PLK1-mediated phosphorylation of WDR62/MCPH2 ensures proper mitotic spindle orientation

Abstract

Primary microcephaly (MCPH) is an autosomal recessive disorder characterized by congenital reduction of head circumference. Here, we identified compound heterozygous mutations c.731 C > T (p.Ser 244 Leu) and c.2413 G > T (p.Glu 805 X) in the WDR62/MCPH2 gene, which encodes the mitotic centrosomal protein WDR62, in two siblings in a Japanese family with microcephaly using whole-exome sequencing. However, the molecular and cellular pathology of microcephaly caused by WDR62/MCPH2 mutation remains unclear. To clarify the physiological role of WDR62, we used the CRISPR/Cas9 system and single-stranded oligonucleotides as a point-mutation-targeting donor to generate human cell lines with knock-in of WDR62/MCPH2 c.731 C > T (p.Ser 244 Leu) missense mutation. In normal metaphase, the mitotic spindle forms parallel to the substratum to ensure symmetric cell division, while WDR62/MCPH2-mutated cells exhibited a randomized spindle orientation caused by the impaired astral microtubule assembly. It was shown that a mitotic kinase, Polo-like kinase 1 (PLK1), is required for the maintenance of spindle orientation through astral microtubule development. In this study, we demonstrated that WDR62 is a PLK1 substrate that is phosphorylated at Ser 897, and that this phosphorylation at the spindle poles promotes astral microtubule assembly to stabilize spindle orientation. Our findings provide insights into the role of the PLK1-WDR62 pathway in the maintenance of proper spindle orientation.

A novel WDR62 missense mutation in microcephaly with abnormal cortical architecture and review of the literature

“…Over 40 pathogenic mutations in WDR62 have already been published. In addition to microcephaly, a wide range of cortical malformations was also described in these patients (Bacino et al 2012;Banerjee et al 2016;Bastaki et al 2016;Bhat et al 2011;Farag et al 2013;Sajid Hussain et al 2013;Kousar et al 2011;McDonell et al 2014;Memon et al 2013;Miyamoto et al 2017;Murdock et al 2011;Najmabadi et al 2011;Nardello et al 2018;Naseer et al 2017;Poulton et al 2014;Rupp et al 2014;Wang et al 2017).…”

The Yin and Yang of Autosomal Recessive Primary Microcephaly Genes: Insights from Neurogenesis and Carcinogenesis

“…Mitotic spindle orientation [43] Cell cycle (gastric cancer, GC) [44] Regulating intermediate neural and glial progenitors [45] Centrosome amplification (ovarian cancer, OC) [46] Cell cycle, centriole biogenesis, and mitotic spindle orientation [17] Cell growth (lung adenocarcinoma) [34] Regulating neural stem cell division [47,48]…”

“…MCPH1 (BRIT1) CHK1, CDC25, CDK1 [16] SLUG, CDK1, p53, CDH1, MDM2, SNAIL [42] p53 [110] CyclinA2, CDK2, CDC25C-cyclinB, CDC2, p53, BCL-2, Bax, Cytochrome c, Caspase-3, PARP-1 [40] MCPH2 (WDR62) JNK [18] N/A AURKA [119] PLK1 [43] MCPH3 (CDK5RAP2) p35 [120] N/A MST1, TAZ (Hippo) [121] MCPH4 (CASC5) BUB1, BUBR, ZWINT-1 [122] BUB1 [52] miR-193b-3p [27] p53 [123] MCPH5 (ASPM) The p53 gene (TP53) is classified as a tumor suppressor gene, and upregulation of its activity led to neural progenitor apoptosis in MCPH gene-deficient mice. Deletion of Mcph5 increases DNA damage and induces postnatal cerebellar progenitor apoptosis that depends on p53 [114].…”

JNK-mediated spindle reorientation in stem cells promotes dysplasia in the aging intestine

“…However, the magnitude of the effect of JNK loss on the association of Wdr62 with the spindle was less compared to the effect of Wdr62 loss on pJNK localization. The more modest decrease may be indicative of a role for other kinases that have been reported to recruit Wdr62 to the centrosome, including Aurora A and Polo-like kinase Lim et al, 2016;Miyamoto et al, 2017;Ramdas Nair et al, 2016). Nonetheless, JNK seems to play a role in stabilizing Wdr62 at the spindle, as JNK over-activation was sufficient to increase Wdr62 affinity with the spindle.…”