DNA Integrity Checkpoint Control

DNA integrity project imageGenome integrity of cells is threatened by DNA damage that is consequence of environmental or endogenous origins. To cope with these stress conditions, cells have developed a set of surveillance mechanisms that monitor the status and structure of DNA during cell cycle progression.

Genome integrity of cells is threatened by DNA damage that is consequence of environmental or endogenous origins. To cope with these stress conditions, cells have developed a set of surveillance mechanisms that monitor the status and structure of DNA during cell cycle progression. In fission yeast and mammals, DNA damage activates the signaling kinases ataxia telangiectasia mutated (ATM) and Rad3-related (ATR) that simultaneously turn on DNA repair complexes and arrest cell division, allowing cells to repair damaged DNA before proceeding into mitosis.

In plants, the role of ATM/ATR-dependent signaling in expression of several DNA repair genes, such as RAD51 and PARP-1 has been demonstrated as well. However, little is known on the molecular players in that modulate plant cell cycle progression in response to DNA stress. We have found that the Arabidopsis WEE1 and SMR genes, encoding negative regulator of the cell cycle, are key plant DNA stress checkpoint regulators that respond to replication defects and DNA breaks, respectively. Currently, we aim to map the consequence of impaired checkpoint control on genome integrity, both in Arabidopsis and crop plants. Additionally, we aim to map the complete pathways that control WEE1 and SMR activation in response to DNA stress, both at the transcriptional and post-transcriptional level.

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DNA damage induced marker gene expression

A particular interest goes to control of the DNA damage response of the stem cells. In contrast to mammals, many plant species live up to several hundreds or even thousands of years. Plants achieve this long lifespan because of their ability to constantly generate new cells at localized regions called meristems, which hold the stem cell niches (SCN). Remarkably, at the centre of the root SCN, a group of mitotically inactive cells are found called the Quiescent Centre (QC). Contrary to their surrounding stem cells, QC cells divide infrequently. Because QC cells appear to be more DNA stress tolerant compared to their surrounding stem cells, it has been speculated that QC cell division is used to repopulate damaged stem cell niches, in such way contributing to the longevity of the stem cell niche. Based on the recent identification of a rate-limiting factor for QC cell division, we apply single cell imaging, profiling, and sequencing techniques to analyse the DNA damage response in stem cells. In particular our objectives are to establish a full understanding on why QC cells are more DNA stress resistant compared to non-QC stem, to identify the molecular components that drive QC cell division, and to unravel the signalling components that activate QC cell division in response to stress. Combined, these results may form the basis for the development of novel innovative strategies to obtain stress resistant crops, thus offering new perspectives towards plant breeding for growth improvement.

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Wild type (left) versus checkpoint mutant (right) plants

Publications:

Hu Z, Cools T, Kalhorzadeh P, Heyman J, De Veylder L. (2015) Deficiency of the Arabidopsis helicase RTEL1 triggers a SOG1-dependent replication checkpoint in response to DNA cross-links. Plant Cell 27, 149-161.

Kalhorzadeh P., Hu Z., Cools T., Amiard S., Willing E.-M., De Winne N., Gevaert K., De Jaeger G., Schneeberger K., White C.I. and De Veylder L. (2014). Arabidopsis thaliana RNase H2 deficiency counteracts the needs for the WEE1 checkpoint kinase but triggers genome instability. Plant Cell 26, 3680-3692.

Yi D., Lessa Alvim Kamei C., Cools T., Vanderauwera S., Takahashi N., Okushima Y., Eekhout T., Yoshiyama K.O., Larkin J., Van den Daele H., Conklin P., Britt A., Umeda M. and De Veylder L. (2014). The Arabidopsis thaliana SIAMESE-RELATED cyclin-dependent kinase inhibitors SMR5 and SMR7 control the DNA damage checkpoint in response to reactive oxygen species. Plant Cell 26, 396-309.

Heyman J., Cools T., Vandenbussche F., Heyndrickx K.S., Van Leene J., Vercauteren I., Vanderauwera S., Vandepoele K., De Jaeger G., Van Der Straeten D. and De Veylder L. (2013). ERF115 controls root quiescent center cell division and stem cell replenishment. Science 342, 860-863.

Cools ,T., Iantcheva A., Weimer A.K., Boens S. Takahashi N., Maes S., Van den Daele H., Van Isterdael G., Schnittger A., De Veylder L. (2011). The Arabidopsis thaliana checkpoint kinase WEE1 protects against premature vascular differentiation during replication stress. Plant Cell 23, 1435-1448.

Cools T., Iantcheva A., Weimer A.K., Boens S. Takahashi N., Maes S., Van den Daele H., Van Isterdael G., Schnittger A. and De Veylder L. (2011). The Arabidopsis thaliana checkpoint kinase WEE1 protects against premature vascular differentiation during replication stress. Plant Cell 23, 1435-1448.

Takahashi N., Quimbaya M., Schubert V., Lammens T., Vandepoele K., Schubert I., Matsui M., Inzé D., Berx G. and De Veylder L. (2010). The MCM-binding protein ETG1 aids sister chromatid cohesion required for postreplicative homologous recombination repair. PLoS Genet. 10, e1000817.

Cools T., Iantcheva A., Maes S., Van den Daele H. and De Veylder L. (2010). A replication stress-induced synchronization method for Arabidopsis thaliana root meristems. Plant J. 64, 705-714.

Cools T. and De Veylder L. (2009). DNA stress checkpoint control and plant development. Curr. Opin. Plant Biol. 12, 23-28.

Takahashi N., Lammens T., Boudolf V., Maes S., Yoshizumi T., De Jaeger G., Witters E., Inzé D. and De Veylder L. (2008). The DNA replication checkpoint aids survival of plants deficient in the novel replisome factor ETG1. EMBO J. 27, 1840-1851.

De Schutter K., Joubès J., Cools T., Verkest A., Corellou F., Babiychuk E., Van Der Schueren E., Beeckman T., Kushnir S., Inzé D. and De Veylder L. (2007). Arabidopsis WEE1 kinase controls cell cycle arrest in response to activation of the DNA integrity checkpoint. Plant Cell 19, 211-225.