【摘要】 14-3-3γ作为蛋白质与蛋白质之间的"桥梁蛋白",广泛参与细胞凋亡、细胞分裂、信号转导和蛋白质合成等生物体所有的重要生命活动的调节。以体外培养的荷斯坦奶牛(Vaccas)乳腺上皮细胞(dairy cow mammary gland epithelial cells,DCMECs)为模型,研究14-3-3γ对奶牛乳腺上皮细胞乳蛋白、乳脂合成的影响。瞬时和稳定转染p GCMV/IRES/EGFP/14-3-3γ即14-3-3γ过表达的奶牛乳腺上皮细胞,应用CASY细胞活力分析仪分析细胞活力的变化,测定细胞中甘油三脂(triglyceride,TG)和β-酪蛋白(β-Casein,CSN2)的合成,并且通过Western blot检测DCMECs中m TOR(mammalian target of rapamycin,m TOR)和p-m TOR的表达量,探究14-3-3γ影响奶牛乳腺上皮细胞生理功能的机制。结果显示,瞬时和稳定转染p GCMV/IRES/EGFP/14-3-3γ的奶牛乳腺上皮细胞相对正常培养的奶牛乳腺上皮细胞,细胞活力极显著增强(P<0.01),CSN2和TG表达量极显著增加(P<0.01),同时m TOR和p-m TOR的表达量也显著增加(P<0.01)。研究结果表明14-3-3γ参与泌乳关键信号通路即m TOR信号通路,能够增强乳腺上皮细胞活力,促进奶牛乳腺上皮细胞CSN2和TG的合成,丰富了奶牛泌乳信号通路,为乳品质研究提供了新的理论依据。更多还原

【Abstract】 As a linker of proteins,14-3-3γ plays critical roles in cell signal transductions,cell growth,differentiation and protein synthesis. Based on cultured dairy cow mammary gland epithelial cells in vitro,the impact of 14-3-3γ on the physiological function of dairy cow mammary gland epithelial cells were ivestigated.Expressing plasmids of 14-3-3γ gene were constructed,then stably and transiently transfected into dairy cow mammary gland epithelial cells. The effect of over-expression on dairy cow mammary gland epithelial cells was analyzed by CASY. Triglyceride was detected by triglyceride GPO-POD assay and beta casein was detected by high performance liquid chromatography. Expression of m TOR and p-m TOR was detected by Western blot.Experimental results show that the viability of dairy cow mammary gland epithelial cells,the expression of triglyceride and β-casein,the expression of m TOR and p-m TOR was identified up-expressed( P < 0. 01). All above results indicated that 14-3-3γ regulates the physiological function of dairy cow mammary gland epithelial cells through m TOR signaling pathway. The result enriches the lactation signaling pathway,which provides theoretical basis for improving the quality of milk.更多还原