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Journal of Bacteriology, February 2000, p. 1062-1073, Vol. 182, No. 4
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

A pAD1-Encoded Small RNA Molecule, mD, Negatively Regulates Enterococcus faecalis Pheromone Response by Enhancing Transcription Termination

Haruyoshi Tomita1,dagger and Don B. Clewell1,2,*

Department of Biologic and Materials Sciences, School of Dentistry,1 and Department of Microbiology/Immunology, School of Medicine,2 The University of Michigan, Ann Arbor, Michigan 48109

Received 27 July 1999/Accepted 24 November 1999

pAD1 is a 60-kb hemolysin-bacteriocin plasmid in Enterococcus faecalis that encodes a conjugative mating response to a peptide sex pheromone, cAD1, secreted by plasmid-free bacteria. The pheromone response is regulated by two proteins: TraE1, which positively regulates all or most conjugative structural genes, and TraA, which negatively regulates traE1. TraA binds to pAD1 DNA at the iad (encoding the inhibitor peptide iAD1) promoter but is released upon binding to imported pheromone. This leads to enhanced transcription through two closely spaced downstream terminators (t1 and t2) into traE1. TraE1 is believed to then upregulate itself from a site located within t2; thus, a small amount of transcription through t1-t2 could lead to overall induction. It is important therefore that the t1-t2 terminators be tightly controlled to keep the response shut down in the absence of pheromone. A small (200-nucleotide) RNA molecule designated mD is encoded just upstream of t1 by a determinant (traD) oriented in the direction opposite to that of transcripts utilizing t1. mD is expressed at high levels in the uninduced state, but it decreases significantly upon induction. Here we present results of genetic studies relating to the activity of t1-t2 and show that mD strongly enhances transcriptional termination at t1. The mD activity is shown to influence transcription well downstream and can affect the determinant for aggregation substance asa1. The phenomenon is specific in that there is no effect of mD on the unrelated pheromone-responding plasmids pPD1 and pCF10.

* Corresponding author. Mailing address: Department of Biologic and Materials Sciences, School of Dentistry, The University of Michigan, Ann Arbor, MI 48109-1078. Phone: (734) 763-0117. Fax: (734) 763-9905. E-mail: dclewell{at}umich.edu.

dagger Present address: Department of Microbiology, Gunma University School of Medicine, Maebashi City, Gunma 371, Japan.

Journal of Bacteriology, February 2000, p. 1062-1073, Vol. 182, No. 4
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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