When neural progenitor cells (NPCs) fail to divide correctly at the right time and location, neurodevelopmental disorders can occur, including gross abnormalities in the size and structure of the brain and more subtle defects in neuronal layering and connectivity. One result of impaired NPC division in the cerebral cortex is microcephaly. The majority of the genetic causes of microcephaly in humans are due to mutations that affect parameters of mitosis in NPCs, but recently defects in cytokinesis have been implicated as well. A novel model of microcephaly studied here, resulting from the loss of the kinesin-6 family member Kif20b, has defects specifically in abscission, the last step in the cytokinesis phase of NPC division. Kif20b protein is detected in the midbodies of NPCs, a structure that connects the two daughter cells' apical membranes until mediating abscission. In Kif02b-/- cortex as well as in Kif20b-/- dissociated NPCs, midbody abnormalities, signifying an abnormal abscission process, and apoptosis have been observed. A common mechanism in the pathogenesis of many genetic and viral causes of microcephaly is p53-dependent apoptosis. However, it is not known whether p53 is activated to cause apoptosis in response to defective abscission in any cell type. To test whether apoptosis in Kif20b-/- mice is p53 dependent, we created mice double mutant for Kif20b and p53. New preliminary data shows that the survival, apoptosis and decreased cortical thickness, at least at early ages, in Kif20b-/- mice are rescued by p53 co-deletion. However, the relationship between midbody defects and p53 activation in Kif20b-/- NPCs is unknown. Additionally, the importance of abscission regulation to produce a cortex of normal size and structure, apart from apoptosis prevention, is unclear. Correct midbody alignment is important for epithelial structure in other organisms, and midbody inheritance has been shown to influence cell fate, but the functional relevance of these observations for cortical development have not been demonstrated. The survival of double mutant mice for many months past birth provides an opportunity to examine the consequences of abnormal abscission for postnatal cortical development. My specific hypothesis is that p53 is activated in response to impaired abscission in Kif20b-/- NPCs to cause apoptosis, and that this response is important to prevent abnormalities in cortical size and structure. To test this hypothesis, I propose to study Kif20b-/- p53-/- mice in vivo and Kif20b-/- NPCs cultured in vitro with and without inhibition of p53. With these techniques and using fixed and live imaging experiments, I will complete the following aims:
Aim 1) Determine the relationship between abscission defects and p53-dependent apoptosis in Kif20b-/- mice.
Aim 2) Investigate the consequence of Kif20b loss for NPCs when p53 is co-deleted. The proposed research is significant in that it will bridge the gap in our knowledge of how apoptosis is regulated in response to abscission defects and whether this is mediated by p53. Additionally, the research will provide insight on normal abscission in NPCs and the consequences for its perturbation in vivo and in vitro.
The proposed research is relevant to public health because it will contribute to our understanding of how errors in neural progenitor cell division can lead to improper brain development, resulting in neurodevelopmental disorders such as microcephaly, brain malformations and/or intellectual disability. Additionally, this research will expand our fundamental knowledge of mechanisms of cytokinesis and p53 activation in neural stem cells and the role of the vertebrate kinesin Kif20b in the cortical neuroepithelium. This knowledge will be informative to our understanding of common and divergent mechanisms in the development of other epithelia and organ systems.
