Jackson D G, Bell J I, Dickinson R, Timans J, Shields J, Whittle N
Molecular Immunology Group, John Radcliffe Hospital, University of Oxford.
J Cell Biol. 1995 Feb;128(4):673-85. doi: 10.1083/jcb.128.4.673.
The CD44 cell surface glycoprotein is expressed on a broad range of different tissues as multiple isoforms containing from one to ten alternatively spliced exons v1-v10 inserted within the extracellular domain. Differential glycosylation generates still further variability, yielding both N- and O-glycan-modified forms of CD44 in addition to proteoglycan-like variants containing chondroitin sulphate and heparan sulphate. These high molecular mass proteoglycan-like variants, previously identified in lymphocytes, melanomas, and keratinocytes have been implicated in cell-matrix adhesion, cell motility, and invasiveness. More recently, monocyte CD44 molecules presumed to carry glycosaminoglycan chains were shown to bind the chemokine MIP-1 beta (Tanaka, Y.,D. H. Adams, S. Hubscher, H. Hirano, U. Siebenlist, and S. Shaw. 1993. Nature (Lond). 361:79-82.) raising the intriguing possibility that proteoglycan-like CD44 variants might play a role in regulating inflammatory responses. Here we have investigated the molecular identity of these proteoglycan-like CD44 variants by generating a panel of recombinant CD44 isoforms using a novel cassette cloning strategy. We show that both chondroitin and heparan sulphate modifications are associated specifically with isoforms (CD44v3-10 and CD44v3,8-10) containing the v3 alternative exon which encodes a consensus motif SGXG for GAG addition. Other isoforms (CD44v10, CD44v8-10, CD44v7-10, and CD44v6-10) are shown to lack these GAG chains but to carry extensive O-glycan modifications, most likely within the mucin-like alternative exon inserts. We also demonstrate that the majority of endogenous GAG-modified CD44 isoforms present in epithelial cells constitute v3 isoforms thus establishing that in these cells the majority of proteoglycan-like CD44 variants are generated by alternative splicing. Finally we present evidence using transfected B lymphoma cells that the GAG-modified CD44 isoforms CD44v3-10 and CD44v3,8-10, unlike CD44H, bind only weakly to hyaluronan. Together with the demonstration in the accompanying paper (Bennett, K., D. G. Jackson, J.C. Simon, E. Tanczos, R. Peach, B. Modrell, I. Stamenkovic, G. Plowman, and A. Aruffo. 1995. J. Cell Biol. 128:687-698.), that CD44 molecules containing the v3 exon bind growth factors, these results highlight a new and potentially important role for CD44 alternative splicing in the control of cell-surface proteoglycan expression.
CD44细胞表面糖蛋白在多种不同组织中表达为多种异构体，这些异构体在细胞外结构域中含有1至10个可变剪接外显子v1 - v10。差异糖基化进一步增加了变异性，除了含有硫酸软骨素和硫酸乙酰肝素的蛋白聚糖样变体之外，还产生了N - 聚糖和O - 聚糖修饰形式的CD44。这些高分子量的蛋白聚糖样变体先前在淋巴细胞、黑色素瘤和角质形成细胞中被鉴定出来，它们与细胞 - 基质粘附、细胞运动性和侵袭性有关。最近，推测携带糖胺聚糖链的单核细胞CD44分子被证明能结合趋化因子MIP - 1β（田中洋、D. H. 亚当斯、S. 胡布舍尔、H. 平野、U. 西本利斯特和S. 肖。1993年。《自然》（伦敦）。361:79 - 82。），这引发了一个有趣的可能性，即蛋白聚糖样CD44变体可能在调节炎症反应中发挥作用。在这里，我们通过使用一种新颖的盒式克隆策略生成一组重组CD44异构体，研究了这些蛋白聚糖样CD44变体的分子特性。我们表明，硫酸软骨素和硫酸乙酰肝素修饰都与含有v3可变外显子的异构体（CD44v3 - 10和CD44v3,8 - 10）特异性相关，v3可变外显子编码一个用于添加GAG的共有基序SGXG。其他异构体（CD44v10、CD44v8 - 10、CD44v7 - 10和CD44v6 - 10）被证明缺乏这些GAG链，但带有广泛的O - 聚糖修饰，最有可能在粘蛋白样可变外显子插入片段内。我们还证明，上皮细胞中存在的大多数内源性GAG修饰的CD44异构体构成v3异构体，从而确定在这些细胞中，大多数蛋白聚糖样CD44变体是通过可变剪接产生的。最后，我们使用转染的B淋巴瘤细胞提供证据表明，与CD44H不同，GAG修饰的CD44异构体CD44v3 - 10和CD44v3,8 - 10与透明质酸的结合很弱。连同随附论文（贝内特、K.、D. G. 杰克逊、J. C. 西蒙、E. 坦佐斯、R. 皮奇、B. 莫德雷尔、I. 斯塔门科维奇、G. 普洛曼和A. 阿鲁福。1995年。《细胞生物学杂志》。128:687 - 698。）中的证明，即含有v3外显子的CD44分子能结合生长因子，这些结果突出了CD44可变剪接在控制细胞表面蛋白聚糖表达方面的一个新的且潜在重要的作用。
