Berón W, Colombo M I, Mayorga L S, Stahl P D
Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110.
Arch Biochem Biophys. 1995 Mar 10;317(2):337-42. doi: 10.1006/abbi.1995.1172.
We have assessed the role of heterotrimeric GTPases on in vitro fusion of phagosomes and endosomes. Highly purified phagosomes were found to contain G alpha s, G alpha i1, G alpha i2, G alpha i3, and G beta subunits of heterotrimeric GTP-binding proteins. A functional role for G proteins was established using an in vitro phagosome-endosome fusion assay. First, addition of AlF4- and purified G beta gamma subunits to the in vitro assay blocked fusion, indicating that heterotrimeric G proteins may play a role, either direct or indirect, in phagosome maturation. Second, a striking inhibitory effect was observed when the vesicles were incubated with peptides that preferentially activate G alpha s. A similar effect on phagosome-endosome fusion was observed with cholera toxin, a reagent known to activate G alpha s. Our results suggest that one or more heterotrimeric G proteins, including Gs, mediate and/or regulate phagosome-endosome fusion.
我们评估了异源三聚体GTP酶在吞噬体与内体体外融合中的作用。发现高度纯化的吞噬体含有异源三聚体GTP结合蛋白的Gαs、Gαi1、Gαi2、Gαi3和Gβ亚基。使用体外吞噬体-内体融合试验确定了G蛋白的功能作用。首先，在体外试验中添加AlF4-和纯化的Gβγ亚基可阻断融合，这表明异源三聚体G蛋白可能直接或间接参与吞噬体成熟。其次，当囊泡与优先激活Gαs的肽一起孵育时，观察到显著的抑制作用。用霍乱毒素（一种已知可激活Gαs的试剂）对吞噬体-内体融合也观察到类似的作用。我们的结果表明，一种或多种异源三聚体G蛋白，包括Gs，介导和/或调节吞噬体-内体融合。
