DNA binding site specificity of the Neurospora global nitrogen regulatory protein NIT2: analysis with mutated binding sites.
NIT2, a positive-acting regulatory protein in Neurospora crassa, activates the expression of a series of unlinked structural genes that encode nitrogen catabolic enzymes. NIT2 binding sites in the promoter regions of nit3, alc and lao have at least two GATA sequence elements. We have examined the binding affinity of the NIT2 protein for the yeast DAL5 wild-type upstream activation sequence UASNTR, which contains two GATA elements, and for a series of mutated binding sites, each differing from the wild-type site by a single base. Substitution for individual nucleotides within 5' or 3' sequences that flank the GATA elements had only modest effects upon NIT2 binding. In contrast, nearly all substitutions within the GATA elements almost completely eliminated NIT2 binding, demonstrating the importance of the GATA sequence for NIT2 binding. Four high-affinity binding sites for the NIT2 protein were found within a central region of the nit-2 gene itself.
Chiang
T Y
TY
Department of Biochemistry, Ohio State University, Columbus 43210.
Rai
R
R
Cooper
T G
TG
Marzluf
G A
GA
D001483:Base Sequence; D001665:Binding Sites; D004271:DNA, Fungal / Q000235:genetics; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005656:Fungal Proteins / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal; D008969:Molecular Sequence Data; D009492:Neurospora crassa / Q000235:genetics* / Q000378:metabolism; D009584:Nitrogen / Q000378:metabolism; D009586:Nitrogen Fixation / Q000235:genetics; D011401:Promoter Regions, Genetic; D014157:Transcription Factors / Q000378:metabolism*; D016335:Zinc Fingers
D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C068100:NIT2 protein, Neurospora crassa; D014157:Transcription Factors; D009584:Nitrogen