Genetic and Molecular Characterization of a Gene Encoding a Wide Specificity Purine Permease of Aspergillus nidulans Reveals a Novel Family of Transporters Conserved in Prokaryotes and Eukaryotes

doi:10.1074/jbc.270.15.8610

Genetic and Molecular Characterization of a Gene Encoding a Wide Specificity Purine Permease of Aspergillus nidulans Reveals a Novel Family of Transporters Conserved in Prokaryotes and Eukaryotes (*)

From the (1) Institut de Génétique et Microbiologie, Unité Associé au Centre National de la Recherche Scientifique 1354, Université de Paris-Sud, Batiment 409, Centre d'Orsay, F-91405, France, the
(2) Department of Infectious Diseases and Bacteriology, Royal Postgraduate Medical School, Du Cane Road, London W12 ONN, United Kingdom, and the
(3) Seccion Bioquimica, Facultad de Ciencias, Universidad de la Republica, Montevideo, Uruguay

^§ Supported by a European Union “Human Capital and Mobility” Post-doctoral grant. To whom correspondence and reprint requests should be addressed. Tel.: 33-1-69416356; Fax: 33-1-69417808.

Abstract

In Aspergillus nidulans, loss-of-function mutations in the uapA and azgA genes, encoding the major uric acid-xanthine and hypoxanthine-adenine-guanine permeases, respectively, result in impaired utilization of these purines as sole nitrogen sources. The residual growth of the mutant strains is due to the activity of a broad specificity purine permease. We have identified uapC, the gene coding for this third permease through the isolation of both gain-of-function and loss-of-function mutations. Uptake studies with wild-type and mutant strains confirmed the genetic analysis and showed that the UapC protein contributes 30% and 8-10% to uric acid and hypoxanthine transport rates, respectively. The uapC gene was cloned, its expression studied, its sequence and transcript map established, and the sequence of its putative product analyzed. uapC message accumulation is: (i) weakly induced by 2-thiouric acid; (ii) repressed by ammonium; (iii) dependent on functional uaY and areA regulatory gene products (mediating uric acid induction and nitrogen metabolite repression, respectively); (iv) increased by uapC gain-of-function mutations which specifically, but partially, suppress a leucine to valine mutation in the zinc finger of the protein coded by the areA gene. The putative uapC gene product is a highly hydrophobic protein of 580 amino acids (M_r= 61,251) including 12-14 putative transmembrane segments. The UapC protein is highly similar (58% identity) to the UapA permease and significantly similar (23-34% identity) to a number of bacterial transporters. Comparisons of the sequences and hydropathy profiles of members of this novel family of transporters yield insights into their structure, functionally important residues, and possible evolutionary relationships.

Footnotes

↵^¶ Supported by a grant from the Universidad de la Republica, Montevideo, Uruguay and a European Union grant.
↵** Supported by a grant from the Universidad de la Republica Montevideo, Uruguay.
↵* This work was supported by the Centre National de la Recherche Scientifique and by a European Union grant. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The nucleotide sequence(s) reported in this paper has been submitted to the GenBank/EMBL Data Bank with accession number(s) X79796.
↵¹ U. Tazebay, V. Sophianopoulou, C. Scazzocchio, and G. Diallinas, unpublished results.
↵² The abbreviations used are:

bp

base pair(s)

kb

kilobase(s).
↵³ A. Ravagnani, T. Langdon, D. Gomez, V. Gavrias, H. N. Arst, Jr., C. Scazzocchio, and B. Cubero, unpublished results.
↵⁴ T. Suárez, M. Vieira de Queiroz, N. Oestreicher, and C. Scazzocchio, manuscript submitted for publication.
↵⁵ G. Diallinas, unpublished results.
↵⁶ V. Sophianopoulou and G. Diallinas, unpublished observations.