Lai Joey, Bernhard Oliver K, Turville Stuart G, Harman Andrew N, Wilkinson John, Cunningham Anthony L
Centre for Virus Research, Westmead Millennium Institute, Westmead Hospital, Westmead, Sydney, New South Wales 2145, Australia.
J Biol Chem. 2009 Apr 24;284(17):11027-38. doi: 10.1074/jbc.M809698200. Epub 2009 Feb 17.
C-type lectin receptors expressed on the surface of dendritic cells and macrophages are able to bind glycoproteins of microbial pathogens via mannose, fucose, and N-acetylglucosamine. Langerin on Langerhans cells, dendritic cell-specific intercellular adhesion molecule 3-grabbing nonintegrin on dendritic cells, and mannose receptor (MR) on dendritic cells and macrophages bind the human immunodeficiency virus (HIV) envelope protein gp120 principally via high mannose oligosaccharides. These C-type lectin receptors can also oligomerize to facilitate enhanced ligand binding. This study examined the effect of oligomerization of MR on its ability to bind to mannan, monomeric gp120, native trimeric gp140, and HIV type 1 BaL. Mass spectrometry analysis of cross-linked MR showed homodimerization on the surface of primary monocyte-derived dendritic cells and macrophages. Both monomeric and dimeric MR were precipitated by mannan, but only the dimeric form was co-immunoprecipitated by gp120. These results were confirmed independently by flow cytometry analysis of soluble monomeric and trimeric HIV envelope and a cellular HIV virion capture assay. As expected, mannan bound to the carbohydrate recognition domains of MR dimers mostly in a calcium-dependent fashion. Unexpectedly, gp120-mediated binding of HIV to dimers on MR-transfected Rat-6 cells and macrophages was not calcium-dependent, was only partially blocked by mannan, and was also partially inhibited by N-acetylgalactosamine 4-sulfate. Thus gp120-mediated HIV binding occurs via the calcium-dependent, non-calcium-dependent carbohydrate recognition domains and the cysteine-rich domain at the C terminus of MR dimers, presenting a much broader target for potential inhibitors of gp120-MR binding.
树突状细胞和巨噬细胞表面表达的C型凝集素受体能够通过甘露糖、岩藻糖和N - 乙酰葡糖胺结合微生物病原体的糖蛋白。朗格汉斯细胞上的朗格蛋白、树突状细胞上的树突状细胞特异性细胞间黏附分子3抓取非整合素以及树突状细胞和巨噬细胞上的甘露糖受体（MR）主要通过高甘露糖寡糖结合人类免疫缺陷病毒（HIV）包膜蛋白gp120。这些C型凝集素受体也能寡聚化以促进增强的配体结合。本研究检测了MR寡聚化对其结合甘露聚糖、单体gp120、天然三聚体gp140和1型HIV BaL能力的影响。交联MR的质谱分析显示在原代单核细胞衍生的树突状细胞和巨噬细胞表面存在同型二聚化。甘露聚糖可沉淀单体和二聚体形式的MR，但只有二聚体形式能被gp120共免疫沉淀。这些结果通过可溶性单体和三聚体HIV包膜的流式细胞术分析以及细胞HIV病毒体捕获试验得到了独立证实。正如预期的那样，甘露聚糖大多以钙依赖的方式结合到MR二聚体的碳水化合物识别结构域。出乎意料的是，gp120介导的HIV与转染MR的大鼠 - 6细胞和巨噬细胞上二聚体的结合不依赖钙，仅部分被甘露聚糖阻断，也部分被4 - 硫酸N - 乙酰半乳糖胺抑制。因此，gp120介导的HIV结合通过MR二聚体C末端的钙依赖、非钙依赖碳水化合物识别结构域和富含半胱氨酸的结构域发生，为gp120 - MR结合的潜在抑制剂提供了更广泛的靶点。
