Identification and characterization of a nitrate transporter gene in Neurospora crassa.
The Neurospora crassa genome database was searched for sequence similarity to crnA, a nitrate transporter in Aspergillus nidulans. A 3.9-kb fragment (contig 3.416, subsequence 183190-187090) was cloned by PCR. The gene coding for this nitrate transporter was termed nit-10. The nit-10 gene specifies a predicted polypeptide containing 541 amino acids with a molecular mass of 57 kDa. In contrast to crnA, which is clustered together with niaD, encoding nitrate reductase, and niiA, encoding nitrite reductase, nit-10 is not linked to nit-3 (nitrate reductase), nit-6 (nitrite reductase), or to nit-2, nit-4 (both are positive regulators of nit-3), or nmr (negative regulator of nit-3) in Neurospora crassa. A nit-10 rip mutant failed to grow in the medium when nitrate (< 10 mM) was used as the sole nitrogen source, but grew similarly to wild type when nitrate concentration was 10 mM or higher. In addition, it showed strong sensitivity to cesium in the presence of nitrate and resistance to chlorate in the presence of alanine, proline, or hypoxanthine. The expression of nit-10 required nitrate induction and was subject to repression by nitrogen metabolites such as glutamine. Expression of nit-10 also required functional products of nit-2 and nit-4. The half-life of nit-10 mRNA was determined to be approximately 2.5 min.
Gao-Rubinelli
Fei
F
Department of Biochemistry, Ohio State University, Columbus, Ohio 43210, USA.
Marzluf
George A
GA
D000409:Alanine / Q000378:metabolism / Q000494:pharmacology; D000595:Amino Acid Sequence; D027321:Anion Transport Proteins / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D002586:Cesium / Q000378:metabolism / Q000494:pharmacology; D002704:Chlorates / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D005973:Glutamine / Q000378:metabolism / Q000494:pharmacology; D006207:Half-Life; D019271:Hypoxanthine / Q000378:metabolism / Q000494:pharmacology; D008969:Molecular Sequence Data; D009154:Mutation; D009492:Neurospora crassa / Q000187:drug effects / Q000235:genetics*; D000091285:Nitrate Transporters; D012150:Polymorphism, Restriction Fragment Length; D011392:Proline / Q000378:metabolism / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
D027321:Anion Transport Proteins; D002704:Chlorates; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C068100:NIT2 protein, Neurospora crassa; D000091285:Nitrate Transporters; D012333:RNA, Messenger; D015534:Trans-Activators; D014157:Transcription Factors; D005973:Glutamine; C071240:NIT-4 protein, Neurospora crassa; D002586:Cesium; D019271:Hypoxanthine; D011392:Proline; D000409:Alanine