Durkin M E, Carlin B E, Vergnes J, Bartos B, Merlie J, Chung A E
Proc Natl Acad Sci U S A. 1987 Mar;84(6):1570-4. doi: 10.1073/pnas.84.6.1570.
Entactin is a widely distributed basement membrane sulfated glycoprotein of approximately equal to 150 kDa. The entactin gene is expressed early in mouse embryogenesis. Two cDNA clones complementary to rat entactin mRNA were isolated by antibody screening of an oligo(dT)-primed cDNA library constructed in the lambda gt11 expression vector. One of the clones, lambda 1E, was subcloned into plasmid pBR322 and further characterized. The clone contained sequences complementary to an mRNA species 6 kilobases in length. This mRNA was translated in rabbit reticulocyte lysates to yield a polypeptide of 143 kDa that was precipitated with anti-entactin antiserum. The cDNA insert, 1328 base pairs long, was sequenced and found to contain an open reading frame of 729 base pairs that coded for 243 amino acids at the carboxyl terminus of entactin. Analysis of the peptide revealed no extended alpha-helical or beta-sheet secondary structures. Radiolabeled probes prepared by nicktranslation of p lambda 1E were used to monitor the steady-state levels of entactin mRNA in F9 embryonal carcinoma cells that were induced to differentiate by exposure to retinoic acid and dibutyryl cyclic AMP. The increase in steady-state levels of entactin mRNA lagged behind the increase in mRNA for the B2 chain of laminin, suggesting that laminin and entactin are independently rather than coordinately regulated.
巢蛋白是一种广泛分布的基底膜硫酸化糖蛋白，分子量约为150 kDa。巢蛋白基因在小鼠胚胎发育早期表达。通过对在λgt11表达载体中构建的oligo(dT)引物cDNA文库进行抗体筛选，分离出两个与大鼠巢蛋白mRNA互补的cDNA克隆。其中一个克隆λ1E被亚克隆到质粒pBR322中并进一步鉴定。该克隆包含与一个长度为6千碱基的mRNA物种互补的序列。这种mRNA在兔网织红细胞裂解物中进行翻译，产生一个143 kDa的多肽，该多肽可被抗巢蛋白抗血清沉淀。对长度为1328个碱基对的cDNA插入片段进行测序，发现其包含一个729个碱基对的开放阅读框，该阅读框编码巢蛋白羧基末端的243个氨基酸。对该肽段的分析未发现延伸的α-螺旋或β-折叠二级结构。通过对pλ1E进行缺口平移制备的放射性标记探针，用于监测在维甲酸和二丁酰环磷腺苷诱导下分化的F9胚胎癌细胞中巢蛋白mRNA的稳态水平。巢蛋白mRNA稳态水平的增加滞后于层粘连蛋白B2链mRNA的增加，这表明层粘连蛋白和巢蛋白是独立而非协同调节的。
