynamics and stability while its loss leads to microtubules that are more dynamic and less susceptible to PTX-induced stabilization conferring taxane resistance.

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Acknowledgements

We would like to thank Dr Geri Kreitzer (Weill Cornell Medical College), Dr Mary Ann Jordan (University of California, Santa Barbara) and Dr Richard Baer (Columbia University) for their kind gifts. We would also like to thank Dr Siddhartha Sen for help with the flow cytometry experiments. Work by Matthew Sung was supported by the National Institutes of Health (T32 CA062948, RO1 CA137020, NCI U54 CA143876).

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  1. Division of Hematology and Medical Oncology, Department of Medicine, Weill Medical College of Cornell University, New York, NY, USA

    M Sung & P Giannakakou

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Correspondence to P Giannakakou.

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Sung, M., Giannakakou, P. BRCA1 regulates microtubule dynamics and taxane-induced apoptotic cell signaling. Oncogene 33, 1418–1428 (2014). https://doi.org/10.1038/onc.2013.85

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