Identification and the role of low molecular weight GTP-binding protein in the rat parotid acinar cells

低分子量GTP结合蛋白Rab4、Rab3D和Rab26主要通过RT-PCR技术从大鼠腮腺提取的mRNA中检测到。在分泌颗粒膜中检测到Rab26和VAMP2(囊泡相关膜蛋白2)，而在上清液和血浆膜中均未检测到。另一方面，Rab4主要在质膜组分中检测到，而在颗粒膜中检测不到。腺泡细胞经异丙肾上腺素处理后，VAMP2从颗粒膜转移到细胞内颗粒中。而Rab26则从颗粒膜上消失，在腺泡细胞中未检测到。免疫组织化学结果也证实了免疫印迹的结果。这些发现表明，Rab26参与了颗粒的调节分泌，功能上属于Rab26组。卡巴胆碱处理腺泡细胞后，Rab4部分从胞内颗粒转移到质膜上。我们刚刚开始揭示Rab4在葡萄糖转运蛋白和/或水通道蛋白5转运中的作用，到目前为止还没有足够的数据来报道。在腮腺分泌中，β-激动剂刺激唾液分泌，而不改变细胞内钙离子。在腮腺中未检测到与Rab3A结合的RabPhilin，提示腮腺的分泌机制不同于钙依赖途径。我们提出了PRA(Prenylated Rab Association Protein)通过磷酸化来调节细胞分泌的机制。

Low moledular weight GTP-binding proteins, Rab4, Rab3D and Rab26, were mainly detected by RT-PCR techniques using mRNA isolated from the rat parotid gland. Rab26 and VAMP2 (vesicle associated membrance protein 2) were detected in secretory granule membrane, but neither in the supernatant nor the plasma membane fractions. On the other hand, Rab4 was mainly detected in the plasma membrane fraction, but not in the granule membrane. After acinar cells were treated with isoproternol, VAMP2 was transferred to the intracellular particles from the granule membrance. However, Rab 26 was disappeared from the granule membrance, and not detected in the acinar cells. The results obtained using western blotting were also confirmed by immuno-histochemistry. These finding suggest that Rab26 participates in the regulated secretion of granules and functionally belongs to the Rab26 group.Treatment of acinar cells with carbachol partially transferred Rab4 to the plasma membrane from the intracellular particle. We have just started to reveal the role of Rab4 on the transportation of glucose transporter and/or aquaporin 5, so far we do not obtained enough data for reporting.In the parotid secretion, beta-agonist stimulates salivary secretion without change of the intracellular calcium ion. And rabphilin, which binds Rab3A, was not detected in the parotid gland, suggesting that the secretory mechanisms in the parotid are different from the calcium dependent pathway. We proposed mechanisms that PRA (prenylated Rab associated protein) regulates the secretion through phosphorylation.