Genomic footprinting of a yeast tRNA gene reveals stable complexes over the 5'-flanking region.
We have shown by genomic footprinting that the 5'-flanking region of the Saccharomyces cerevisiae tRNASUP53 gene is protected from DNase I digestion. The protected region has a 5' boundary at -40 (relative to the transcription initiation site) and extends into the coding region of the gene, with a 3' boundary at approximately +15. Although the DNase I protection over this region was much greater than at the A- and B-box internal promoters, point mutations within the A or B box that reduced transcription in vitro eliminated the upstream DNase I protection. This implies that formation of a stable complex over the 5'-flanking region is dependent on interaction of the gene with transcription factor IIIC but that stability of the complex may not require continued interaction with this factor. The DNase I protection under varied growth conditions further suggested that the upstream complex is composed of two or more components. The region over the transcription initiation site (approximately +15 to -10) was less protected in stationary-phase cultures, whereas the more upstream region (approximately -10 to -40) was protected in both exponential- and stationary-phase cultures.
Huibregtse
J M
JM
Department of Biological Chemistry, University of Michigan, Ann Arbor 48109-0606.
Engelke
D R
DR
D001483:Base Sequence; D004252:DNA Mutational Analysis; D003850:Deoxyribonuclease I; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D009698:Nucleoproteins / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D012331:RNA, Fungal / Q000235:genetics*; D012343:RNA, Transfer / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D014157:Transcription Factors / Q000032:analysis
D009698:Nucleoproteins; D012331:RNA, Fungal; D014157:Transcription Factors; D012343:RNA, Transfer; D003850:Deoxyribonuclease I