Distinct transcriptional responses of RNA polymerases I, II and III to aptamers that bind TBP.
The TATA-binding protein (TBP) is a general factor that is involved in transcription by all three types of nuclear RNA polymerase. To delineate the roles played by the DNA-binding surface of TBP in these transcription reactions, we used a set of RNA aptamers directed against TBP and examined their ability to perturb transcription in vitro by the different RNA polymerases. Distinct responses to the TBP aptamers were observed for transcription by different types of polymerase at either the initiation, reinitiation or both stages of the transcription cycle. We further probed the TBP interactions in the TFIIIB*DNA complex to elucidate the mechanism for the different sensitivity of Pol III dependent transcription before and after preinitiation complex (PIC) formation. Lastly, the aptamers were employed to measure the time required for Pol III PIC formation in vitro. This approach can be generalized to define the involvement of a particular region on the surface of a protein at particular stages in a biological process.
Fan
Xiaochun
X
Department of Molecular Biology and Genetics, Cornell University Ithaca, NY 14853, USA.
Shi
Hua
H
Lis
John T
JT
D045304:Cytochromes c / Q000235:genetics; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D007700:Kinetics; D009843:Oligoribonucleotides / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D012318:RNA Polymerase I / Q000378:metabolism; D012319:RNA Polymerase II / Q000378:metabolism; D012320:RNA Polymerase III / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics; D035181:TATA-Box Binding Protein / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology*; D014158:Transcription, Genetic
C014059:CYC1 protein, S cerevisiae; D009843:Oligoribonucleotides; D029701:Saccharomyces cerevisiae Proteins; D035181:TATA-Box Binding Protein; D045304:Cytochromes c; D012319:RNA Polymerase II; D012321:DNA-Directed RNA Polymerases; D012318:RNA Polymerase I; D012320:RNA Polymerase III