Dynamics of Rad9 chromatin binding and checkpoint function are mediated by its dimerization and are cell cycle-regulated by CDK1 activity.
Saccharomyces cerevisiae Rad9 is required for an effective DNA damage response throughout the cell cycle. Assembly of Rad9 on chromatin after DNA damage is promoted by histone modifications that create docking sites for Rad9 recruitment, allowing checkpoint activation. Rad53 phosphorylation is also dependent upon BRCT-directed Rad9 oligomerization; however, the crosstalk between these molecular determinants and their functional significance are poorly understood. Here we report that, in the G1 and M phases of the cell cycle, both constitutive and DNA damage-dependent Rad9 chromatin association require its BRCT domains. In G1 cells, GST or FKBP dimerization motifs can substitute to the BRCT domains for Rad9 chromatin binding and checkpoint function. Conversely, forced Rad9 dimerization in M phase fails to promote its recruitment onto DNA, although it supports Rad9 checkpoint function. In fact, a parallel pathway, independent on histone modifications and governed by CDK1 activity, allows checkpoint activation in the absence of Rad9 chromatin binding. CDK1-dependent phosphorylation of Rad9 on Ser11 leads to specific interaction with Dpb11, allowing Rad53 activation and bypassing the requirement for the histone branch.
Granata
Magda
M
Dipartimento di Scienze Biomolecolari e Biotecnologie, Università degli Studi di Milano, Milano, Italy.
Lazzaro
Federico
F
Novarina
Daniele
D
Panigada
Davide
D
Puddu
Fabio
F
Abreu
Carla Manuela
CM
Kumar
Ramesh
R
Grenon
Muriel
M
Lowndes
Noel F
NF
Plevani
Paolo
P
Muzi-Falconi
Marco
M
D016203:CDC2 Protein Kinase / Q000235:genetics / Q000378:metabolism*; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002843:Chromatin / Q000235:genetics / Q000378:metabolism*; D004249:DNA Damage; D019281:Dimerization; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000166:cytology* / Q000235:genetics / Q000378:metabolism*
D018797:Cell Cycle Proteins; D002843:Chromatin; C069930:rad9 protein; D016203:CDC2 Protein Kinase