Regulation of spindle assembly checkpoint signalling and silencing during mitosis

Abstract

The development, growth and maintenance of a healthy body depends on structural replenishment and renewal of cells, the units of which our body is composed. Through cell division or mitosis, one mother cell splits into two identical daughter cells. Hereditary material, known as sister chromatids, are segregated by microtubule-bundles which are part of the mitotic spindle. Errors in sister chromatid segregation can contribute or cause diseases such as cancer or have a negative impact on the growth and development of an organism. Two essential mechanisms, the error correction pathway and the spindle assembly checkpoint (SAC), warrant the correct segregation of sister chromatids during mitosis. The enzyme MPS1 activates the spindle assembly checkpoint and plays an important role in the regulation of the error correction pathway. Work described in this dissertation advances our understanding of the fundamental workings of the SAC and the regulation of the upstream SAC-activator MPS1. We investigated the fundamental workings of the SAC-mechanism and showed that the formation of attachments to sister chromatid kinetochores is sufficient to satisfy the SAC and not the pulling forces exerted by microtubule-bundles. Our work settled a longstanding debate in the field and directed subsequent studies to investigate the function of pulling forces that kinetochores endure. Multiple molecular events contribute to silencing the SAC and we showed that the removal of MAD1 from kinetochores is the critical molecular step. Relocating MAD1 to kinetochores is sufficient to reactivate the SAC-signal and this suggested that the upstream SAC-activator MPS1 remains active on kinetochores with correct kinetochore-microtubule attachments. We developed a tool to monitor MPS1 activity at defined locations within single cells at high temporal resolution and showed that indeed MPS1 is not fully switched off. Our work uncovered new insights and regulatory mechanisms that orchestrate the activation of MPS1 and modulate its activity during mitosis.