Identification, purification, and cloning of a polypeptide (PRTF/GRM) that binds to mating-specific promoter elements in yeast.
In yeast the alpha-specific regulators, alpha 1 and alpha 2 have been proposed to be DNA-binding proteins, both of which have to interact with an additional factor called PRTF or GRM, respectively, to exert their biological functions. Although the cis-acting sequence requirements for alpha 1 and alpha 2 are different, their target sequences share a common motif. PRTF or GRM is thought to act via this common DNA sequence; therefore, it has been suggested that they represent the same factor. I purified a protein that binds to this common promoter element by DNA affinity chromatography. The purified protein is able to recruit the alpha-specific activator alpha 1 to its binding sites, suggesting that it is indeed PRTF. Further evidence is presented to show that PRTF and GRM are the same protein and that PRTF plays a role in the activation of a-specific genes. Specific antibodies to the purified protein were obtained. They identify the protein as a component of DNA-protein complexes that formed with cell-type-specific promoter sequences. Using these antibodies, the gene encoding the protein was cloned from a yeast lambda gt11 expression library. The DNA sequence established that the gene encoding PRTF/GRM is identical with a previously described gene, FUN80 (essential factor of unknown function) or MCM1 (minichromosome maintenance). Sequence comparison showed further that PRTF/GRM shares similarities with a repressor from yeast, ARGRI, and the mammalian transcription factor SRF.
Ammerer
G
G
MRC Laboratory of Molecular Biology, Cambridge, UK.
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000302:isolation & purification*; D005656:Fungal Proteins / Q000235:genetics / Q000302:isolation & purification*; D005800:Genes, Fungal; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011956:Receptors, Cell Surface / Q000235:genetics / Q000302:isolation & purification*; D051964:Receptors, Mating Factor; D018000:Receptors, Peptide; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors; D014158:Transcription, Genetic
D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D011956:Receptors, Cell Surface; D051964:Receptors, Mating Factor; D018000:Receptors, Peptide; D014157:Transcription Factors