Regulation of chk1

Abstract

Chk1 is a serine/threonine protein kinase that is the effector of the G2 DNA damage checkpoint. Chk1 homologs have a highly conserved N-terminal kinase domain, and a less conserved C-terminal regulatory domain of ~200 residues. In response to a variety of genomic lesions, a number of proteins collaborate to activate Chk1, which in turn ensures that the mitotic cyclin-dependent kinase Cdc2 remains in an inactive state until DNA repair is completed. Chk1 activation requires the phosphorylation of residues in the C-terminal domain, and this is catalyzed by the ATR protein kinase. How phosphorylation of the C-terminal regulatory domain activates the N-terminal kinase domain has not been elucidated, though some studies have suggested that this phosphorylation relieves an inhibitory intramolecular interaction between the N- and C-termini. However, recent studies in the fission yeast Schizosaccharomyces pombe have revealed that there is more to Chk1 regulation than this auto-inhibition model, and we review these findings and their implication to the biology of this genome integrity determinant.

Figure 1

S. pombe Chk1 domain structure. Chk1 has an N-terminal kinase domain (blue) and a C-terminal regulatory domain (yellow). Regions required for 14-3-3 interaction and nuclear localization [42] are indicated below the schematic. The position of S345 (orange), the site of activating phosphorylation is shown, as are the two regions of the C-terminal domain conserved across species that have been analyzed by mutagenesis [45]. These are "region 1" (green; residues 394–400, RLTRFYS in S. pombe, RMTRFYS in humans), and "region 2" (red; residues 468–477, GDPLEWRKFF in S. pombe, GDPLEWRKFY in humans).

Figure 2

Auto-inhibition model of Chk1 regulation. A. Using the same color scheme as in Figure 1, in its inactive (unphosphorylated) state, the C-terminal domain physically interacts with the kinase domain, precluding it from phosphorylating substrates. B. This intramolecular interaction is blocked by S345 phosphorylation, opening the molecule to uncover the kinase domain.

Figure 3

Model of regulatory effect of Chk1 mutations. Using the same color scheme as in Figure 1: A. a postulated domain organization is shown. B. In its inactive state, the C-terminal domain interacts with the kinase domain to occlude the active site catalytic cleft. C. Phosphorylation opens the molecule to expose the catalytic cleft. D. Activating mutations such as E472D (red, in region 2) block the intramolecular interactions that occlude the catalytic cleft, activating Chk1's kinase activity. E. The E472D suppressor mutations in the kinase domain (Cyan) restore the intramolecular interaction to again occlude the active site. F. The molecule containing both E472D and a suppressor mutation can still be opened by S345 phosphorylation, restoring normal regulation in response to DNA damage.