Phosphate Assimilation in Rhizobium ( Sinorhizobium ) meliloti : Identification of a pit -Like Gene

Abstract

ABSTRACT Rhizobium meliloti mutants defective in the phoCDET -encoded phosphate transport system form root nodules on alfalfa plants that fail to fix nitrogen (Fix − ). We have previously reported that two classes of second-site mutations can suppress the Fix − phenotype of phoCDET mutants to Fix + . Here we show that one of these suppressor loci ( sfx1 ) contains two genes, orfA and pit , which appear to form an operon transcribed in the order orfA-pit . The Pit protein is homologous to various phosphate transporters, and we present evidence that three suppressor mutations arose from a single thymidine deletion in a hepta-thymidine sequence centered 54 nucleotides upstream of the orfA transcription start site. This mutation increased the level of orfA-pit transcription. These data, together with previous biochemical evidence, show that the orfA-pit genes encode a P i transport system that is expressed in wild-type cells grown with excess P i but repressed in cells under conditions of P i limitation. In phoCDET mutant cells, orfA-pit expression is repressed, but this repression is alleviated by the second-site suppressor mutations. Suppression increases orfA-pit expression compensating for the deficiencies in phosphate assimilation and symbiosis of the phoCDET mutants.