Highly sensitive saturation labeling reveals changes in abundance of cell cycle-associated proteins and redox enzyme variants during oocyte maturation in vitro.
Oocyte maturation is a complex process and a critical issue in assisted reproduction techniques (ART) in humans and other mammals. We used a sensitive 2-D DIGE saturation labeling approach including an internal pooled standard for quantitative proteome profiling of immature versus in vitro matured bovine oocytes in six independent samples. The study comprised 48 2D gel images representing 24 DIGE experiments. From 250 ng sample analyzed per gel, quantitative analysis revealed an average of 2244 spots in pH 4-7 images and 1291 spots in pH 6-9 images. Thirty-eight spots with different intensities were detected in total. Spots of a preparative gel from 2200 oocytes were identified by nano-LC-MS/MS analysis. The ten spots which could be unambiguously identified include the Ca2+-binding protein translationally controlled tumor protein, enzymes of the Krebs and pentose phosphate cycles, clusterin, 14-3-3 epsilon, elongation factor-1 gamma, and redox enzymes such as polymorphic forms of GST Mu 5 and peroxiredoxin-3. The cellular distribution of two proteins was determined by confocal laser scanning microscopy. The interesting protein candidates identified by this study may help to improve the in vitro maturation process in order to increase the rate of successful in vitro fertilization and other ART in cattle and other mammals.
Berendt
Frank J
FJ
Laboratory for Functional Genome Analysis (LAFUGA), Gene Center, LMU Munich, Munich, Germany.
Fröhlich
Thomas
T
Bolbrinker
Patrick
P
Boelhauve
Marc
M
Güngör
Tuna
T
Habermann
Felix A
FA
Wolf
Eckhard
E
Arnold
Georg J
GJ
D000818:Animals; D002417:Cattle; D018797:Cell Cycle Proteins / Q000032:analysis*; D002853:Chromatography, Liquid; D015180:Electrophoresis, Gel, Two-Dimensional; D004798:Enzymes / Q000032:analysis*; D005260:Female; D018613:Microscopy, Confocal; D009866:Oogenesis / Q000502:physiology*; D010084:Oxidation-Reduction; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods*
D018797:Cell Cycle Proteins; D004798:Enzymes; D020543:Proteome