A high-throughput screening system for genes extending life-span.
We developed a high-throughput functional genomic screening system that allows identification of genes prolonging life-span in the baker's yeast Saccharomyces cerevisiae. The method is based on isolating yeast mother cells with extended number of cell divisions as indicated by the increased number of bud scars on their surface. Fluorescently labelled Wheat Germ Agglutinin was used for specific staining of chitin, a major component of bud scars. Screening of a human HepG2 cDNA expression library in yeast resulted in the isolation of 12 yeast transformants with a potentially prolonged life-span. The transgene in one of the lines was identified as ferritin light chain (FTL) and studied in more detail. Yeast cells containing FTL showed an enhanced iron and H(2)O(2) resistance, a reduced cell death rate and an increased number of cell divisions. Overexpression of FTL in the nematode Caenorhabditis elegans resulted in a life-span increase of 8% confirming our yeast observations in a multicellular organism. Our data demonstrate that this method permits a fast screening of libraries for hunting genes involved in ageing processes.
Chen
Cuiying
C
Fundamental and Applied Molecular Biology, Ghent University and Flanders Interuniversity Institute for Biotechnology, Ghent University, B-9000, Ghent, Belgium.
Dewaele
Sylviane
S
Braeckman
Bart
B
Desmyter
Liesbeth
L
Verstraelen
Jan
J
Borgonie
Gaetan
G
Vanfleteren
Jacques
J
Contreras
Roland
R
D005293:Ferritins / Q000235:genetics; D005434:Flow Cytometry; D015723:Gene Library; D005800:Genes, Fungal; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007501:Iron / Q000494:pharmacology; D008136:Longevity / Q000235:genetics; D008954:Models, Biological; D018384:Oxidative Stress; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development; D013997:Time Factors; D019076:Transgenes; D014909:Wheat Germ Agglutinins
D014909:Wheat Germ Agglutinins; D005293:Ferritins; D006861:Hydrogen Peroxide; D007501:Iron