Distinct D-box Motifs in SPD-2 Mediate APC/C FZR-1 -Dependent Degradation and Centrosomal Localization in Caenorhabditis elegans Embryos

Abstract Centrosome duplication must be tightly regulated to maintain genomic stability. In Caenorhabditis elegans, the APC/C and co-activator FZR-1 function as negative regulators of centrosome duplication by targeting specific substrates for proteolytic degradation. While C. elegans SAS-5 and ZYG-1 have been identified as substrates of APC/CFZR-1, the mechanism by which APC/CFZR-1-dependent degradation influences centrosome assembly remains unclear. Here, we identified SPD-2, the conserved homolog of human CEP192, as a substrate of APC/CFZR-1. We show that loss of APC/CFZR-1 increases both cellular and centrosomal SPD-2 levels, and that SPD-2 physically associates with FZR-1 in vivo. Functional analyses of canonical D-box motifs reveal that D-box1, D-box2, and D-box3 each contribute to SPD-2 degradation, each with different functional consequences. Mutation of D-box3 alone partially rescued zyg-1 mutant phenotypes by restoring centrosome duplication and embryonic viability through increased centrosomal SPD-2 and ZYG-1. In contrast, mutating D-box1 or D-box2 elevated cellular SPD-2 but did not rescue zyg-1, with the D-box1 mutation further reducing centrosomal SPD-2 and exacerbating duplication defects and lethality in zyg-1 mutants. Our results reveal a conserved mechanism for APC/CFZR-1-dependent degradation of SPD-2 and show that its degron motifs have dual functions in degradation and centrosomal localization, ensuring robust control of centrosome assembly during C. elegans embryogenesis. Competing Interest Statement The authors have declared no competing interest. Footnotes The results and discussion section, as well as the figure legends, have been revised in this revision.