Rhizobium leguminosarum bv. trifolii rosR gene encodes a transcriptional regulator involved in the positive regulation of exopolysaccharide synthesis. Transcription of rosR is directed by two promoters, distal P1 and proximal P2, of different strengths. We demonstrated that rosR P1 functions as the main promoter and, besides the -35 and -10 sequences, it contains two other important regulatory elements, an extended -10 motif and an upstream promoter element, that play a significant role in the initiation of transcription. Two cAMP-CRP binding sites (I and II) have been identified upstream of P1, both necessary for optimal rosR expression. cAMP-CRP binding site III is located within the P2 promoter and also influences rosR transcription. rosR transcription levels are dependent both on the presence of the cAMP-CRP complex and on the carbon source, indicating regulation of transcription and exopolysaccharide production by catabolite repression.