Comparative Proteomics Studies Of Follicular Fluid During The Different Periods Of Follicular Development In Tianzhu White Yak And Bactrian Camel

Tianzhu white yak and Bactrian camel are two important livestock and geneticresources in China. Tianzhu white yak inhabits cold and alpine prairies, Bactriancamel inhabits in the desert and semi-desert areas. Facing such an adversecircumstances, they have their own reproductive characteristics, but the reproductiveperformance is relatively low. Follicular fluid is helpful for understanding oocytequality, fertilization and embryo development potential, and the mechanism offollicular differentiation and development. Proteomics is also a powerful tool toexplain mammalian reproductive physiological mechanism.To explore the method of constructing2-DE map, Tianzhu white yak follicular fluidwas used to compare the effect of different sample preparation methods and differentconcentrations of SDS-PAGE gels on follicular fluid proteins of separation in2-DE.Follicles were divided into different groups based on follicular diameter, and then thefull scale protein expression maps of Tianzhu white yak and Bactrian camel follicularfluid by2-DE were established. The maps were used to detect differentially expressedprotein spots by PDQuest8.0software. Differentially expressed proteins spots wereidentified by mass-spectrometric technique. The normal follicular fluid proteomedatabase was developed based on the follicular fluid proteome data published byNCBI and our own experimental results.It was found that direct solubilization and12％homogeneous gel can be used toconstruct the2-DE map of white yaks follicular fluid proteome.34differentiallyexpressed spots were found during four classes of developmental ovarian follicle inTianzhu white yak. Among them,14were excised from the gels and identified byMALDI-TOF/TOF-MS as: arsenic (+3) methyltransferase, vitamin D binding protein,ankyrin repeat domain-containing protein-6, C10orf107protein, multidrugresistance-associated protein-4-like, mitogen activated protein kinase-2-associatedprotein-1, argonaute-3, profilin-β-actin, SPT2homolog, α-1-acid glycoprotein,obscurin-like-1, aldose reductase, E3ubiquitin-protein ligase RE1A, and one unnamed protein.13differentially expressed spots were found during six classes ofdevelopmental ovarian follicle in Bactrian camel. These spots were identified byLC-MS/MS and found that in addition to one point have not result other pointsrepresent7different proteins: hemoglobin, toll-like receptor9, antithrombin,recombination activating protein1, polyubiquitin, transmembrane and coiled coildomain protein3and γ fibrinogen. At present, follicular fluid proteome of8differentspecies has been studied, which include people, cattle, sheep, pig, dog, horse, Bactriancamel and Tianzhu white yak. The normal follicular fluid proteome database contains324proteins. There are82proteins detected in two or more species. No protein wasfound in8species at the same time. The follicular fluid proteomics have been studiedmost are human being, and human follicular fluid proteome account for78.4％ofentire normal follicular fluid proteome database.It was concluded there was a significant difference at protein level in Tianzhu yhiteyak during follicular development. In these14proteins, arsenic (+3)methyltransferase, mitogen activated protein kinase-2-associated protein-1, α-1-acidglycoprotein, E3ubiquitin-protein ligase BRE1A may be associated with folliculardevelopment. Vitamin D binding protein, argonaute-3may related to oocyte growth.Ankyrin repeat domain-containing protein-6, profilin-β-actin maybe associated withthe second meiotic division of oocyte, need further verification. The function ofaldose reductase, multidrug resistance-associated protein4-like, obscurin-like1,protein SPT2homolog, C10orf107protein and an unnamed protein in reproductionremains ambiguous, need to be further studied.There was a significant difference at protein level in Bactrian camel duringfollicular development. In these7proteins, toll-like receptor9may be associated withfollicular development, ovulation and the developing of embryo. Polyubiquitin andrecombination activating protein1may be related to oocyte growth. Antithrombinmay be related to the coagulation of follicular fluid. γ fibrinogen may be associatedwith pregnancy maintenance, but need further verification. Hemoglobin andtransmembrane and coiled coil domain protein3in reproduction remains ambiguous,need to be further studied. According to analyse follicular fluid datas, we found that so far there is no proteinhave been found at8species, not because there is no common protein between them,but there are few studies on follicular fluid of the other species, and need to be furtherstudied.