A basal promoter element recognized by free RNA polymerase sigma subunit determines promoter recognition by RNA polymerase holoenzyme.
During transcription initiation by bacterial RNA polymerase, the sigma subunit recognizes the -35 and -10 promoter elements; free sigma, however, does not bind DNA. We selected ssDNA aptamers that strongly and specifically bound free sigma(A) from Thermus aquaticus. A consensus sequence, GTA(C/T)AATGGGA, was required for aptamer binding to sigma(A), with the TA(C/T)AAT segment making interactions similar to those made by the -10 promoter element (consensus sequence TATAAT) in the context of RNA polymerase holoenzyme. When in dsDNA form, the aptamers function as strong promoters for the T. aquaticus RNA polymerase sigma(A) holoenzyme. Recognition of the aptamer-based promoters depends on the downstream GGGA motif from the aptamers' common sequence, which is contacted by sigma(A) region 1.2 and directs transcription initiation even in the absence of the -35 promoter element. Thus, recognition of bacterial promoters is controlled by independent interactions of sigma with multiple basal promoter elements.
Feklistov
Andrey
A
Institute of Molecular Genetics, Moscow 123182, Russia.
Barinova
Nataliya
N
Sevostyanova
Anastasiya
A
Heyduk
Ewa
E
Bass
Irina
I
Vvedenskaya
Irina
I
Kuznedelov
Konstantin
K
Merkiene
Egle
E
Stavrovskaya
Elena
E
Klimasauskas
Saulius
S
Nikiforov
Vadim
V
Heyduk
Tomasz
T
Severinov
Konstantin
K
Kulbachinskiy
Andrey
A
D052157:Aptamers, Nucleotide / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D012321:DNA-Directed RNA Polymerases / Q000235:genetics* / Q000378:metabolism; D020035:Holoenzymes / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011487:Protein Conformation; D012808:Sigma Factor / Q000235:genetics* / Q000378:metabolism; D013824:Thermus / Q000201:enzymology
D052157:Aptamers, Nucleotide; D001426:Bacterial Proteins; D020035:Holoenzymes; D012808:Sigma Factor; C063505:RNA polymerase sigma A; D012321:DNA-Directed RNA Polymerases; C455879:Taq sigma A