| Abstract: | We have determined the DNA sequence of the oxi-3 gene and its 5' flanking region in the extranuclear [mi-3] mutant of Neurospora crassa. The oxi-3 gene encodes subunit 1 of cytochrome c oxidase, a protein known to be altered in the [mi-3] mutant (Bertrand, H., and Werner, S. (1979) Eur. J. Biochem. 98, 9-18). When the sequence from [mi-3] was compared to previously published sequences of the same region of mtDNA from wild-type N. crassa, a total of five differences was found. Four of these differences can be accounted for as either genetic polymorphisms or previous errors in DNA sequence determination. The remaining difference is a G/C to T/A transversion that changes a codon specifying an aspartic acid residue (GAC) to one that would specify tyrosine (TAC) at amino acid 448 of the 555 amino acid mature subunit 1 protein. This alteration was also found in the mtDNA of two separate heterokaryotic strains that had acquired the [mi-3] phenotype after repeated subculturing of heterokaryons forced between an [mi-3] strain and a strain containing a wild-type cytoplasm. The particular aspartic acid residue that would be affected by the mutation observed in [mi-3] is conserved in a diversity of species as either aspartic acid or glutamic acid, suggesting that an acidic residue at this position is important for the correct function of the subunit 1 protein. For these reasons, we consider it likely that the observed missense mutation is responsible for the [mi-3] phenotype. |
