Mad1 Serine-214 Phosphorylation Is Essential For The Spindle Assembly Checkpoint

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL The spindle assembly checkpoint (SAC), which blocks anaphase onset until all chromosomes have bioriented, is one of the key self-monitoring systems of the eukaryotic cell cycle. The mitotic arrest-deficient protein 1 (Mad1), a critical component of the SAC, is hyperphosphorylated in mitosis. However, the kinases responsible for Mad1 phosphorylation and its functional significance are not fully understood. Here we report that Mad1 is phosphorylated on Serine-214 in vivo and in vitro by the ATM kinase, a critical DNA damage response protein also activated in mitosis and required for the SAC. This phosphorylation facilitates the homodimerization of Mad1 and heterodimerzation with Mad2. Cells expressing the mutation of Serine-214 to Alanine show a defective SAC and shorter mitotic timing. Introduction of the Serine-214 to glutamic acid mutant Mad1 partially complements the SAC defect in ATM-deficient cells. Immunofluorescence experiments show that Mad1-Ser214 phosphorylation has a similar localization pattern to ATM Serine-1403 phosphorylation during mitosis. In clinical samples, we find that Mad1 Serine-214 phosphorylation is highly expressed in malignant breast cancer tissues and is significantly correlated with the mitotic index (p<0.001, Pearson Correlation Test), indicating the physiological significance of Mad1 Serine-214 phosphorylation. Together, these findings reveal an important role of ATM-mediated Mad1 phosphorylation in mitosis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3058. doi:1538-7445.AM2012-3058