The role of a mitotic gene expression for the function of the mitotic spindle assembly checkpoint
Final Report Abstract
In our previous w/ork, we found surprisingly that potent antibiotic inhibitors for transcription (e.g. actinomycin-D) and translation (e.g. cycloheximide, puromycin or hygromycin) can efficiently override a spindle assembly checkpoint (SAC) mediated mitotic arrest after the induction of spindle damage. Therefore, we suggested that transcription and/or translation might be required for the maintenance of the SAC, a hypothesis that was successfully investigated in detail in our laboratory during the funding period by one Ph.D student funded by the DFG. First, we verified our preliminary results and showed that treatment of mitotic cells with actinomycin-D indeed overrides the SAC and induces a premature exit from mitosis. The effect of actinomycin-D, however, is not due to inhibition of gene transcription, but rather due to its so far unrecognized activity to mislocalize components of the chromosomal passenger complex (CPC) from centromeres onto chromosomes arms. We used this unique activity of actinomycin-D to demonstrate that a centromeric localization of the mitotic Aurora-B kinase, which is part of the CPC, is essential for the maintenance of the SAC. Thus, this result supports the view that the CPC actively contributes to the function of the SAC and was published. We further verified our preliminary results by demonstrating that treatment with translation inhibitors including cyloheximide, puromycin and hygromycin also efficiently overrides the SAC after spindle damage. By performing 35S-labeling experiments with mitotic cells we found that cells in mitosis retain considerable mRNA translation and protein synthesis. Moreover, we obtained evidence that the majority of these mRNAs are translated in a 5'-capindependent manner during mitosis. By analyzing candidates that might participate in the SAC we found that cyclin A, TPX2, Bub1 and Cdc20 are indeed continuously synthesized during a SAC mediated mitotic arrest. In future studies we are further evaluating the physiological relevance of this remarkable finding, which suggest that a subset of stable mRNAs need to be translated during a prolonged mitotic delay in order to maintain an activated SAC.
Publications
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2007. Mechanisms of mitotic cell death induced by chemotherapy-mediated G2 checkpoint abrogation. Cancer Research 67: 339 - 345
Vogel, C., Hager, C., Bastians, H.
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2009. Determinants for the efficiency of anticancer drugs targeting either Aurora-A or Aurora-B kinases in human colon carcinoma cells. Molecular Cancer Therapeutics 8: 2046 - 2056
Kaestner, P., Stolz, A., Bastians, H.
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2009. Pharmacologic abrogation of the mitotic spindle checkpoint by an indolocarbazole discovered by cellular screening efficiently kills cancer cells. Cancer Research 69: 3874 - 3883
Stolz, A., Vogel, C., Schneider, V., Ertych, N., Kienitz, A., Yu, H., Bastians, H.
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2010. Centromere localization of INCENP-Aurora B is sufficient to support spindle checkpoint function. Cell Cycle, 9:1360 - 1372
Becker, M., Stolz, A., Ertych, N., Bastians, H.
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2010. The CHK2-BRCA1 tumour suppressor pathway ensures chromosomal stability in human somatic cells. Nature Cell Biology, 12: 492 - 499
Stolz, A., Ertych, N., Kienitz, A., Vogel, C., Schneider, V., Fritz, B., Jacob, R., Dittmar, G., Weichert, W., Petersen, I., Bastians, H.
