Overexpression of WEE1 kinase promotes resistance to DNA damage in Chlamydomonas reinhardtii

Kateřina Bišová1, Monika Hlavová1, Dáša Umysová1, James G. Umen2, and Vilém Zachleder1

1Laboratory of Cell Cycles of Algae, Institute of Microbiology, ASCR, Třeboň, Czech Republic 2Plant Molecular Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, USA

WEE1 kinase is a major regulator of mitosis in all known eukaryotes. It phosphorylates and inactivates cyclin dependent kinase (CDK), the key activator of cell cycle progression, during S and G2 phase and delays mitosis in response to DNA damage and developmental signals during G2. In mammals and higher plants, WEE1 is a part of DNA damage checkpoint that ensures maintenance of cell viability in cells, which have suffered genomic damage. Chlamydomonas genome encodes a single homolog of WEE1 kinase; its transcription peaks during the S/M phase of the cell cycle. We cloned a GFP tagged CrWEE1 cDNA under Ni inducible Cyc6 promoter. Recovered transformants were screened for a growth phenotype in the presence and absence of NiCl2. Transformants in the presence of NiCl2 grew slower comparing to untreated controls. Detailed analysis showed that the cells started to grow significantly more slowly within six hour after NiCl2 addition; however, they eventually reached the same mother cell size as the control cells. The cells with overexpressed GFP-WEE1 divided later with smaller division number leading to bigger daughter cell size. The cells also showed significant increase in the survival rate in the presence of zeocin as a DNA damaging agent. The data implies that Chlamydomonas WEE1 kinase is part of both cell sizing mechanism and DNA damage checkpoint. The work was supported by the GACR (grant no. 204/06/0102), the GAAS CR (grant no. IAA500200614) and the Institutional Research Concept (no. AV0Z50200510).

e-mail address of presenting author: bisova@alga.cz