Respiratory chain complex I is essential for sexual development in neurospora and binding of iron sulfur clusters are required for enzyme assembly

We have cloned and disrupted in vivo, by repeat-induced point mutations, th e nuclear gene coding for an iron sulfur subunit of complex I from Neurospo ra crassa, homologue of the mammalian TYKY protein. Analysis of the obtaine d mutant nuo21.3c revealed that complex I fails to assemble. The peripheral arm of the enzyme is disrupted while its membrane arm accumulates. Further more, mutated 21.3c-kD proteins, in which selected cysteine residues were s ubstituted with alanines or serines, were expressed in mutant nuo21.3c. The phenotypes of these strains regarding the formation of complex I are simil ar to that of the original mutant, indicating that binding of iron sulfur c enters to protein subunits is a prerequisite for complex I assembly. Homozy gous crosses of nuo21.3c strain, and of other complex I mutants, are unable to complete sexual development. The crosses are blocked at an early develo pmental stage, before fusion of the nuclei of opposite mating types. This p henotype can be rescued only by transformation with the intact gene. Our re sults suggest that this might be due to the compromised capacity of complex I-defective strains in energy production.