Mcm1 promotes replication initiation by binding specific elements at replication origins.
Minichromosome maintenance protein 1 (Mcm1) is required for efficient replication of autonomously replicating sequence (ARS)-containing plasmids in yeast cells. Reduced DNA binding activity in the Mcm1-1 mutant protein (P97L) results in selective initiation of a subset of replication origins and causes instability of ARS-containing plasmids. This plasmid instability in the mcm1-1 mutant can be overcome for a subset of ARSs by the inclusion of flanking sequences. Previous work showed that Mcm1 binds sequences flanking the minimal functional domains of ARSs. Here, we dissected two conserved telomeric X ARSs, ARS120 (XARS6L) and ARS131a (XARS7R), that replicate with different efficiencies in the mcm1-1 mutant. We found that additional Mcm1 binding sites in the C domain of ARS120 that are missing in ARS131a are responsible for efficient replication of ARS120 in the mcm1-1 mutant. Mutating a conserved Mcm1 binding site in the C domain diminished replication efficiency in ARS120 in wild-type cells, and increasing the number of Mcm1 binding sites stimulated replication efficiency. Our results suggest that threshold occupancy of Mcm1 in the C domain of telomeric ARSs is required for efficient initiation. We propose that origin usage in Saccharomyces cerevisiae may be regulated by the occupancy of Mcm1 at replication origins.
Chang
Victoria K
VK
Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853, USA.
Donato
Justin J
JJ
Chan
Clarence S
CS
Tye
Bik K
BK
D001483:Base Sequence; D001665:Binding Sites; D015825:Chromosomes, Fungal; D004247:DNA / Q000378:metabolism*; D018983:DNA Footprinting; D004261:DNA Replication; D026341:Minichromosome Maintenance 1 Protein / Q000378:metabolism*; D008957:Models, Genetic; D008969:Molecular Sequence Data; D010957:Plasmids; D011485:Protein Binding; D012045:Regulatory Sequences, Nucleic Acid; D018741:Replication Origin; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism
D026341:Minichromosome Maintenance 1 Protein; D029701:Saccharomyces cerevisiae Proteins; D004247:DNA