Functional characterization of a maize purine transporter by expression inAspergillus nidulans

We have characterized the function of Leaf Permease1 (LPE1), a protein that is necessary for proper chloroplast development in maize, by functional ex pression in the filamentous fungus Aspergillus nidulans. The choice of this ascomycete was dictated by the similarity of its endogenous purine transpo rters to LPE1 and by particular genetic and physiological features of purin e transport and metabolism in A. nidulans. When Lpe1 was expressed in a pur ine transport-deficient A. nidulans strain, the capacity for uric acid and xanthine transport was acquired. This capacity was directly dependent on Lp e1 copy number and expression level. Interestingly, overexpression of LPE1 from >10 gene copies resulted in transformants with pleiotropically reduced growth rates on various nitrogen sources and the absolute inability to tra nsport purines. Kinetic analysis established that LPE1 is a high-affinity ( K-m = 30 +/- 2.5 muM), high-capacity transporter specific for the oxidized purines xanthine and uric acid. Competition studies showed that high concen trations of ascorbic acid (>30 mM) competitively inhibit LPE1-mediated puri ne transport. This work defines the biochemical function of LPE1, a plant r epresentative of a large and ubiquitous transporter family. In addition, A. nidulans is introduced as a novel model system for the cloning and/or func tional characterization of transporter genes.