Lallier T, Bronner-Fraser M
Developmental Biology Center, University of California, Irvine 92717.
Development. 1991 Dec;113(4):1069-84. doi: 10.1242/dev.113.4.1069.
The mechanisms of neural crest cell interaction with laminin were explored using a quantitative cell attachment assay. With increasing substratum concentrations, an increasing percentage of neural crest cells adhere to laminin. Cell adhesion at all substratum concentrations was inhibited by the CSAT antibody, which recognizes the chick beta 1 subunit of integrin, suggesting that beta 1-integrins mediate neural crest cell interactions with laminin. The HNK-1 antibody, which recognizes a carbohydrate epitope, inhibited neural crest cell attachment to laminin at low coating concentrations (greater than 1 microgram ml-1; Low-LM), but not at high coating concentration of laminin (10 micrograms ml-1; High-LM). Attachment to Low-LM occurred in the absence of divalent cations, whereas attachment to High-LM required greater than 0.1 mM Ca2+ or Mn2+. Neural crest cell adherence to the E8 fragment of laminin, derived from its long arm, was similar to that on intact laminin at high and low coating concentrations, suggesting that this fragment contains the neural crest cell binding site(s). The HNK-1 antibody recognizes a protein of 165,000 Mr which is also found in immunoprecipitates using antibodies against the beta 1 subunit of integrin and is likely to be an integrin alpha subunit or an integrin-associated protein. Our results suggest that the HNK-1 epitope on neural crest cells is present on or associated with a novel or differentially glycosylated form of beta 1-integrin, which recognizes laminin in the apparent absence of divalent cations. We conclude that neural crest cells have at least two functionally independent means of attachment to laminin which are revealed at different substratum concentrations and/or conformations of laminin.
利用定量细胞黏附试验探究了神经嵴细胞与层粘连蛋白相互作用的机制。随着底物浓度的增加，黏附于层粘连蛋白的神经嵴细胞百分比也增加。在所有底物浓度下，细胞黏附均被CSAT抗体抑制，该抗体识别整合素的鸡β1亚基，这表明β1整合素介导神经嵴细胞与层粘连蛋白的相互作用。识别碳水化合物表位的HNK - 1抗体在低包被浓度（大于1微克/毫升；低浓度层粘连蛋白，Low - LM）时抑制神经嵴细胞与层粘连蛋白的黏附，但在高层粘连蛋白包被浓度（10微克/毫升；高浓度层粘连蛋白，High - LM）时则不抑制。在没有二价阳离子的情况下发生与低浓度层粘连蛋白的黏附，而与高浓度层粘连蛋白的黏附需要大于0.1 mM的Ca2+或Mn2+。神经嵴细胞对源自层粘连蛋白长臂的E8片段的黏附，在高、低包被浓度下与对完整层粘连蛋白的黏附相似，这表明该片段包含神经嵴细胞结合位点。HNK - 1抗体识别一种165,000 Mr的蛋白质，该蛋白质也存在于使用抗整合素β1亚基抗体的免疫沉淀物中，并且可能是一种整合素α亚基或一种整合素相关蛋白。我们的结果表明，神经嵴细胞上的HNK - 1表位存在于β1整合素的一种新的或糖基化不同的形式上或与之相关联，这种形式在明显没有二价阳离子的情况下识别层粘连蛋白。我们得出结论，神经嵴细胞至少有两种功能上独立的与层粘连蛋白黏附的方式，这在不同的底物浓度和/或层粘连蛋白构象下得以揭示。
