Loss of the mammalian APC/C activator FZR1 shortens G1 and lengthens S phase but has little effect on exit from mitosis

Abstract

The anaphase-promoting complex/cyclosome (APC/C) is essential for progression through mitosis. At anaphase onset, the APC/C requires the activator protein CDC20 to target securin and cyclin B1 for proteasome-dependent degradation, but then depends on the CDC20-related protein FZR1 (also known as CDH1) to remain active until the onset of the next S phase. To investigate the role of FZR1 in mammalian cells, we used RNAi in human cell lines and conditional gene targeting in mouse embryonic fibroblasts. In neither case was FZR1 required for exit from mitosis, but in cells lacking FZR1, the G1 phase was shortened and the S phase was prolonged. In several normal and transformed human cell lines, loss of FZR1 function induced DNA-damage responses and impaired proliferation independently of the p53 status. Constitutive knockdown of p53 in U2OS cells with inducible FZR1 siRNA also failed to restore their proliferative capacity. Thus, the proliferation defects are a direct consequence of the genetic damage inflicted by loss of FZR1 function and are largely independent of p53. In summary, mammalian FZR1 is not required for the completion of mitosis, but is an important regulator of G1 phase and is required for efficient DNA replication in human and mouse somatic cells.

APC/C is an essential regulator of centrosome clustering

“…Although CDH1 is known to target for degradation, a wide range of mitotic proteins including motor kinesins, kinases and CDC20 itself 8,24 , in our knowledge there is no mitotic phenotype associated with APC/C-CDH1 inhibition. However, inhibition of APC/C-CDH1 is suggested to impair DNA damage response to genotoxic stress 38 , shorten G1 and prolong S phase 39 , induce replicative stress 40 and impair cell motility by reducing stress fibre formation 41 . Moreover, it has been proposed that in a therapeutic approach that involves targeting APC/C, inhibition of APC/C-CDH1, although not highly cytotoxic, may have an adjuvant role to APC/C-CDC20 inhibition 40 .…”

Spatiotemporal organization of Aurora-B by APC/CCdh1 after mitosis coordinates cell spreading via FHOD1

“…Cdh1 exerts spatial control on AurB kinase activity at the end of mitosis Previous studies have indicated that, at the end of mitosis, AurB disappears from the cell later than other APC/C substrates (Floyd et al, 2008;Lindon and Pines, 2004;Sigl et al, 2009), and indeed in vivo time-lapse analysis of AurB-GFP degradation reveals Cdh1-dependent proteolysis of AurB continuing over a window of time that extends well into G1 phase (C.M., M.M. and C.L., unpublished data).…”

The APC/C activator FZR1 coordinates the timing of meiotic resumption during prophase I arrest in mammalian oocytes

“…A more detailed analysis of the potential targets of APC/C FZR1 has to be set against the small numbers of oocytes that can be collected for analysis. There is much interest in what constitutes the physiological set of FZR1 substrates, with contrasting observations on what is stabilized following Fzr1 knockdown or knockout (Floyd et al, 2008;Garcia-Higuera et al, 2008;Li et al, 2008;Sigl et al, 2009). Although these other studies have failed to detect any differences in the levels of cyclin B1 following loss of FZR1, it should be noted that they were performed on mitotically dividing cells that do have the capacity to degrade cyclin B1 through APC/C CDC20 activity.…”

“…Low CDK1 activity promotes APC/C FZR1 activation, such that APC/C CDC20 activity is replaced by APC/C FZR1 in late mitosis/G1. Recent somatic cell knockout studies have implicated FZR1 in G1/S phase timing, DNA replication and, consequently, genomic stability (Garcia-Higuera et al, 2008;Sigl et al, 2009), in addition to roles in the differentiation of various cell lineages including neurons (for a review, see Wasch et al, 2010). The recent identification of APC/C FZR1 activity in vitro during prophase I of the oocyte therefore represents a temporally unique role for this APC/C co-activator.…”