A highly representative two-hybrid genomic library for the yeast Yarrowia lipolytica.
Since its description by Fields and Song in 1989 (Nature 340, 245-246), the yeast two-hybrid system has been used extensively to study protein-protein interactions, becoming increasingly efficient with technological and methodological improvements. Here, we report the construction of a highly representative two-hybrid genomic library for the dimorphic yeast Yarrowia lipolytica based on the system described by James et al. (1996. Genetics 144, 1425-1436). The endoplasmic reticulum protein Slslp was then used as a bait in a functional test of the library. Indeed, we previously showed that the SLS1 gene product is involved in protein translocation across the endoplasmic reticulum membrane and interacts physically in a two-hybrid assay with Kar2p, an essential luminal member of the HSP70 family (Boisramé et al., 1998. J. Biol. Chem. 273, 30 903-30 908). We developed a mating strategy similar to that used for the Saccharomyces cerevisiae FRYL library (Fromont-Racine et al., 1997. Nat. Genet. 16, 277-282). No other partner than Kar2p was identified in this screen. As an interesting result, Kar2p interacts with Slslp through its ATPase domain, supporting our hypothesis that Slslp is a cofactor of the chaperone protein, modulating its activity during the HSP70 cycle. Our results indicate that we have constructed a new and powerful tool for the study of Yarrowia lipolytica, which we believe is a good alternative model to investigate such complex biological processes as secretion pathways.
Kabani
M
M
Laboratoire de Génétique Moléculaire et Cellulaire, INRA-INA.PG-CNRS BP 01 78850, Thiverval-Grignon, France. kabani@platon.grignon.inra.fr
Boisramé
A
A
Beckerich
J M
JM
Gaillardin
C
C
D002352:Carrier Proteins / Q000378:metabolism; D004262:DNA Restriction Enzymes; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005656:Fungal Proteins / Q000378:metabolism; D005822:Genetic Vectors; D015698:Genomic Library; D008565:Membrane Proteins; D024101:Mitochondrial Proteins; D011485:Protein Binding; D029701:Saccharomyces cerevisiae Proteins; D004718:Saccharomycetales / Q000235:genetics*; D013045:Species Specificity; D020798:Two-Hybrid System Techniques
D002352:Carrier Proteins; D005656:Fungal Proteins; D008565:Membrane Proteins; D024101:Mitochondrial Proteins; C492046:SLS1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D004262:DNA Restriction Enzymes