Disruption of the nuclear gene encoding the 20.8-kDa subunit of NADH:ubiquinone reductase of Neurospora mitochondria

• Type: Article in International Scientific Journal
• Date: 1996

Authors

dasilva, mv (Author) ICBAS; alves, pc (Author) FCUP; duarte, m (Author) Other; mota, n (Author) Other; lobodacunha, a (Author) Other; harkness, taa (Author) Other; nargang, fe (Author) Other; videira, a (Author) ICBAS

Journal

Title: Molecular and General Genetics

Vol. 252

Pages: 177-183

ISSN: 0026-8925

Publisher: Springer

Indexing

ISI Web of Knowledge - 22 Citations

Scopus - 21 Citations

Scientific classification

FOS: Natural sciences > Biological sciences

Other information

Authenticus ID: P-001-DV2

DOI: 10.1007/s004389670020

Abstract (EN): The nuclear gene coding for the 20.8-kDa subunit of the membrane arm of respiratory chain NADH:ubiquinone reductase (Complex I) from Neurospora, nuo-20.8, was localized on linkage group I of the fungal genome. A genomic DNA fragment containing this gene was cloned and a duplication was created in a strain of N. crassa by transformation. To generate RIP (repeat-induced point) mutations in the duplicated sequence, the transformant was crossed with another strain carrying an auxotrophic marker on chromosome I. To increase the chance of finding an isolate with a non-functional nuo-20.8 gene, random progeny from the cross were selected against this auxotrophy since RIP of the target gene will only occur in the nucleus carrying the duplication. Among these, we isolated and characterised a mutant strain that lacks the 20.8 kDa mitochondrial protein, indicating that this cysteine-rich polypeptide is not essential. Nevertheless, the absence of the 20.8-kDa subunit prevents the full assembly of complex I. It appears that the peripheral arm and two intermediates of the membrane arm of the enzyme are still formed in the mutant mitochondria. The NADH:ubiquinone reductase activity of sonicated mitochondria from the mutant is rotenone insensitive. Electron microscopy of mutant mitochondria does not reveal any alteration in the structure or numbers of the organelles.

Language: English

Type (Professor's evaluation): Scientific

No. of pages: 7