Chung Jooho, Serezani Carlos H, Huang Steven K, Stern Joel N H, Keskin Derin B, Jagirdar Rajesh, Brock Thomas G, Aronoff David M, Peters-Golden Marc
Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, University of Michigan Health System, Ann Arbor, MI 48109, USA.
J Immunol. 2008 Oct 15;181(8):5501-9. doi: 10.4049/jimmunol.181.8.5501.
Phagocytosis of IgG-opsonized microbes via the Fc gamma receptor (Fc gammaR) requires the precise coordination of a number of signaling molecules, including the low-molecular mass GTPases. Little is known about the Ras-family GTPase Rap1 in this process. We therefore investigated its importance in mediating Fc gammaR-dependent phagocytosis in NR8383 rat alveolar macrophages. Pulldown of active Rap1 and fluorescence microscopic analysis of GFP-RalGDS (Ral guanine dissociation stimulator)-transfected macrophages revealed that Rap1 is indeed activated by Fc gammaR crosslinking. Inhibition of Rap1 activity, both by Rap1GAP (GTPase-activating protein) expression and liposome-delivered blocking Ab, severely impaired the ability of cells to ingest IgG-opsonized targets. Fc gammaR-induced Rap1 activation was found to be independent of both cAMP and Ca(2+), suggesting a role for the second messenger-independent guanosine exchange factor, C3G. This was supported by the facts that 1) liposome-delivered blocking Ab against C3G inhibited both Fc gammaR-dependent phagocytosis and Rap1 activation, and 2) both active Rap1GTP and C3G were found to translocate to the phagosome. Taken together, our data demonstrate a novel role for Rap1 and its exchange factor C3G in mediating Fc gammaR-dependent phagocytosis.
通过Fcγ受体（FcγR）对IgG调理的微生物进行吞噬作用需要多种信号分子的精确协调，包括低分子量GTP酶。在此过程中，关于Ras家族GTP酶Rap1的了解甚少。因此，我们研究了其在NR8383大鼠肺泡巨噬细胞介导FcγR依赖性吞噬作用中的重要性。对活性Rap1的下拉分析以及对转染了GFP-RalGDS（Ral鸟嘌呤解离刺激因子）的巨噬细胞的荧光显微镜分析表明，Rap1确实通过FcγR交联而被激活。通过Rap1GAP（GTP酶激活蛋白）表达和脂质体递送的阻断抗体抑制Rap1活性，严重损害了细胞摄取IgG调理靶标的能力。发现FcγR诱导的Rap1激活与cAMP和Ca²⁺均无关，提示第二信使非依赖性鸟嘌呤交换因子C3G发挥了作用。以下事实支持了这一点：1）脂质体递送的针对C3G的阻断抗体抑制了FcγR依赖性吞噬作用和Rap1激活；2）发现活性Rap1GTP和C3G均易位至吞噬体。综上所述，我们的数据证明了Rap1及其交换因子C3G在介导FcγR依赖性吞噬作用中的新作用。
