Structure, exon pattern, and chromosome mapping of the gene for cytosolic copper-zinc superoxide dismutase (sod-1) from Neurospora crassa

Abstract

A 4.8-kilobase BamHI-HindIII fragment encoding the entire Neurospora crassa CuZn superoxide dismutase gene (herein designated sod-1) was isolated from a genomic library using two 60-base deoxyoligonucleotide probes corresponding to the published N. crassa amino acid sequence. The nucleotide sequence of the gene encodes an amino acid sequence matching the published protein sequence at 152 of 153 positions. Codon preference shows an unusually strong bias such that only 32 of the possible 61 codons are used, with no codons ending in A. Codon usage is that of highly expressed N. crassa genes. The gene contains three introns, none of which corresponds to any of the introns previously identified in the human gene. Analysis of the intron positions provides support for the hypothesis that CuZn superoxide dismutases evolved by gene duplication and fusion followed by the addition of exons encoding an N-terminal beta-hairpin and a zinc-binding subdomain. The N. crassa gene has an intron mapping to amino acid residue 114 in a sequence-conserved region of the protein whereas the human gene has an intron mapping to a similar but not identical position at residue 118. The discordant position of these introns suggests that one of them was inserted relatively recently. The first N. crassa intron contains a sequence that is similar to the transcriptional regulatory site, UAS1, of the yeast CYC1 (iso-1-cytochrome c) gene and to a putative UAS from the yeast manganese superoxide dismutase gene. A 10-nucleotide portion of this region also matches exactly a sequence in intron 2 of the con-10 gene of N. crassa. sod-1 was mapped to the left arm of chromosome I by following the segregation of a restriction fragment length polymorphism in a sexual cross. Although results indicate that there is a single gene for cytosolic CuZn superoxide dismutase, two additional, perhaps distantly related, sequences were identified that hybridized weakly to both oligonucleotide probes.