A NEW REGULATORY ELEMENT MODULATES HOMOSERINE LACTONE-MEDIATED AUTOINDUCTION OF TI PLASMID CONJUGAL TRANSFER

Conjugal transfer of the Agrobacterium tumefaciens nopaline-type Ti pl asmid pTiC58 is induced by agrocinopines A and B, opines secreted by c rown gall tumors induced by the bacterium. This regulation functions t hrough the transcriptional repressor, AccR. However, actual transcript ion of the tra genes is regulated by autoinduction through the activat or TraR and the substituted homoserine lactone second messenger. Agrob acterium autoinducer (AAI). We have identified a new regulatory elemen t that modulates the response of TraR to AAI. The gene, called traM, s uppresses TraR-AAI activation of transcription of tra genes carried on recombinant clones. The suppression could be relieved by increasing t he expression of TraR but not by increasing AAI levels. traM is locate d between traR and traAF on pTiC58 and is transcribed in the clockwise direction. The 306-bp gene encodes an 11.2-kDa protein showing no sig nificant relatedness to other proteins in the databases. Mutations in traM in pTiC58 conferred a transfer-constitutive phenotype, and strain s harboring the Ti plasmid produced easily detectable amounts of AAI, These same mutations engineered into the transfer-constitutive Ti plas mid pTiC58 Delta nccR conferred a hyperconjugal phenotype and very hig h levels of AAI production. Expression of traM required TraR, indicati ng that transcription of the gene is regulated by the autoinduction sy stem. TraM had no effect on the expression of traR, demonstrating that the suppressive effect is not due to repression of the gene encoding the activator. These results suggest that TraM is not a direct transcr iptional regulator. Since the suppressive effect is demonstrable only when traM is overexpressed with respect to traR, we suggest that TraM functions to sequester TraR from the very small amounts of AAI produce d under conditions when the agrocinopines are not present.