The green alga Chlamydomonas reinhardtii: a new model system to unravel the biogenesis of respiratory complexes

The general purpose of this research is to contribute to a better understanding of the mitochondrial NADH: ubiquinone oxidoreductase (Complex I) and to illustrate the current view of its assembly process. I propose to use the green alga Chlamydomonas reinhardtii as a novel model system to carry out the molecular dissection of Complex I assembly. The main objective is to discover novel genes controlling the assembly process of this multimeric enzyme.

Several reasons, including patient death at young age, strict regulations, and ethical concerns, make the study of Complex I on humans highly difficult. Although traditionally well established model systems such as the fungi Yarrowia lipolytica and Neurospora crassa and human cell lines have proven useful to understand the role of specific subunits in the assembly process, forward genetic approaches leading to discovery of novel assembly factors have been limited by the lack of straightforward screening methodologies (REMACLE et al. 2008). The process of plant Complex I assembly is at present largely obscure, despite the progress achieved in recent years. In plants, similarly to other eukaryotes, there is no simple strategy to approach the issue of assembly. Thus, the development of a model system where both mitochondrial and nuclear genomes can be manipulated is highly desirable for the study of Complex I.

Chapter 2 shows the use of Chlamydomonas reinhardtii as a forward genetics tool to screen for Complex I deficiency mutants and the finding of six new loci that result in defects in assembly. Based on large level of conservation of Complex I subunits in eukaryotes as well as the presence of homologs of currently proposed assembly factors, this work follows the hypothesis that the assembly process is also conserved among eukaryotes. Chapter 3 refers to the molecular identification of one of these genes as the Complex I subunit NUOB10, homolog of the bovine PSDW subunit. Also, a molecular mapping approach for another mutant suggests that the mutation does not affect a structural subunit of Complex I and that the final identification of this gene will lead to the discovery of a novel assembly factor. Finally, Chapter 4 provides an initial evaluation for the overall functional conservation of two known assembly factors: CIA30 and OXA1, both of which have a ortholog gene in Chlamydomonas.