Chakravarti S, Stallings R L, SundarRaj N, Cornuet P K, Hassell J R
Department of Ophthalmology, School of Medicine, University of Pittsburgh, Pennsylvania 15213, USA.
Genomics. 1995 Jun 10;27(3):481-8. doi: 10.1006/geno.1995.1080.
A human corneal fibroblast cDNA library was screened with a bovine lumican cDNA probe to obtain three clones. Sequencing of the longest clone (1.75 kb) yielded an open reading frame of 1014 bp coding for a 338-amino-acid core protein. Amino acid sequencing of a tryptic peptide resulted in a 9-amino-acid match with the derived primary structure, confirming the identity of these clones. Human lumican displays all of the features of small interstitial proteoglycans: N- and C-terminal domains with highly conserved cysteines and a central domain containing nine repeats of slight variations of the leucine motif LXXLXLXXNXL. Like bovine lumican, the human core protein contains four possible N-glycosylation sites in the central domains, all or some of which are substituted with keratan sulfate side chains. At the amino acid level, it is 90% identical with bovine and 72% identical with the chicken core protein. The gene (LUM) was localized to human chromosome 12 by hybridizing a cDNA probe to a Southern blot containing a human/hamster monochromosomal mapping panel DNA. Further sublocalization to 12q21.3-q22 was performed by the fluorescence in situ hybridization technique using a lumican P1 genomic clone. By immunohistochemical staining, we show lumican's presence, not only in the corneal stroma as shown previously, but also in the dermal area of the skin, indicating a wider distribution of this proteoglycan.
用人角膜成纤维细胞cDNA文库与牛角膜蛋白聚糖cDNA探针进行筛选,获得了三个克隆。对最长的克隆(1.75 kb)进行测序,得到一个1014 bp的开放阅读框,编码一个338个氨基酸的核心蛋白。对胰蛋白酶肽段进行氨基酸测序,结果显示与推导的一级结构有9个氨基酸匹配,证实了这些克隆的身份。人角膜蛋白聚糖具有小间隙蛋白聚糖的所有特征:N端和C端结构域含有高度保守的半胱氨酸,中央结构域含有9个重复的亮氨酸基序LXXLXLXXNXL的轻微变体。与人角膜蛋白聚糖一样,人核心蛋白在中央结构域含有四个可能的N-糖基化位点,其中全部或部分被硫酸角质素侧链取代。在氨基酸水平上,它与牛的同源性为90%,与鸡的核心蛋白同源性为72%。通过将cDNA探针与包含人/仓鼠单染色体定位板DNA的Southern印迹杂交,将该基因(LUM)定位到人类12号染色体上。使用角膜蛋白聚糖P1基因组克隆,通过荧光原位杂交技术进一步将其亚定位到12q21.3-q22。通过免疫组织化学染色,我们发现角膜蛋白聚糖不仅如先前所示存在于角膜基质中,还存在于皮肤的真皮区域,表明这种蛋白聚糖分布更广泛。