Rosenberg R D, de Agostini A I
Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
Adv Exp Med Biol. 1992;313:307-16. doi: 10.1007/978-1-4899-2444-5_30.
Mammalian cells synthesize heparan sulfate proteoglycans (HSPG) which consist of core proteins with covalently linked glycosaminoglycans (GAGs) of 50-150 disaccharide units. The GAGs exhibit great structural diversity which arise from differing arrangements of alternate disaccharide units. It has been hypothesized that HSPG may be involved in regulating the most basic aspects of cell biologic systems such as adhesion, proliferation and differentiation. However, considerable doubt exists about the specific nature of the above interactions because of a failure to isolate GAGs of unique monosaccharide sequence with appropriate biologic activities. We have demonstrated that mouse LTA cells synthesize cell surface heparan sulfate proteoglycans with regions of defined monosaccharide sequence that specifically interact with antithrombin (HSPGact). However, it remains unclear how HSPGact can be generated by a biosynthetic pathway with no simple template for directing the ordered assembly of monosaccharide units. To examine this issue, we treated LTA cells with ethylmethane sulfonate and then identified mutants that exhibit decreased antithrombin binding to heparan sulfate chains but possess no gross defects in glycosaminoglycan biosynthesis. After screening 40,000 colonies, we isolated 7 stable mutants which synthesize 8-27% of the wild type HSPGact but produce normal amounts of other HSPG. These mutants are recessive in nature, and fall into at least two different complementation groups. The delineation of the molecular basis of these defects should greatly improve our understanding of how cells synthesize HSPG with regions of defined monosaccharide sequence.
哺乳动物细胞合成硫酸乙酰肝素蛋白聚糖（HSPG），其由核心蛋白和共价连接的50 - 150个二糖单位的糖胺聚糖（GAG）组成。GAG表现出极大的结构多样性，这源于交替二糖单位的不同排列。据推测，HSPG可能参与调节细胞生物系统的最基本方面，如黏附、增殖和分化。然而，由于未能分离出具有适当生物活性的独特单糖序列的GAG，上述相互作用的具体性质存在相当大的疑问。我们已经证明，小鼠LTA细胞合成具有特定单糖序列区域的细胞表面硫酸乙酰肝素蛋白聚糖，该区域与抗凝血酶特异性相互作用（HSPGact）。然而，尚不清楚HSPGact如何通过没有简单模板来指导单糖单位有序组装的生物合成途径产生。为了研究这个问题，我们用甲基磺酸乙酯处理LTA细胞，然后鉴定出抗凝血酶与硫酸乙酰肝素链结合减少但糖胺聚糖生物合成无明显缺陷的突变体。在筛选了40,000个菌落之后，我们分离出7个稳定的突变体，它们合成的HSPGact为野生型的8 - 27%，但其他HSPG的产量正常。这些突变体本质上是隐性的，并且至少分为两个不同的互补组。对这些缺陷分子基础的描述应该会大大提高我们对细胞如何合成具有特定单糖序列区域的HSPG的理解。
