Two mcm3 mutations affect different steps in the initiation of DNA replication.
Mcm3 is a subunit of the hexameric MCM2-7 complex required for the initiation and elongation of DNA replication in eukaryotes. We have characterized two mutant alleles, mcm3-1 and mcm3-10, in Saccharomyces cerevisiae and showed that they are defective at different steps of the replication initiation process. Mcm3-10 contains a P118L substitution that compromises its interaction with Mcm5 and the recruitment of Mcm3 and Mcm7 to a replication origin. P118 is conserved between Mcm3, Mcm4, Mcm5, and Mcm7. An identical substitution of this conserved residue in Mcm5 (P83L of mcm5-bob1) strengthens the interaction between Mcm3 and Mcm5 and allows cells to enter S phase independent of Cdc7-Dbf4 kinase (Hardy, C. F., Dryga, O., Pahl, P. M. B., and Sclafani, R. A. (1997) Proc. Natl. Acad. Sci. U. S. A. 94, 3151-3155). Mcm3-1 contains a G246E mutation that diminishes the efficiency of replication initiation (Yan, H., Merchant, A. M., and Tye, B. K. (1993) Genes Dev. 7, 2149-2160) but not its interaction with Mcm5 or recruitment of the MCM2-7 complex to replication origin. These observations indicate that Mcm3-10 is defective in a step before, and Mcm3-1 is defective in a step after the recruitment of the MCM2-7 complex to replication origins.
Lei
Ming
M
Department of Microbiology and Molecular Genetics, Medical College of Wisconsin, Milwaukee, WI 53226, USA. mlei@mcw.edu
Cheng
Irene H
IH
Roberts
Louis A
LA
McAlear
Michael A
MA
Tye
Bik K
BK
D000483:Alleles; D000595:Amino Acid Sequence; D018797:Cell Cycle Proteins / Q000502:physiology*; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000502:physiology; D064168:Minichromosome Maintenance Complex Component 7; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000502:physiology; D016196:S Phase; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; C085711:mcm5 protein, S pombe; C491088:MCM7 protein, S cerevisiae; D064168:Minichromosome Maintenance Complex Component 7