Quantitative variations of the mitochondrial proteome and phosphoproteome during fermentative and respiratory growth in Saccharomyces cerevisiae.
The yeast Saccharomyces cerevisiae is a facultative aerobe able to adapt its metabolism according to the carbon substrate. The mechanisms of these adaptations involve at least partly the mitochondria but are not yet well understood. To address the possible role of protein phosphorylation event in their regulation, it is necessary in a first instance to determine precisely the phosphorylation sites that show changes depending on the carbon source. In this aim we performed an overall quantitative proteomic and phosphoproteomic study of isolated mitochondria extracted from yeast grown on fermentative (glucose or galactose) and respiratory (lactate) media. Label free quantitative analysis of protein accumulation revealed significant variation of 176 mitochondrial proteins including 108 proteins less accumulated in glucose medium than in lactate and galactose media. We also showed that the responses to galactose and glucose are not similar. Stable isotope dimethyl labeling allowed the quantitative comparison of phosphorylation levels between the different growth conditions. This study enlarges significantly the map of yeast mitochondrial phosphosites as 670 phosphorylation sites were identified, of which 214 were new and quantified. Above all, we showed that 90 phosphosites displayed a significant variation according to the medium and that variation of phosphorylation level is site-dependent.


CONCLUSIONS
This proteomic and phosphoproteomic study is the first extensive study providing quantitative data on phosphosites responses to different carbon substrates independent of the variations of protein quantities in the yeast S. cerevisiae mitochondria. The significant changes observed in the level of phosphorylation according to the carbon substrate open the way to the study of the regulation of mitochondrial proteins by phosphorylation in fermentative and respiratory media. In addition, the identification of a large number of new phosphorylation sites show that the characterization of mitochondrial phosphoproteome is not yet completed.
CNRS, UMR8221, F-91191 Gif-sur-Yvette, France; CEA, IBITECS, SB(2)SM, LPM, F-91191 Gif-sur-Yvette, France.
Margaux
M
Renvoisé
INRA, PAPPSO, UMR de Génétique Végétale, Gif-sur-Yvette, France.
Ludovic
L
Bonhomme
CNRS, PAPPSO, UMR de Génétique Végétale, Gif-sur-Yvette, France.
Marlène
M
Davanture
INRA, PAPPSO, UMR de Génétique Végétale, Gif-sur-Yvette, France.
Benoit
B
Valot
CNRS, PAPPSO, UMR de Génétique Végétale, Gif-sur-Yvette, France.
Michel
M
Zivy
CNRS, UMR8221, F-91191 Gif-sur-Yvette, France; CEA, IBITECS, SB(2)SM, LPM, F-91191 Gif-sur-Yvette, France. Electronic address: claire.lemaire@cea.fr.
Claire
C
Lemaire
D002244:Carbon / Q000737:chemistry; D002852:Chromatography, Ion Exchange; D002853:Chromatography, Liquid; D005285:Fermentation; D005690:Galactose / Q000737:chemistry; D005947:Glucose / Q000737:chemistry; D007477:Ions; D007773:Lactates / Q000737:chemistry; D008670:Metals / Q000737:chemistry; D008928:Mitochondria / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000378:metabolism*; D010085:Oxidative Phosphorylation; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D020543:Proteome; D040901:Proteomics; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D053719:Tandem Mass Spectrometry; D014357:Trypsin / Q000737:chemistry
D007477:Ions; D007773:Lactates; D008670:Metals; D024101:Mitochondrial Proteins; D010750:Phosphoproteins; D020543:Proteome; D002244:Carbon; D014357:Trypsin; D005947:Glucose; D005690:Galactose
Galactose; Glucose; Lactate; OXPHOS; Phosphoproteomic; Proteomic