Laurent M, Martin G R, Sobel M E
Biochim Biophys Acta. 1987 Apr 29;908(3):241-50. doi: 10.1016/0167-4781(87)90104-7.
The biosynthetic products of the Engelbreth-Holm-Swarm (EHS) tumor and the cell-free translation products of EHS tumor cell RNA were characterized. Six distinct gene products (three laminin polypeptides, entactin/nidogen, and two collagen IV chains) comprising the basement membrane matrix were identified by a combination of proteolytic digestion and immunologic techniques. Analysis of the cell-free translation products using EHS tumor RNA precipitated by anti-laminin serum confirms earlier evidence that there are at least two B chains encoded by different genes. The anti-laminin serum also recognized entactin/nidogen, which was further identified by specific immunoprecipitation with anti-entactin serum. Radiolabeled laminin A chains, synthesized by the EHS tumor in organ culture, were also identified by the anti-laminin serum but were not detected among the cell-free translation products of EHS tumor RNA. Pulse-chase studies of EHS tumor in organ culture as well as in vitro translation of EHS tumor RNA suggest that the precursor forms of alpha 1(IV) and alpha 2(IV) collagen chains are nearly identical in size, with apparent molecular weights of 170,000. The mRNAs encoding these two polypeptides migrate differently on sucrose gradients. It is likely that glycosylation and hydroxylation of collagen IV account for the major differences in molecular weight of mature alpha 1(IV) and alpha 2(IV) chains in the EHS tumor matrix.
对恩格尔布雷特-霍尔姆-斯旺（EHS）肿瘤的生物合成产物以及EHS肿瘤细胞RNA的无细胞翻译产物进行了表征。通过蛋白水解消化和免疫学技术相结合，鉴定出了构成基底膜基质的六种不同基因产物（三种层粘连蛋白多肽、巢蛋白/巢素以及两条IV型胶原链）。使用抗层粘连蛋白血清沉淀的EHS肿瘤RNA对无细胞翻译产物进行分析，证实了早期的证据，即至少有两条B链由不同基因编码。抗层粘连蛋白血清还识别巢蛋白/巢素，通过用抗巢蛋白血清进行特异性免疫沉淀进一步鉴定。EHS肿瘤在器官培养中合成的放射性标记层粘连蛋白A链也被抗层粘连蛋白血清识别，但在EHS肿瘤RNA的无细胞翻译产物中未检测到。对EHS肿瘤在器官培养中的脉冲追踪研究以及EHS肿瘤RNA的体外翻译表明，α1(IV)和α2(IV)胶原链的前体形式大小几乎相同，表观分子量为170,000。编码这两种多肽的mRNA在蔗糖梯度上迁移方式不同。IV型胶原的糖基化和羟基化可能是EHS肿瘤基质中成熟α1(IV)和α2(IV)链分子量主要差异的原因。
