APC/C^FZR-1Controls 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 co-activator of the anaphase promoting complex/cyclosome (APC/C), an E3 ubiquitin ligase, functions as a negative regulator of centrosome duplication in the Caenorhabditis 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 Caenorhabditis elegans.

APC/CFZR-1Controls ZYG-1 Levels to Regulate Centrosome Assembly

“…Thus, loss of FZR-1 leads to increased levels of ZYG-1 at centrosomes, which should compensate for impaired ZYG-1 function and rescue the zyg-1 mutant phenotypes observed previously (Kemp et al, 2007, Medley et al, 2017a. These results support a hypothesis that ZYG-1 levels are regulated by APC/C FZR-1 -dependent proteasomal degradation.…”

“…Prior study identified SAS-5 as a substrate of APC/C FZR-1 , and implicated that APC/C FZR-1 might target additional centrosome regulators in C. elegans embryos (Medley et al, 2017a). The APC/C coactivator, FZR-1, was identified as a genetic suppressor of ZYG-1 since loss of fzr-1 restores embryonic viability and centrosome duplication to the zyg-1(it25) mutant (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 a substrate of APC/C FZR-1 .…”

“…Since the C. elegans SAS-5 is an APC/C FZR-1 substrate and mutating the SAS-5:KENbox leads to the zyg-1 suppression (Medley et al, 2017a), we asked if mutating the ZYG-1:D-box3 and SAS-5:KEN-box degrons simultaneously produces the effects comparable to those resulting from the fzr-1 mutation. Instead, mutating both ZYG-1 and SAS-5 degrons leads to the zyg-1 suppression at a level nearly equivalent to the ZYG-1:DB3(2A) mutation alone (Fig.…”

“…The ubiquitin-proteasome system provides a key mechanism to control centrosome protein levels (Nakayama and Nakayama, 2006). Levels of core centrosome factors are shown to be regulated by proteasomal degradation through E3 ubiquitin ligases, including the Anaphase Promoting Complex/Cyclosome (APC/C) and the SKP1-CUL1-F-box-protein (SCF), and their regulatory mechanisms appear to be conserved (Arquint et al, 2012;Cunha-Ferreira et al, 2009;Guderian et al, 2010;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 factors. In C. elegans, FZR-1 (Cdh1 in human), a coactivator of APC/C, negatively influences centrosome assembly, and APC/C FZR-1 -dependent proteolysis regulates SAS-5 levels through a KEN-box motif (Kemp et al, 2007;Medley et al, 2017a), which is conserved in humans and flies where APC/C Cdh1/Fzr targets core 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). In this study, we investigated ZYG-1 as a potential substrate of APC/C FZR-1 , and how two E3 ubiquitin ligases APC/C FZR-1 and SCF Slimb/bTrCP influence ZYG-1 levels in centrosome assembly.…”