ASPM and CITK regulate spindle orientation by affecting the dynamics of astral microtubules

Abstract

Correct orientation of cell division is considered an important factor for the achievement of normal brain size, as mutations in genes that affect this process are among the leading causes of microcephaly. Abnormal spindle orientation is associated with reduction of the neuronal progenitor symmetric divisions, premature cell cycle exit, and reduced neurogenesis. This mechanism has been involved in microcephaly resulting from mutation of ASPM, the most frequently affected gene in autosomal recessive human primary microcephaly (MCPH), but it is presently unknown how ASPM regulates spindle orientation. In this report, we show that ASPM may control spindle positioning by interacting with citron kinase (CITK), a protein whose loss is also responsible for severe microcephaly in mammals. We show that the absence of CITK leads to abnormal spindle orientation in mammals and insects. In mouse cortical development, this phenotype correlates with increased production of basal progenitors. ASPM is required to recruit CITK at the spindle, and CITK overexpression rescues ASPM phenotype. ASPM and CITK affect the organization of astral microtubules (MT), and low doses of MT-stabilizing drug revert the spindle orientation phenotype produced by their knockdown. Finally, CITK regulates both astral-MT nucleation and stability. Our results provide a functional link between two established microcephaly proteins.

Tissue specific vulnerability to mitotic defects caused by mutations in theDrosophilaASPM homologue, Asp

“…These data demonstrate tissue specific susceptibility to disrupted metaphase spindle formation. In mammals, the loss of ASPM function has been thought to specifically impede neurogenesis because the specialized cellular morphology of neural progenitors renders them extremely susceptible to alterations in spindle orientation (Fish et al 2006;Gai et al 2016). However, two recent studies have provided evidence uncoupling spindle orientation from the microcephaly phenotype, pointing instead to apoptosis (Insolera et al 2014) or impaired centriole duplication (Jayaraman et al 2016).…”

E2F-2 Promotes Nuclear Condensation and Enucleation of Terminally Differentiated Erythroblasts

“…Using RNA sequencing to compare the transcriptomes of E2F-2 knockout and wild-type progenitors and differentiated erythroblasts, we found E2F-2-dependent erythroid induction of CRIK, a mitotic kinase known to interact with Rho family GTPases, which are linked to the control of enucleation (31,37). In mitotic cells, CRIK regulates astral microtubule length and spindle orientation (36), and during cytokinesis, CRIK colocalizes with RhoA and mediates contractile function at the cleavage furrow (32,33,35). Previous studies of erythroid enucleation describe a critical role for Rac GTPases in organizing CAR formation and lipid raft clusters between the pyrenocyte and incipient reticulocyte (37).…”

“…To determine the mechanism by which E2F-2 regulates nuclear condensation and subsequent erythroid enucleation, we focused on genes deregulated in E2F-2 knockout differentiated erythroid cells that were associated with chromosome organization (see Table S1 in the supplemental material). We found that citron Rho-interacting kinase (CRIK), which plays an established role in late mitosis and cytokinesis (32)(33)(34)(35)(36), is induced over the course of differentiation from wild-type progenitors to orthochromatic erythroblasts. Notably, Cit gene expression decreases in E2F-2 knockout erythroid cells (Fig.…”

Regulation of cytokinesis during corticogenesis: focus on the midbody

“…Loss of CitK results in widespread apoptosis and the presence of [168,169] many binucleated and multiciliated neurons throughout the brain [81][82][83]. RGCs in CitK-null mice have randomized spindle poles, increased cell cycle exit, and elevated production of basal progenitors, indicating a higher rate of differentiative divisions [84]. Increased DNA damage and activation of apoptosis independent of cytokinesis failure have been reported in CitK-null mice, presenting an additional mechanism for the widespread apoptosis seen in CitK-null brains [85].…”