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 (> or =850 kDa) complex, found in nondamaged cells, contains hypophosphorylated Rad9, whereas the smaller (560 kDa) complex, which forms after DNA damage, contains hyperphosphorylated Rad9 and Rad53. This smaller Rad9 complex is capable of catalyzing phosphorylation and release of active Rad53 kinase, a process requiring the kinase 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-dependent hyperphosphorylation of Rad9 results in formation of the smaller Rad9 complex and recruitment of Rad53. This complex then catalyzes activation of Rad53 by acting as a scaffold that brings Rad53 molecules into close proximity, facilitating Rad53 in trans autophosphorylation and subsequent release of activated Rad53.
Gilbert
C S
CS
Imperial Cancer Research Fund, Clare Hall Laboratories, CDC Laboratory, South Mimms, EN6 3LD, Hertfordshire, United Kingdom.
Green
C M
CM
Lowndes
N F
NF
D000255:Adenosine Triphosphate / Q000378:metabolism*; D002453:Cell Cycle / Q000502:physiology; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D064447:Checkpoint Kinase 2; D004260:DNA Repair; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015151:Immunoblotting; D047908:Intracellular Signaling Peptides and Proteins; D046911:Macromolecular Substances; D008954:Models, Biological; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D004718:Saccharomycetales / Q000502:physiology*
D018797:Cell Cycle Proteins; D005656:Fungal Proteins; D047908:Intracellular Signaling Peptides and Proteins; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C069930:rad9 protein; D000255:Adenosine Triphosphate; D064447:Checkpoint Kinase 2; C088937:MEC1 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; C067022:RAD53 protein, S cerevisiae