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Abstract:
Eukaryotic cells respond to DNA damage by activating
checkpoint signalling pathways. Checkpoint signals are
transduced by a protein kinase cascade that also requires
non-kinase mediator proteins. One such mediator is the
Saccharomyces cerevisiae Dpb11 protein, which binds to
and activates the apical checkpoint kinase, Mec1. Here, we
show that a ternary complex of Dpb11, Mec1 and another
key mediator protein Rad9 is required for efficient Rad9
phosphorylation by Mec1 in vitro, and for checkpoint
activation in vivo. Phosphorylation of Rad9 by cyclindependent
kinase (CDK) on two key residues generates a
binding site for tandem BRCT repeats of Dpb11, and is
thereby required for Rad9 recruitment into the ternary
complex. Checkpoint signalling via Dpb11, therefore, does
not efficiently occur during G1 phase when CDK is inactive.
Thus, Dpb11 coordinates checkpoint signal transduction
both temporally and spatially, ensuring the initiator
kinase is specifically activated in proximity of one of its
critical substrates.
