Analysis of a distal cluster of binding elements and other unusual features of the promoter of the highly regulated nit-3 gene of Neurospora crassa.
In Neurospora crassa, the expression of the nit-3 gene (nitrate reductase) is dependent upon nitrogen derepression and nitrate induction and is regulated by two positive-acting transcription factors, NIT2 and NIT4, and a negative regulator, NMR. The presence of a tightly linked cluster of NIT2 and NIT4 binding sites suggested that their close spacing might be required for a synergistic interaction of the NIT2 and NIT4 proteins. We show here that the NIT2 and NIT4 binding sites can be separated without affecting either the expression level or the precise regulation of the nit-3 gene. Studies conducted on the NIT2 site II, which contains only a single GATA element and yet plays a major role in nit-3 gene expression, showed that nucleotides both 5' and 3' of the GATA sequence were important for strong DNA binding in vitro and its activation function in vivo. The nit-3 promoter contains two long AT-rich sequences, one of which is located just upstream of the transcription start sites and is required for optimal promoter function. The nit-3 transcript contains eight TACC repeats in its 5' noncoding region which appear to be involved in mRNA instability. Deletion of these TACC repeats led to a significant increase in the stability of nit-3 mRNA.
Tao
Y
Y
Department of Biochemistry, Molecular Cellular and Developmental Biology Program, The Ohio State University, Columbus 43210, USA.
Marzluf
G A
GA
D001665:Binding Sites / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics*; D005656:Fungal Proteins; D015966:Gene Expression Regulation, Fungal; D005810:Multigene Family; D009492:Neurospora crassa / Q000201:enzymology / Q000235:genetics*; D050901:Nitrate Reductase; D009565:Nitrate Reductases / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012045:Regulatory Sequences, Nucleic Acid; D012091:Repetitive Sequences, Nucleic Acid; D015534:Trans-Activators; D014157:Transcription Factors / Q000235:genetics
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C068100:NIT2 protein, Neurospora crassa; D015534:Trans-Activators; D014157:Transcription Factors; C071240:NIT-4 protein, Neurospora crassa; D009565:Nitrate Reductases; D050901:Nitrate Reductase