Published online 4 June 2004
Nucleic Acids Research, 2004, Vol. 32, No. 10 3093-3100
© 2004 Oxford University Press
Sequence-specific Rho–RNA interactions in transcription termination
Department of Biology and Department of Chemistry, Indiana University, Bloomington, IN 47405, USA and Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, KY 40202, USA
*Tel: +1 502 852 5351; Fax: +1 502 852 5350; Email: j.graham{at}louisville.edu
Received as resubmission May 10, 2004; Accepted May 11, 2004
The bacteriophage 
tR1 terminator encodes a region of the nascent cro transcript containing RNA residues recognized by termination factor Rho. To identify ribonucleotide–protein interactions contributing to termination, a library of reporter gene plasmids was constructed containing predominantly single-nucleotide substitutions in a 24 nt region previously shown to be critical for efficient termination. Screening 16 822 bacterial transformants identified 110 terminator mutants, most of which contained two or more nucleotide substitutions. Although the vast majority of single base changes did not reduce tR1 function, 11 specific single-nucleotide substitutions at eight positions interspersed in the upstream part of the target region (5'-ATAACCCCGCTCTT ACACATTCCA-3') did reduce termination. About half of these substitutions also reduced Rho-dependent termination on cro gene templates transcribed by purified RNA polymerase, indicating specific residues critical for optimal terminator function. Other termination defects were not reproduced in these in vitro assays, and likely resulted from indirect effects of altering interactions between tR1 and additional cellular factors capable of attenuating Rho function. Our results indicate that while Rho is able to recognize a wide variety of similar rut site sequences by interacting with alternate nucleotides at critical positions, interactions with specific individual ribonucleotides of the tR1 transcript provide highly efficient Rho-dependent termination.
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  M. Kaczanowska and M. Ryden-Aulin Ribosome Biogenesis and the Translation Process in Escherichia coli Microbiol. Mol. Biol. Rev., September 1, 2007; 71(3): 477 - 494. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. S. Ciampi Rho-dependent terminators and transcription termination. Microbiology, September 1, 2006; 152(Pt 9): 2515 - 2528. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. Hinde, P. Deighan, and C. J. Dorman Characterization of the Detachable Rho-Dependent Transcription Terminator of the fimE Gene in Escherichia coli K-12 J. Bacteriol., December 15, 2005; 187(24): 8256 - 8266. [Abstract] [Full Text] [PDF]
|
|