Functional Study of Cdk12 in Neural Development and Axonal Growth

• Main Authors: Hong-Ru Chen, 陳虹如
• Other Authors: Ming-Ji Fann
• Format: Others
• Language: en_US
• Published: 2015
• Online Access: http://ndltd.ncl.edu.tw/handle/24163789841443830359

博士 === 國立陽明大學 === 生命科學系暨基因體科學研究所 === 103 === Neural development proceeds sequentially, including the generation and differentiation of neurons from precursor cells, the migration of immature neurons from their birthplaces to their final positions, outgrowth of axons and dendrites from neurons. Cdk12 is a protein kinase with abundant expression in the nervous system. In order to characterize what the physiological function of Cdk12 is during neural development, I employ (1) P19 cells and cultured cortical neurons as an in vitro assay system, and (2) Cdk12 conditional knockout mice as an in vivo model. I explored the roles of Cdk12 in neuronal differentiation by using P19 cells, which are mouse embryonic carcinoma cells and are able to differentiate into neurons when there is overexpression of the proneural gene Ascl1. Upon knockdown of Cdk12, no effect on differentiated cell numbers was detected, but a substantial decrease of numbers of neurons with long neurites was identified. Similarly, knockdown of Cdk12 in primarily cultured cortical neurons shortens the averaged axonal length. Depletion of Cdk12 in P19 cells significantly reduces Cdk5 expression. Overexpression of Cdk5 protein in P19 cells was able to partially rescue the neurite outgrowth defect observed when Cdk12 is depleted. Together, I conclude that Cdk12 regulates axonal elongation through modulating Cdk5 expression. To define the role of Cdk12 during neural development in vivo, the Nestin-Cdk12-cKO mice were used. Nestin-Cdk12-cKO mice showed microcephaly, reduced thickness of the cortical layers, and a loss of the corpus callosum in cerebral cortex. The reduced neuronal numbers in the neocortex results from apoptosis of newborn progenitors, which is due to down-regulation of DDR genes that are important for DNA repair during DNA replication. Although cortical layer organization is largely preserved in Nestin-Cdk12-cKO mice, CUX+ cells that normally locate in layer II-IV are misplaced in layer II-VI evenly, suggesting that deficiency of Cdk12 also leads to abnormally of neuronal migration. To further investigate neuronal migration after deletion of Cdk12, I examined the migration defect following deletion of Cdk12 by in utero electroporation of Cre-expression plasmids in to Cdk12f/f mice between embryonic day 14 ~ 16 (E14-16). Cdk12del/del neurons showed dramatic migration defects from E15, which are due to the morphological transition defects of late-born migrating cells. In summary, Cdk12’s function is crucial for several aspects of neuronal development, including neurogenesis, late-born neuron migration, and axon growth.