Journal of Bacteriology, July 2001, p. 4190-4201, Vol. 183, No. 14
0021-9193/01/$04.00+0 DOI: 10.1128/JB.183.14.4190-4201.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
andInstituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, 2781-901 Oeiras,1 and Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, Quinta da Torre, 2825 Monte de Caparica,2 Portugal
Received 26 February 2001/Accepted 27 April 2001
The proteins involved in the utilization of L-arabinose by Bacillus subtilis are encoded by the araABDLMNPQ-abfA metabolic operon and by the araE/araR divergent unit. Transcription from the ara operon, araE transport gene, and araR regulatory gene is induced by L-arabinose and negatively controlled by AraR. The purified AraR protein binds cooperatively to two in-phase operators within the araABDLMNPQ-abfA (ORA1 and ORA2) and araE (ORE1 and ORE2) promoters and noncooperatively to a single operator in the araR (ORR3) promoter region. Here, we have investigated how AraR controls transcription from the ara regulon in vivo. A deletion analysis of the ara promoters region showed that the five AraR binding sites are the key cis-acting regulatory elements of their corresponding genes. Furthermore, ORE1-ORE2 and ORR3 are auxiliary operators for the autoregulation of araR and the repression of araE, respectively. Analysis of mutations designed to prevent cooperative binding of AraR showed that in vivo repression of the ara operon requires communication between repressor molecules bound to two properly spaced operators. This communication implicates the formation of a small loop by the intervening DNA. In an in vitro transcription system, AraR alone sufficed to abolish transcription from the araABDLMNPQ-abfA operon and araE promoters, strongly suggesting that it is the major protein involved in the repression mechanism of L-arabinose-inducible expression in vivo. The ara regulon is an example of how the architecture of the promoters is adapted to respond to the particular characteristics of the system, resulting in a tight and flexible control.
Present address: Instituto de Histologia e Embriologia, Faculdade
de Medicina de Lisboa, 1649-028, Lisbon, Portugal.
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