Open AccessArchaeal Intrinsic Transcription Termination In Vivo
Author(s) -
Thomas J. Santangelo,
Ľubomíra Čuboňová,
Katherine M. Skinner,
John N. Reeve
Publication year - 2009
Publication title -
journal of bacteriology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.652
H-Index - 246
eISSN - 1067-8832
pISSN - 0021-9193
DOI - 10.1128/jb.00982-09
Subject(s) - biology , thermococcus , transcription (linguistics) , rna polymerase , terminator (solar) , microbiology and biotechnology , gene , dna , palindromic sequence , reporter gene , rna , genetics , gene expression , palindrome , archaea , genome , ionosphere , linguistics , philosophy , physics , astronomy
Thermococcus kodakarensis (formerlyThermococcus kodakaraensis ) strains have been constructed with synthetic and natural DNA sequences, predicted to function as archaeal transcription terminators, identically positioned between a constitutive promoter and a β-glycosidase-encoding reporter gene (TK1761). Expression of the reporter gene was almost fully inhibited by the upstream presence of 5′- (T8 ) and was reduced >70% by archaeal intergenic sequences that contained oligo(T) sequences. An archaeal intergenic sequence (tmcrA ) that conforms to the bacterial intrinsic terminator motif reduced TK1761 expression ∼90%, but this required only the oligo(T) trail sequence and not the inverted-repeat and loop region. Template DNAs were amplified from eachT. kodakarensis strain, and transcription in vitro byT. kodakarensis RNA polymerase was terminated by sequences that reduced TK1761 expression in vivo. Termination occurred at additional sites on these linear templates, including at a 5′- (A8 ) sequence that did not reduce TK1761 expression in vivo. When these sequences were transcribed on supercoiled plasmid templates, termination occurred almost exclusively at oligo(T) sequences. The results provide the first in vivo experimental evidence for intrinsic termination of archaeal transcription and confirm that archaeal transcription termination is stimulated by oligo(T) sequences and is different from the RNA hairpin-dependent mechanism established for intrinsic bacterial termination.