Probing the in vivo function of Mad1:C-Mad2 in the spindle assembly checkpoint |
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Authors: | Fava Luca L Kaulich Manuel Nigg Erich A Santamaria Anna |
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Institution: | Growth and Development, Biozentrum, University of Basel, Basel, Switzerland. |
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Abstract: | The spindle assembly checkpoint (SAC) restrains anaphase until all chromosomes become bi-oriented on the mitotic spindle. The SAC protein Mad2 can fold into two distinct conformers, open (O) and closed (C), and can asymmetrically dimerize. Here, we describe a monoclonal antibody that specifically recognizes the dimerization interface of C-Mad2. This antibody revealed several conformation-specific features of Mad2 in human cells. Notably, we show that Mad2 requires association with Mad1 to adopt the closed conformation and that the activity of the Mad1:C-Mad2 complex undergoes regulation by p31comet-dependent 'capping'. Furthermore, C-Mad2 antibody microinjection caused an abrupt termination of the SAC and accelerated mitotic progression. Remarkably, microinjection of a Mad1-neutralizing antibody triggered a comparable mitotic acceleration. Our study provides direct in vivo evidence for the model that a kinetochore complex of Mad1:C-Mad2 acts as a template to sustain the SAC and it challenges the distinction between SAC and mitotic timer. |
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Keywords: | closed‐Mad2 Mad2 template model mitotic timing p31comet spindle assembly checkpoint |
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