APC/CFZR-1 Controls SAS-5 Levels To Regulate Centrosome Duplication inCaenorhabditis elegans

Abstract

As the primary microtubule-organizing center, centrosomes play a key role in establishing mitotic bipolar spindles that secure correct transmission of genomic content. For the fidelity of cell division, centrosome number must be strictly controlled by duplicating only once per cell cycle. Proper levels of centrosome proteins are shown to be critical for normal centrosome number and function. Overexpressing core centrosome factors leads to extra centrosomes, while depleting these factors results in centrosome duplication failure. In this regard, protein turnover by the ubiquitin-proteasome system provides a vital mechanism for the regulation of centrosome protein levels. Here, we report that FZR-1, the Caenorhabditis elegans homolog of Cdh1/Hct1/Fzr, a coactivator of the anaphase promoting complex/cyclosome (APC/C), an E3 ubiquitin ligase, functions as a negative regulator of centrosome duplication in the C. elegans embryo. During mitotic cell division in the early embryo, FZR-1 is associated with centrosomes and enriched at nuclei. Loss of fzr-1 function restores centrosome duplication and embryonic viability to the hypomorphic zyg-1(it25) mutant, in part, through elevated levels of SAS-5 at centrosomes. Our data suggest that the APC/CFZR-1 regulates SAS-5 levels by directly recognizing the conserved KEN-box motif, contributing to proper centrosome duplication. Together, our work shows that FZR-1 plays a conserved role in regulating centrosome duplication in C. elegans.

APC/CFZR-1 regulates centrosomal ZYG-1 to limit centrosome number

“…At anaphase, the ZYG-1 DB3(2A) mutation leads to elevated centrosomal SAS-5 (1.26±0.39 fold; mean±s.d.) compared to WT controls (1.00 ±0.27), which is nearly equivalent to the increase in SAS-5 KEN(3A) mutants (1.26±0.37 fold) where APC/C FZR-1 -mediated degradation of SAS-5 is blocked (Medley et al, 2017a). However, centrosomal SAS-5 levels are decreased in ZYG-1 DB2(2A) mutants (0.70±0.19 fold).…”

“…Prior study has identified SAS-5 as an APC/C FZR-1 substrate, and implicated APC/C FZR-1 as targeting additional centrosome regulators in C. elegans (Medley et al, 2017a). FZR-1 was identified as a genetic suppressor of ZYG-1 since loss of FZR-1 restores embryonic viability and centrosome duplication to zyg-1(it25) mutants (Kemp et al, 2007;Medley et al, 2017a). Given the strong genetic interaction between zyg-1 and fzr-1, a compelling possibility is that ZYG-1 might be an APC/C FZR-1 substrate.…”

“…The ubiquitin-proteasome system provides a key mechanism to control centrosome protein levels (Nakayama and Nakayama, 2006). Levels of centrosome factors are regulated by proteasomal degradation through E3 ubiquitin ligases, including the anaphase promoting complex/cyclosome (APC/C) and SKP1-CUL1-F-box-protein complex (SCF), and their regulatory mechanisms appear to be conserved (Arquint et al, 2012;Cunha-Ferreira et al, 2009;Holland et al, 2010;Medley et al, 2017a;Meghini et al, 2016;Peel et al, 2012;Puklowski et al, 2011;Rogers et al, 2009;Strnad et al, 2007;Tang et al, 2009). In C. elegans, the kinase ZYG-1 is a key centrosome regulator and ZYG-1 levels are critical for normal centrosome number and function (O'Connell et al, 2001;Song et al, 2008).…”

“…Another E3 ubiquitin ligase, APC/C, also regulates levels of centrosome regulators. In C. elegans, FZR-1 (Cdh1 in humans), a coactivator of APC/C, negatively influences centrosome assembly, and together, denoted APC/C FZR-1 , regulate SAS-5 levels through a KEN box (Kemp et al, 2007;Medley et al, 2017a), which is the conserved motif in humans and flies where APC/C Cdh1/Fzr targets centrosome factors (human STIL/SAS-5, SAS-6, CPAP/SAS-4 and Drosophila Spd2) via KEN-box (Arquint and Nigg, 2014;Meghini et al, 2016;Strnad et al, 2007;Tang et al, 2009). Here, we investigate ZYG-1 as a potential APC/C FZR-1 substrate, and how the E3 ubiquitin ligases APC/C FZR-1 and SCF Slimb/βTrCP influence ZYG-1 levels during centrosome assembly.…”

Mitotic Cell Division in Caenorhabditis elegans

“…Finally, negative regulation of SAS-5 levels also is important for the control of centriole duplication. The same screen that led to the identification of a role for PP1 in limiting ZYG-1 levels also recovered two loss-of-function alleles of the FZR-1/Cdh1 subunit of the anaphase promoting complex/cyclosome (APC/C) E3 ligase as suppressors of a TS zyg-1 centriole duplication defect (Medley et al 2017a). Further investigation showed that FZR-1/Cdh1 and other APC/C subunits are required to limit SAS-5 levels ( Figure 3B).…”