Two polypeptide chains in yeast transcription factor tau interact with DNA.
Yeast transcription factor tau interacts with the A and B blocks of the intragenic promoter of tRNA genes. The structure of tau was investigated by identifying the polypeptide chains specifically complexed to the tRNA3Glu gene. Highly purified factor, obtained by an improved purification procedure, contained several polypeptide chains, four of which (Mr = 145,000, 135,000, 100,000 and 65,000) comigrated with tau-DNA complex by polyacrylamide gel electrophoresis. Antibodies raised against the 145- and 100-kDa components altered the migration of tau-DNA complexes in band shift assays and inhibited tRNA synthesis in a reconstituted transcription system. These components are immunologically unrelated proteins. By UV cross-linking to 32P-body-labeled tDNA followed by extensive DNase treatment, two polypeptides of the same size (145 and 100 kDa) were found to be radioactively labeled. Factor tau, therefore, appears to be a multisubunit DNA-binding protein with two distinct polypeptides contributing to DNA recognition. Limited proteolysis of tau generated a protease-resistant tau B (tau B) domain that binds solely to the B block. tau B-tDNA complexes were recognized by anti-145 IgG and contained a 120-kDa polypeptide that could originate from the 145-kDa component by proteolysis. These results strongly suggest that the 145-kDa polypeptide belongs to tau B and is responsible for B block binding.
Gabrielsen
O S
OS
Département de Biologie, Centre d'Etudes Nucléaires de Saclay, Gif-sur-Yvette, France.
Marzouki
N
N
Ruet
A
A
Sentenac
A
A
Fromageot
P
P
D000937:Antigen-Antibody Reactions; D015153:Blotting, Western; D002846:Chromatography, Affinity; D003432:Cross-Linking Reagents; D004258:DNA Polymerase III; D004271:DNA, Fungal / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D005800:Genes, Fungal; D066298:In Vitro Techniques; D046911:Macromolecular Substances; D008970:Molecular Weight; D012352:RNA, Transfer, Glu / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000502:physiology*; D014157:Transcription Factors / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D014158:Transcription, Genetic; D014466:Ultraviolet Rays
D003432:Cross-Linking Reagents; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D012352:RNA, Transfer, Glu; D014157:Transcription Factors; C041273:transcription factor tau; D004258:DNA Polymerase III