Department of Experimental Diabetology, German Institute of Human Nutrition Potsdam-Rehbruecke (DIfE), 14558 Nuthetal, Germany; German Center for Diabetes Research, München-Neuherberg, 85764 Neuherberg, Germany.
Department of Pharmacology, Max Planck Institute for Heart and Lung Research, 61231 Bad Nauheim, Germany.
Mol Metab. 2018 Feb;8:167-179. doi: 10.1016/j.molmet.2017.11.011. Epub 2017 Nov 22.
Intracellular vesicle trafficking maintains cellular structures and functions. The assembly of cargo-laden vesicles at the trans-Golgi network is initiated by the ARF family of small GTPases. Here, we demonstrate the role of the trans-Golgi localized monomeric GTPase ARFRP1 in endosomal-mediated vesicle trafficking of mature adipocytes.
Control (Arfrp1) and inducible fat-specific Arfrp1 knockout (Arfrp1) mice were metabolically characterized. In vitro experiments on mature 3T3-L1 cells and primary mouse adipocytes were conducted to validate the impact of ARFRP1 on localization of adiponectin and the insulin receptor. Finally, secretion and transferrin-based uptake and recycling assays were performed with HeLa and HeLa M-C1 cells.
We identified the ARFRP1-based sorting machinery to be involved in vesicle trafficking relying on the endosomal compartment for cell surface delivery. Secretion of adiponectin from fat depots was selectively reduced in Arfrp1 mice, and Arfrp1-depleted 3T3-L1 adipocytes revealed an accumulation of adiponectin in Rab11-positive endosomes. Plasma adiponectin deficiency of Arfrp1 mice resulted in deteriorated hepatic insulin sensitivity, increased gluconeogenesis and elevated fasting blood glucose levels. Additionally, the insulin receptor, undergoing endocytic recycling after ligand binding, was less abundant at the plasma membrane of adipocytes lacking Arfrp1. This had detrimental effects on adipose insulin signaling, followed by insufficient suppression of basal lipolytic activity and impaired adipose tissue expansion.
Our findings suggest that adiponectin secretion and insulin receptor surface targeting utilize the same post-Golgi trafficking pathways that are essential for an appropriate systemic insulin sensitivity and glucose homeostasis.
细胞内囊泡运输维持着细胞的结构和功能。货物装载囊泡在高尔基网络的组装是由 ARF 家族的小分子 GTP 酶启动的。在这里，我们证明了高尔基体定位的单体 GTP 酶 ARFRP1 在成熟脂肪细胞的内体介导的囊泡运输中的作用。
对对照（Arfrp1）和诱导型脂肪特异性 Arfrp1 敲除（Arfrp1）小鼠进行代谢特征分析。在成熟的 3T3-L1 细胞和原代小鼠脂肪细胞中的体外实验，验证了 ARFRP1 对脂联素和胰岛素受体定位的影响。最后，使用 HeLa 和 HeLa M-C1 细胞进行分泌和转铁蛋白摄取和回收实验。
我们确定了 ARFRP1 为基础的分拣机制参与了依赖于内体区室进行细胞表面递呈的囊泡运输。脂肪组织中脂联素的分泌在 Arfrp1 小鼠中被选择性地减少，而 Arfrp1 耗尽的 3T3-L1 脂肪细胞中，脂联素在 Rab11 阳性内体中积累。Arfrp1 小鼠的血浆脂联素缺乏导致肝胰岛素敏感性恶化、糖异生增加和空腹血糖水平升高。此外，配体结合后经历内吞体循环的胰岛素受体在缺乏 Arfrp1 的脂肪细胞的质膜中含量较少。这对脂肪胰岛素信号传递产生了不利影响，随后导致基础脂肪分解活性的抑制不足和脂肪组织扩张受损。
我们的研究结果表明，脂联素的分泌和胰岛素受体的表面靶向作用利用了相同的高尔基后运输途径，这对于适当的全身胰岛素敏感性和葡萄糖稳态至关重要。
