Heparan sulphate proteoglycans interact with neurocan and promote neurite outgrowth from cerebellar granule cells
Heparan sulphate proteoglycans interact with neurocan and promote neurite outgrowth from cerebellar granule cells
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DOI：
10.1042/bj20040585
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发表时间：
2004-10-01
影响因子：
4.1
通讯作者：
Nakada, H
中科院分区：
文献类型：
--
作者：
Akita, K;Toda, M;Nakada, H
我们发现，神经多糖是一种主要的脑软骨素硫酸蛋白多糖，与Syndecan-3和Glypcan-1等HSPG(硫酸乙酰肝素蛋白多糖)相互作用。用肝素酶I和II处理HSPG可阻止这些HSPG与神经胶原结合，但不能用软骨素酶ABC处理神经胶原。Scatchard图分析表明，NeuroCan与这些具有不同亲和力的HSPG有两个结合位点。众所周知，随着年龄的增长，啮齿动物大脑中的神经钙素被蛋白质降解为N-末端和C-末端片段。当将从新生小鼠脑中制备的完整的神经聚糖和N-端和C-端片段或重组的N-端和C-端片段的混合物用于肝素柱时，整个分子以及N-端和C-端片段都与肝素结合。细胞黏附实验表明，N端和C端的神经钙调素片段均能与细胞表面表达的HSPG相互作用。为了研究热休克蛋白G-神经钙调素相互作用的生物学意义，将表达这些热休克蛋白原的小脑颗粒细胞培养在重组神经钙调素底物上。在涂有C-末端神经粘连蛋白片段的孔上观察到神经突起生长速度显著增加，而在N-末端涂布的孔上没有观察到明显的增加。用肝素处理神经钙调素底物或在培养液中加入肝素酶I均可抑制轴突生长促进活性。这些结果表明，Syndecan-3和Glypcan-1等HSPG是神经钙素的细胞表面受体，这些HSPG通过其C-末端结构域与神经钙素的相互作用参与促进神经突起的生长。
We found that neurocan, a major brain chondroitin sulphate proteoglycan, interacts with HSPGs (heparan sulphate proteoglycans) such as syndecan-3 and glypican-1. Binding of these HSPGs to neurocan was prevented by treatment of the HSPGs with heparitinases I and II, but not by treatment of neurocan with chondroitinase ABC. Scatchard plot analysis indicated that neurocan has two binding sites for these HSPGs with different affinities. It is known that neurocan in the rodent brain is proteolytically processed with aging into N- and C-terminal fragments. When a mixture of whole neurocan and N- and C-terminal fragments prepared from neonatal mouse brains or recombinant N- and C-terminal fragments was applied to a heparin column, the whole molecule and both the N- and C-terminal fragments bound to heparin. A centrifugation cell adhesion assay indicated that both the N- and C-terminal neurocan fragments could interact with these HSPGs expressed on the cell surface. To examine the bio-logical significance of the HSPG-neurocan interaction, cerebellar granule cells expressing these HSPGs were cultured on the recombinant neurocan substrate. A significant increase in the rate of neurite outgrowth was observed on the wells coated with the C-terminal neurocan fragment, but not with the N-terminal one. Neurite outgrowth-promoting activity was inhibited by pretreatment of neurocan substrate with heparin or the addition of heparitinase I to culture medium. These results suggest that HSPGs such as syndecan-3 and glypican-1 serve as the cell-surface receptor of neurocan, and that the interaction of these HSPGs with neurocan through its C-terminal domain is involved in the promotion of neurite outgrowth.