Identification of a transcription factor IIIA-interacting protein.
Transcription factor IIIA (TFIIIA) activates 5S ribosomal RNA gene transcription in eukaryotes. The protein from vertebrates has nine contiguous Cys(2)His(2)zinc fingers which function in nucleic acid binding, and a C-terminal region involved in transcription activation. In order to identify protein partners for TFIIIA, yeast two-hybrid screens were performed using the C-terminal region of Xenopus TFIIIA as an attractor and a rat cDNA library as a source of potential partners. A cDNA clone was identified which produced a protein in yeast that interacted with Xenopus TFIIIA but not with yeast TFIIIA. This rat clone was sequenced and the primary structure of the human homolog (termed TFIIIA-intP for TFIIIA-interacting protein) was determined from expressed sequence tags. In vitro interaction of recombinant human TFIIIA-intP with recombinant Xenopus TFIIIA was demonstrated by immuno-precipitation of the complex using anti-TFIIIA-intP antibody. Interaction of rat TFIIIA with rat TFIIIA-intP was indicated by co-chromatography of the two proteins on DEAE-5PW following fractionation of a rat liver extract on cation, anion and gel filtration resins. In a HeLa cell nuclear extract, recombinant TFIIIA-intP was able to stimulate TFIIIA-dependent transcription of the Xenopus 5S ribosomal RNA gene but not TFIIIA-independent transcription of the human adenovirus VA RNA gene.
Moreland
R J
RJ
Department of Biochemistry and Molecular Biology, University of Oklahoma College of Medicine, Oklahoma City, OK 73104, USA.
Dresser
M E
ME
Rodgers
J S
JS
Roe
B A
BA
Conaway
J W
JW
Conaway
R C
RC
Hanas
J S
JS
D000595:Amino Acid Sequence; D000818:Animals; D018797:Cell Cycle Proteins; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D008110:Liver Extracts / Q000737:chemistry / Q000378:metabolism; D026901:Membrane Transport Proteins; D008969:Molecular Sequence Data; D011485:Protein Binding; D012341:RNA, Ribosomal, 5S / Q000235:genetics; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D012441:Saccharomyces cerevisiae / Q000235:genetics; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D014158:Transcription, Genetic; D020798:Two-Hybrid System Techniques; D014981:Xenopus
D018797:Cell Cycle Proteins; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D008110:Liver Extracts; D026901:Membrane Transport Proteins; C409460:OPTN protein, human; D012341:RNA, Ribosomal, 5S; D011993:Recombinant Fusion Proteins; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors