Patrick Dessi | PubMed Health

PubMed2005

A combined approach exploring gene function based on worm-human orthology.

BACKGROUND: Many aspects of the nematode Caenorhabditis elegans biology are conserved between invertebrates and vertebrates establishing this particular organism as an excellent genetic model. Because of its small size, large populations and self-fertilization of the hermaphrodite, functional predictions carried out by genetic modifications as well as RNAi screens, can be rapidly tested. RESULTS: In order to explore the function of a set of C. elegans genes of unknown function, as well as their potential functional roles in the human genome, we performed a phylogenetic analysis to select the most probable worm orthologs. A total of 13 C. elegans genes were subjected to down-regulation via RNAi and characterization of expression profiles using GFP strains. Previously unknown distinct expression patterns were observed for four of the analyzed genes, as well as four visible RNAi phenotypes. In addition, subcellular protein over-expression profiles of the human orthologs for seven out of the thirteen genes using human cells were also analyzed. CONCLUSION: By combining a whole-organism approach using C. elegans with complementary experimental work done on human cell lines, this analysis extends currently available information on the selected set of genes.

Animals↗

PubMed2003

Investigations on the in vitro import ability of mitochondrial precursor proteins synthesized in wheat germ transcription-translation extract.

Mitochondrial precursor proteins synthesized in rabbit reticulocyte lysate (RRL) are readily imported into mitochondria, whereas the same precursors synthesized in wheat germ extract (WGE) fail to be imported. We have investigated factors that render import incompetence from WGE. A precursor that does not require addition of extramitochondrial ATP for import, the F(A)d ATP synthase subunit, is imported from WGE. Import of chimeric constructs between precursors of the F(A)d protein and alternative oxidase (AOX) with switched presequences revealed that the mature domain of the F(A)d precursor defines the import competence in WGE as only the construct containing the presequence of AOX and mature portion of F(A)d (pAOX-mF(A)d) could be imported. Import competence of F(A)d and pAOX-mF(A)d correlated with solubility of these precursors in WGE, however, solubilization of import-incompetent precursors with urea did not restore import competence. Addition of RRL to WGE-synthesized precursors did not stimulate import but addition of WGE to the RRL-synthesized precursors or to the over-expressed mitochondrial precursor derived from the F1beta ATP synthase precursor inhibited import into mitochondria. The dual-targeted glutathione reductase precursor synthesized in WGE was imported into chloroplasts, but not into mitochondria. Antibodies against the 14-3-3 guidance complex characterized for chloroplast targeting were able to immunoprecipitate all of the precursors tested except the F(A)d ATP synthase precursor. Our results point to the conclusion that the import incompetence of WGE-synthesized mitochondrial precursors is not presequence dependent and is a result of interaction of WGE inhibitory factors with the mature portion of precursor proteins.

14-3-3 Proteins↗