The Schizosaccharomyces pombe rad3 checkpoint gene.
The rad3 gene of Schizosaccharomyces pombe is required for checkpoint pathways that respond to DNA damage and replication blocks. We report the complete rad3 gene sequence and show that rad3 is the homologue of Saccharomyces cerevisiae ESR1 (MEC1/SAD3) and Drosophila melanogaster mei-41 checkpoint genes. This establishes Rad3/Mec1 as the only conserved protein which is required for all the DNA structure checkpoints in both yeast model systems. Rad3 is an inessential member of the 'lipid kinase' subclass of kinases which includes the ATM protein defective in ataxia telangiectasia patients. Mutational analysis indicates that the kinase domain is required for Rad3 function, and immunoprecipitation of overexpressed Rad3 demonstrates an associated protein kinase activity. The previous observation that rad3 mutations can be rescued by a truncated clone lacking the kinase domain may be due to intragenic complementation. Consistent with this, biochemical data suggest that Rad3 exists in a complex containing multiple copies of Rad3. We have identified a novel human gene (ATR) whose product is closely related to Rad3/Esr1p/Mei-41. ATR can functionally complement esr1-1 radiation sensitivity in S. cerevisiae. Together, the structural conservation and functional complementation suggest strongly that the mechanisms underlying the DNA structure checkpoints are conserved throughout evolution.
Bentley
N J
NJ
MRC Cell Mutation Unit, Sussex University, Falmer, UK.
Holtzman
D A
DA
Flaggs
G
G
Keegan
K S
KS
DeMaggio
A
A
Ford
J C
JC
Hoekstra
M
M
Carr
A M
AM
D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D064007:Ataxia Telangiectasia Mutated Proteins; D001483:Base Sequence; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000378:metabolism; D003001:Cloning, Molecular; D004249:DNA Damage; D004265:DNA Helicases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017931:DNA Primers; D004261:DNA Replication; D004307:Dose-Response Relationship, Radiation; D004331:Drosophila melanogaster / Q000235:genetics; D005800:Genes, Fungal; D005816:Genetic Complementation Test; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009838:Oligodeoxyribonucleotides; D010770:Phosphotransferases / Q000378:metabolism; D010802:Phylogeny; D017346:Protein Serine-Threonine Kinases; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D012568:Schizosaccharomyces / Q000166:cytology* / Q000235:genetics* / Q000528:radiation effects; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid
D018797:Cell Cycle Proteins; D017931:DNA Primers; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D010770:Phosphotransferases; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases; D000251:Adenosine Triphosphatases; C061897:Rad3 protein, S cerevisiae; D004265:DNA Helicases