Functional characterization of the two nucleobase cation symporters of Physcomitrella patens reveals their solute specificity

The two genes PpNCS1A and PpNCS1B of the moss Physcomitrella patens encode for nucleobase transporters and putative members of the purine-related transporter (PRT) or nucleobase:cation symporter 1 (NCS1) family. Previously characterized members of this family include the uracil transporter of Saccharomyces cerevisiae (FUR4), the adenine-guanine-hypoxanthine-cytosine transporter of S. cerevisiae (FCY2) and the recently characterized adenine-guanine-uracil transporters of Arabidopsis thaliana (AtNCS1) and Chlamydomonas reinhardtii (CrNCS1). The coding regions of two NCS1 genes of P. patens were cloned into yeast expression vectors and expressed in yeast strains lacking their own NCS1 genes. Transport profiles of the two P. patens NCS1 genes were characterized by radiolabeled nucleobase uptake and competition assays, and toxic analog growth studies. Kinetic parameters were determined by radionucleobase homologous competition assays. Interestingly, the two genes exhibit identical transport profiles, but in many cases transport their respective substrates with drastically different affinities. Also of note is the relatively broad substrate specificity exhibited by the two transporter proteins as compared to other members of the NCS1 family. PpNCS1A and PpNCS1B move adenine, cytosine, guanine, uracil, hypoxanthine and xanthine. The phylogenetic position of P. patens raises the possibility of making inferences about the ancestral state and evolution of nucleobase transporters following divergence of land plants