Strand-specific modulation of UV photoproducts in 5S rDNA by TFIIIA binding and their effect on TFIIIA complex formation.
The relationship between UV-induced photoproduct formation and transcription factor binding was studied in a 214 bp fragment containing the entire Xenopus borealis 5S rRNA gene. DNA mobility shift and DNase I footprinting show a strong inhibition of TFIIIA binding to UV-damaged 5S rDNA. An average of approximately 2 cyclobutane pyrimidine dimers (CPDs) per 214 bp fragment, and a lesser amount of pyrimidine-pyrimidone (6-4) dimers, reduced the fraction of TFIIIA bound by approximately 70%. Furthermore, irradiation of the TFIIIA/5S rDNA complex displaces TFIIIA at doses of 0.8-2 CPDs/fragment, indicating the complex is unable to accommodate UV photoproducts. UV photofootprinting of the 50 bp TFIIIA binding region of 5S rDNA (or ICR) shows that TFIIIA binding modulates photoproduct formation primarily in the template strand. Formation of CPDs at six different sites is strongly inhibited, while another CPD site is strongly enhanced, by TFIIIA binding. Most of these sites are located in one of three boxes (A, IE, or C) designated as TFIIIA contact sites in the ICR, while one site is between these boxes. Formation of (6-4) dimers is also inhibited at several sites in the template strand by TFIIIA binding. However, formation of photoproducts in the nontemplate strand is much less affected by TFIIIA binding, where only one CPD site is inhibited in the complex. These data indicate that formation of UV photoproducts in 5S rDNA can be markedly affected by TFIIIA binding, and complex formation is inhibited by UV photoproducts.
Liu
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Department of Biochemistry and Biophysics, Washington State University, Pullman 99164-4660, USA.
Conconi
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A
Smerdon
M J
MJ
D001665:Binding Sites / Q000528:radiation effects; D004249:DNA Damage; D018983:DNA Footprinting; D004275:DNA, Ribosomal / Q000378:metabolism / Q000528:radiation effects*; D004268:DNA-Binding Proteins / Q000378:metabolism* / Q000528:radiation effects*; D011485:Protein Binding / Q000528:radiation effects; D011740:Pyrimidine Dimers / Q000378:metabolism; D012341:RNA, Ribosomal, 5S / Q000378:metabolism / Q000528:radiation effects*; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors / Q000378:metabolism* / Q000528:radiation effects*; D014466:Ultraviolet Rays
D004275:DNA, Ribosomal; D004268:DNA-Binding Proteins; D011740:Pyrimidine Dimers; D012341:RNA, Ribosomal, 5S; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors