CHARACTERIZATION OF TWO NEW GENES THAT REGULATE CONJUGATIVE PLASMID TRANSFER ON RHIZOBIA

CASTAGNO, L.; LUCHETTI, A.; NILSSON, J; PISTORIO, M.; TORRES TEJERIZO, G. A.

Congreso; Reunión conjunta SAIB-SAMIGE 2020; 2020

Rhizobia are gram-negative bacteria that are able to establish a symbiotic interaction with leguminous plants. Due to their nitrogen fixing capacity, the study of these microorganisms has acquired great relevance for the agriculture. Rhizobia usually harbor many plasmids in their genome. Frequently, some of these elements can be transferred to other strains by conjugation. Two main mechanisms of regulation of rhizobial plasmid transfer have been described: Quorum sensing (QS) and rctA/rctB system. The QS regulation mechanism is usually based on the production of a signal molecule (in most of the cases, an acyl homoserin lactone, AHL) by the traI gene. This molecule accumulates in the environment, and when it reaches a certain concentration, the molecule enters the cell and binds to the TraR regulator, which will allow expression of conjugative genes in response to population density. In plasmids regulated by the rctA/rctB system, the product of rctA inhibits conjugative genes expression. To allow conjugation, the inhibitory function of RctA must be reduced by the action of RctB, but conditions for RctB expression are not known yet. In the last years, several molecules and new genes that modulate conjugative transfer have been described, even in those systems regulated by QS or rctA/rctB, demonstrating that new actors can tightly regulate the process. pLPU83a is a plasmid from Rhizobium favelukesii LPU83, an acid tolerant rhizobia able to nodulate different species of leguminous plants. This plasmid harbors a traR gene in the conjugation locus, so it seems to be regulated by a QS system. Nevertheless, there is no traI within the conjugative region. In this work, we demonstrate that conjugative transfer of this plasmid does not respond to population density in the same way that other QS regulated plasmids, and that production of different AHLs does not affect the conjugative process. Furthermore, we use bioinformatics and molecular biology tools to describe two hypothetical genes located between conjugative genes, which are involved in the regulation of the process. One of the mentioned genes is essential for conjugation, and the other one is involved in transcriptional regulation of conjugative genes. Previously, we showed that pLPU83a is able to switch between different transfer machineries depending on its genomic background. The hypothetical genes are also involved in the conjugative transfer from different genomic backgrounds. Thus, we introduce new elements of a different mechanism of conjugative transfer regulation, which affect plasmid transfer behavior