Site-Specific Phosphorylation of ZYG-1 Regulates ZYG-1 Stability and Centrosome Number

Abstract

Spindle bipolarity is critical for genomic integrity. Given that centrosome number often dictates mitotic bipolarity, tight control of centrosome assembly is vital for the fidelity of cell division. The kinase ZYG-1/Plk4 is a master centrosome factor that is integral for controlling centrosome number and is modulated by protein phosphorylation. While autophosphorylation of Plk4 has been extensively studied in other systems, the mechanism of ZYG-1 phosphorylation in C. elegans remains largely unknown. In C. elegans, Casein Kinase II (CK2) negatively regulates centrosome duplication through control of centrosome-associated ZYG-1 levels. In this study, we investigated ZYG-1 as a potential substrate of CK2 and the functional impact of ZYG-1 phosphorylation on centrosome assembly. First, we show that CK2 directly phosphorylates ZYG-1 in vitro and physically interacts with ZYG-1 in vivo. Intriguingly, depleting CK2 or blocking ZYG-1 phosphorylation at putative CK2 target sites leads to centrosome amplification. In the non-phosphorylatable (NP)-ZYG-1 mutant embryo, total ZYG-1 levels are increased, leading to elevated centrosomal ZYG-1 and downstream factors, providing a possible mechanism of the NP-ZYG-1 mutation to drive centrosome amplification. Moreover, inhibiting the 26S proteasome blocks degradation of the phospho-mimetic (PM)-ZYG-1, whereas the NP-ZYG-1 is partially resistant to proteasomal degradation. Our data suggest that site-specific phosphorylation of ZYG-1, in part by CK2, regulates ZYG-1 levels through proteasomal degradation to limit centrosome number. We provide a mechanism linking CK2 kinase activity to centrosome duplication through direct phosphorylation of ZYG-1, which is critical for the integrity of centrosome number.