Department of Dermatology, University of Wisconsin, Madison, WI 53706, USA.
Arch Biochem Biophys. 2011 Apr 15;508(2):227-33. doi: 10.1016/j.abb.2011.01.021. Epub 2011 Feb 1.
By virtue of the presence of multiple protein-protein interaction and signaling domains, PDZ proteins play important roles in assembling protein complexes that participate in diverse cell biological processes. GIPC is a versatile PDZ protein that binds a variety of target proteins in different cell types. In previous studies we showed that, in epidermal melanocytes, GIPC interacts with newly synthesized melanosomal protein TRP1 in the Golgi region and proposed that this interaction may facilitate intracellular trafficking of TRP1. However, since GIPC contains a single PDZ domain and no other known protein interaction motifs, it is not known how GIPC-TRP1 interaction affects melanosome biogenesis and/or melanin pigmentation. Here, we show that in human primary melanocytes GIPC interacts with AKT-binding protein APPL (adaptor protein containing pleckstrin homology, leucine zipper and phosphotyrosine binding domains), which readily co-precipitates with newly synthesized TRP1. Knockdown of either GIPC or APPL inhibits melanogenesis by decreasing tyrosinase protein levels and enzyme activity. In melanocytes, APPL exists in a complex with GIPC and phospho-AKT. Inhibition of AKT phosphorylation using a PI3-kinase inhibitor abolishes this interaction and results in retardation TRP1 in the Golgi. These data suggest that interactions between TRP1-GIPC and GIPC-APPL-AKT provide a potential link between melanogenesis and PI3 kinase signaling.
PDZ 蛋白因其具有多个蛋白-蛋白相互作用和信号结构域，在组装参与多种细胞生物学过程的蛋白质复合物方面发挥着重要作用。GIPC 是一种多功能的 PDZ 蛋白，可在不同的细胞类型中结合多种靶蛋白。在之前的研究中，我们表明，在表皮黑素细胞中，GIPC 与高尔基体区域中新合成的黑素体蛋白 TRP1 相互作用，并提出这种相互作用可能有助于 TRP1 的细胞内运输。然而，由于 GIPC 只含有一个 PDZ 结构域，没有其他已知的蛋白质相互作用模体，因此尚不清楚 GIPC-TRP1 相互作用如何影响黑素体的生物发生和/或黑色素沉着。在这里，我们表明在人原代黑素细胞中，GIPC 与 AKT 结合蛋白 APPL（含有 pleckstrin 同源结构域、亮氨酸拉链和磷酸酪氨酸结合结构域的衔接蛋白）相互作用，APPL 可与新合成的 TRP1 轻易共沉淀。敲低 GIPC 或 APPL 均可通过降低酪氨酸酶蛋白水平和酶活性来抑制黑色素生成。在黑素细胞中，APPL 与 GIPC 和磷酸化 AKT 存在复合物。使用 PI3-激酶抑制剂抑制 AKT 磷酸化会破坏这种相互作用，并导致 TRP1 在高尔基体中积累。这些数据表明，TRP1-GIPC 和 GIPC-APPL-AKT 之间的相互作用为黑色素生成和 PI3 激酶信号之间提供了一个潜在的联系。
