ArabidopsisWEE1 Kinase Controls Cell Cycle Arrest in Response to Activation of the DNA Integrity Checkpoint

Abstract

Upon the incidence of DNA stress, the ataxia telangiectasia-mutated (ATM) and Rad3-related (ATR) signaling kinases activate a transient cell cycle arrest that allows cells to repair DNA before proceeding into mitosis. Although the ATM-ATR pathway is highly conserved over species, the mechanisms by which plant cells stop their cell cycle in response to the loss of genome integrity are unclear. We demonstrate that the cell cycle regulatory WEE1 kinase gene of Arabidopsis thaliana is transcriptionally activated upon the cessation of DNA replication or DNA damage in an ATR-or ATM-dependent manner, respectively. In accordance with a role for WEE1 in DNA stress signaling, WEE1-deficient plants showed no obvious cell division or endoreduplication phenotype when grown under nonstress conditions but were hypersensitive to agents that impair DNA replication. Induced WEE1 expression inhibited plant growth by arresting dividing cells in the G2-phase of the cell cycle. We conclude that the plant WEE1 gene is not rate-limiting for cycle progression under normal growth conditions but is a critical target of the ATR-ATM signaling cascades that inhibit the cell cycle upon activation of the DNA integrity checkpoints, coupling mitosis to DNA repair in cells that suffer DNA damage.

The plant‐specific CDKB 1‐ CYCB 1 complex mediates homologous recombination repair in Arabidopsis

“…For this analysis, wee1 was used as a positive control and, consistent with previous data, was found to be highly sensitive to HU, whereas it shows no growth abnormalities on control medium (De Schutter et al , 2007; Cools et al , 2011). In contrast, root growth on HU of all tested cycb1 mutants was comparable to the growth of wild‐type plants (Fig 1C and D).…”

“…T‐DNA insertion line cycb1;3 (pst15850) was obtained from the RIKEN collection (Yokohama, Japan) and cycb1;4 from the GABI‐KAT collection (Bielefeld, Germany) (Kleinboelting et al , 2012). The ku70 (Riha et al , 2002), wee1 (De Schutter et al , 2007), and rad51 (Li et al , 2004) mutants were described previously. The reporter line uidA/IC9C was used as described in Molinier et al (2004) and the pCDKB1;1:GUS line as in Boudolf et al (2004).…”

“…While homologs of ATM and its sister kinase ATR (ATM‐ and Rad3‐related) predominantly involved in replication stress response by sensing single‐stranded DNA have also been identified in Arabidopsis (Garcia et al , 2000; Culligan et al , 2004; Culligan & Britt, 2008), no homologs of Chk2 or its sister kinase Chk1 could be found in plants to date. Furthermore, even though a homolog of the yeast Wee1 kinase exists in Arabidopsis and other plants, its function appears to be different as Arabidopsis WEE1 was found to act during S phase after hydroxyurea (HU)‐induced replication stress and not in repressing CDK activity during mitosis or blocking cell division after DSB formation (De Schutter et al , 2007; Cools et al , 2011). Moreover, transgenic plants expressing a mutant version of CDKA;1, the Arabidopsis homolog of mammalian Cdk1 and Cdk2, in which the putative WEE1 target sites were replaced with non‐phosphorylatable amino acids, were not hypersensitive to HU indicated that cell‐cycle arrest after DNA damage is differently regulated in plants (Dissmeyer et al , 2009, 2010).…”

The regulatory network of cell-cycle progression is fundamentally different in plants versus yeast or metazoans

“…3B and D). 27,29 This suggested that perhaps a regulation of CDKA;1 by P-loop phosphorylation is restricted to checkpoint activity in Arabidopsis.…”

“…23 Consistent with this, wee1 mutants in Arabidopsis were not found to display growth defects. 27 However, RNAimediated downregulation of WEE1 expression in tomato resulted in reduced plant growth and a smaller fruit size, 28 possibly indicating that WEE1 controls growth in a species-specific manner.…”