CRISPR/Cas9 editing of two adenine phosphoribosyl transferase coding genes reveals the functional specialization of adenine salvage proteins in common bean

Adenine metabolism is important for common bean (Phaseolus vulgaris L.) productivity since this legume uses ureides derived from the oxidation of purine nucleotides as its primary nitrogen storage molecules. Purine nucleotides are produced from de novo synthesis or through salvage pathways. Adenine phosphoribosyl transferase (APRT) is the enzyme dedicated to adenine nucleobase salvage for nucleotide synthesis, but it can also convert active cytokinin bases into their inactive nucleotide forms. In common bean, APRT is encoded by four genes. Gene expression analysis, biochemical properties, and subcellular location indicated functional differences among the common bean APRT isoforms. CRISPR/Cas9 targeted down-regulation of two of the four PvAPRTs followed by metabolomic and physiological analyses of targeted hairy roots revealed that, although the two proteins have redundant functions, PvAPRT1 mostly participated in the salvage of adenine, whereas PvAPRT5 was the predominant form in the regulation of cytokinin homeostasis and stress responses with a high impact in root and nodule growth.