DNA-protein interactions at the S.cerevisiae alpha 2 operator in vivo.
Two homodimeric proteins, alpha 2 and MCM1, are required to repress transcription of a-cell type specific genes in haploid yeast alpha-cells. In vitro studies by others of the interactions of these proteins with operator DNA have suggested that MCM1 binds to the middle and alpha 2 to the ends of the 31 bp operator. We have previously shown that alpha 2 organizes chromatin structure adjacent to the operator; in the presence of alpha 2 repressor, a precisely positioned nucleosome abuts the operator in both minichromosomes and the genome. We present in vivo footprinting evidence consistent with occupancy of the operator by MCM1 in both a- and alpha-cells and by alpha 2 repressor in alpha-cells. Interestingly, our in vivo results differ from previous in vitro work in detail. In contrast to the broad block of reagent accessibility to DNA by the factors seen in vitro, we find a pattern of strand-specific protection or augmented reactivity in vivo. The in vivo results are consistent with genetic data concerning transcriptional regulation of a-cell specific genes and corroborate the crystallographic data of others.
Murphy
M R
MR
Laboratory of Cellular and Developmental Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892.
Shimizu
M
M
Roth
S Y
SY
Dranginis
A M
AM
Simpson
R T
RT
D001483:Base Sequence; D004271:DNA, Fungal / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003850:Deoxyribonuclease I; D005656:Fungal Proteins / Q000378:metabolism*; D018398:Homeodomain Proteins; D008745:Methylation; D026341:Minichromosome Maintenance 1 Protein; D008969:Molecular Sequence Data; D009875:Operator Regions, Genetic; D011485:Protein Binding; D012097:Repressor Proteins; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors / Q000378:metabolism*; D014466:Ultraviolet Rays
D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D018398:Homeodomain Proteins; C062928:MATA2 protein, S cerevisiae; D026341:Minichromosome Maintenance 1 Protein; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D003850:Deoxyribonuclease I