Budding yeast Rad9 is an ATP-dependent Rad53 activating machine

We find budding yeast Rad9 in two distinct, large, and soluble complexes in cell extracts. The larger (greater than or equal to 850 kDa) complex, foun d in nondamaged cells, contains hypophosphorylated Rad9, whereas the smalle r (560 kDa) complex, which forms after DNA damage, contains hyperphosphoryl ated Rad9 and Rad53. This smaller Rad9 complex is capable of catalyzing pho sphorylation and release of active Rad53 kinase, a process requiring the ki nase activity of Rad53. However, Mec1 and Tel1 are no longer required once the 560 kDa complex has been formed. We propose a model whereby Mec1/Tel1-d ependent hyperphosphorylation of Rad9 results in formation of the smaller R ad9 complex and recruitment of Rad53. This complex then catalyzes activatio n of Rad53 by acting as a scaffold that brings Rad53 molecules into close p roximity, facilitating Rad53 in trans autophosphorylation and subsequent re lease of activated Rad53.