Identification and characterization of transcription factor IIIA from Schizosaccharomyces pombe.
Transcription factor IIIA (TFIIIA) is specifically required for transcription of 5S rRNA genes and is the archetypal C2H2 zinc finger protein. All known vertebrate TFIIIAs have a similar organization: nine zinc fingers, followed by a C-terminal domain of unknown structure. The zinc fingers of Saccharomyces cerevisiae TFIIIA are interrupted between fingers eight and nine by an 81-amino acid spacer. Aside from the amino acids required for zinc finger folding, TFIIIAs from different species are remarkably divergent, whereas the natural binding site, the internal control region of the 5S rRNA gene, is well conserved. We now describe the identification and characterization of TFIIIA from Schizosaccharomyces pombe. This protein is organized differently from its known homologs, in that it contains eight closely spaced zinc fingers, a ninth zinc finger missing a C-terminal Zn2+-coordinating histidine, a 53- amino acid spacer, and an unprecedented tenth zinc finger. We have confirmed the identity of this divergent protein as TFIIIA by showing that it binds specifically and with high affinity to the S.pombe 5S rRNA gene. Comparison of DNase I protection patterns produced by TFIIIA from multiple species suggests a novel mode of DNA recognition by the S.pombe protein. Recombinant S.pombe TFIIIA was also shown to support specific transcription of the 5S rRNA gene in vitro.
Schulman
Deborah B
DB
Department of Molecular Biology and Microbiology, School of Medicine, Case Western Reserve University, 2109 Adelbert Road, Cleveland, OH 44106, USA.
Setzer
David R
DR
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism; D004275:DNA, Ribosomal / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics*; D024202:Electrophoretic Mobility Shift Assay; D008969:Molecular Sequence Data; D011485:Protein Binding; D012341:RNA, Ribosomal, 5S / Q000235:genetics; D012568:Schizosaccharomyces / Q000235:genetics*; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors / Q000235:genetics*; D014158:Transcription, Genetic
D018076:DNA, Complementary; D004271:DNA, Fungal; D004275:DNA, Ribosomal; D004268:DNA-Binding Proteins; D012341:RNA, Ribosomal, 5S; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors