Cloning and characterization of genes involved in the regulation of the alternative oxidase and mitochondrial DNA replication in Neurospora crassa.

摘要

The mitochondria of Neurospora crassa contain two respiratory pathways, the normal electron-transport chain and an inducible cyanide-resistant alternative oxidase pathway. In Neurospora, the assembly of the alternative oxidase pathway requires the expression of two genes, aod-1, which encodes the alternative oxidase, and aod-2, which is involved in the transcriptional regulation of aod-1. The objective of this work was the cloning and characterization of the aod-2 gene. An attempt was made to isolate the aod-2 + DNA in two different ways: chromosome walking, and by complementation of an aod-2 mutant using aod-1 promoter-driven expression of the bacterial hygromycin resistance gene as a method for selecting transformants;Most of the proteins involved in mitochondrial gene replication and expression are encoded by nuclear genes. I have cloned, sequenced and partially characterized the DNA polymerase-gamma gene (mip-1) that codes for the presumptive mitochondrial DNA polymerase of Neurospora crassa. Sequence analysis showed that mip-1 includes a 3918-nucleotide open reading frame encoding a 1305 amino-acid, 146 kDa protein that has a mitochondrial targeting element at the amino terminus.