Characterization of an unusual, sequence-specific termination signal for T7 RNA polymerase.
We have characterized an unusual type of termination signal for T7 RNA polymerase that requires a conserved 7-base pair sequence in the DNA (ATCTGTT in the non-template strand). Each of the nucleotides within this sequence is critical for function, as any substitutions abolish termination. The primary site of termination occurs 7 nucleotides downstream from this sequence but is context-independent (that is, the sequence around the site of termination, and in particular the nucleotide at the site of termination, need not be conserved). Termination requires the presence of the conserved sequence and its complement in duplex DNA and is abolished or diminished if the signal is placed downstream of regions in which the non-template strand is missing or mismatched. Under the latter conditions, much of the RNA product remains associated with the template. The latter results suggest that proper resolution of the transcription bubble at its trailing edge and/or displacement of the RNA product are required for termination at this class of signal.
He
B
B
Department of Microbiology and Immunology, Morse Institute for Molecular Genetics, State University of New York, Health Science Center, Brooklyn, New York 11203-2098, USA.
Kukarin
A
A
Temiakov
D
D
Chin-Bow
S T
ST
Lyakhov
D L
DL
Rong
M
M
Durbin
R K
RK
McAllister
W T
WT
D001483:Base Sequence; D002874:Chromosome Mapping; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D008969:Molecular Sequence Data; D010281:Parathyroid Hormone / Q000235:genetics*; D011498:Protein Precursors / Q000235:genetics*; D006163:Ribonuclease T1 / Q000378:metabolism; D012259:Ribonuclease, Pancreatic / Q000378:metabolism; D017384:Sequence Deletion; D013728:Terminator Regions, Genetic; D014764:Viral Proteins
D010281:Parathyroid Hormone; D011498:Protein Precursors; D014764:Viral Proteins; C019056:preproparathormone; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases; D006163:Ribonuclease T1; D012259:Ribonuclease, Pancreatic