Analysis of the DNA-binding and dimerization activities of Neurospora crassa transcription factor NUC-1.
NUC-1, a positive regulatory protein of Neurospora crassa, controls the expression of several unlinked target genes involved in phosphorus acquisition. The carboxy-terminal end of the NUC-1 protein has sequence similarity to the helix-loop-helix family of transcription factors. Bacterially expressed and in vitro-synthesized proteins, which consist of the carboxy-terminal portion of NUC-1, bind specifically to upstream sequences of two of its target genes, pho2+ and pho-4+. These upstream sequences contain the core sequence, CACGTG, a target for many helix-loop-helix proteins. A large loop region (47 amino acids) separates the helix I and helix II domains. Mutations and deletion within the loop region did not interfere with the in vitro or in vivo functions of the protein. Immediately carboxy-proximal to the helix II domain, the NUC-1 protein contains an atypical zipper domain which is essential for function. This domain consists of a heptad repeat of alanine and methionine rather than leucine residues. Analysis of mutant NUC-1 proteins suggests that the helix II and the zipper domains are essential for the protein dimerization, whereas the basic and the helix I domains are involved in DNA binding. The helix I domain, even though likely to participate in dimer formation while NUC-1 is bound to DNA, is not essential for in vitro dimerization.
Department of Biomolecular Chemistry, University of Wisconsin-Madison 53706.
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D001483:Base Sequence; D004271:DNA, Fungal / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005656:Fungal Proteins / Q000737:chemistry / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal; D018257:Helix-Loop-Helix Motifs; D008969:Molecular Sequence Data; D009492:Neurospora crassa / Q000235:genetics*; D010710:Phosphates / Q000378:metabolism; D011485:Protein Binding; D015183:Restriction Mapping; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*
D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C079787:NUC-1 protein, Neurospora crassa; D010710:Phosphates; D014157:Transcription Factors