nit-4, a pathway-specific regulatory gene of Neurospora crassa, encodes a protein with a putative binuclear zinc DNA-binding domain.
nit-4, a pathway-specific regulatory gene in the nitrogen circuit of Neurospora crassa, is required for the expression of nit-3 and nit-6, the structural genes which encode nitrate and nitrite reductase, respectively. The complete nucleotide sequence of the nit-4 gene has been determined. The predicted NIT4 protein contains 1,090 amino acids and appears to possess a single Zn(II)2Cys6 binuclear-type zinc finger, which may mediate DNA binding. Site-directed mutagenesis studies demonstrated that cysteine and other conserved amino acid residues in this possible DNA-binding domain are necessary for nit-4 function. A stretch of 27 glutamines, encoded by a CAGCAA repeating sequence, occurs in the C terminus of the NIT4 protein, and a second glutamine-rich domain occurs further upstream. A NIT4 protein deleted for the polyglutamine region was still functional in vivo. However, nit-4 function was abolished when both the polyglutamine region and the glutamine-rich domain were deleted, suggesting that the glutamine-rich domain might function in transcriptional activation. The homologous regulatory gene from Aspergillus nidulans, nirA, encodes a protein whose amino-terminal half has approximately 60% amino acid identity with NIT4 but whose carboxy terminus is completely different. A hybrid nit-4-nirA gene was constructed and found to function in N. crassa.
Yuan
G F
GF
Department of Biochemistry and Molecular Genetics, Ohio State University, Columbus 43210.
Fu
Y H
YH
Marzluf
G A
GA
D000595:Amino Acid Sequence; D001483:Base Sequence; D002872:Chromosome Deletion; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics / Q000302:isolation & purification; D004268:DNA-Binding Proteins / Q000235:genetics*; D005656:Fungal Proteins; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal; D005809:Genes, Regulator; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D050901:Nitrate Reductase; D009565:Nitrate Reductases / Q000235:genetics; D009572:Nitrite Reductases / Q000235:genetics; D009841:Oligonucleotides; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D012689:Sequence Homology, Nucleic Acid; D015534:Trans-Activators; D016335:Zinc Fingers / Q000235:genetics*
D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D009841:Oligonucleotides; D015534:Trans-Activators; C071240:NIT-4 protein, Neurospora crassa; D009565:Nitrate Reductases; D009572:Nitrite Reductases; D050901:Nitrate Reductase