Department of Dermatology, Venereology and Allergology, University Medical Center and Medical Faculty Mannheim, Heidelberg University and Center of Excellence in Dermatology, Mannheim, Germany.
Biochem Biophys Res Commun. 2013 Apr 26;434(1):22-7. doi: 10.1016/j.bbrc.2013.03.073. Epub 2013 Apr 2.
Liver endothelial differentiation-associated protein-1 (LEDA-1/PIANP) is a type-I-transmembrane protein first identified by us as a putative junctional protein in liver sinusoidal endothelial cells. Others have shown that LEDA-1/PIANP binds and activates immune inhibitory receptor PILRα in trans, a process that requires sialidation of LEDA-1/PIANP. Here we show that LEDA-1/PIANP is subject to O-glycosylation and sialidation as demonstrated in brain tissue as well as in LEDA-1 expressing cell lines by using anti-LEDA-1/PIANP C-terminal antibodies. In addition, analysis of LEDA-1/PIANP processing with His-tags inserted at different positions in the extracellular domain revealed that multiple steps of proteolytic cleavage occur during maturation of the protein. Proteolytic cleavage between aa59 and aa83 preceded sorting of the protein to the plasma membrane. Deletion of aa75-79 and inhibition with Furin inhibitor I confirmed that LEDA-1/PIANP is processed by a Furin-like proprotein convertase. In summary, these findings show that Furin-like proprotein convertase-dependent processing precedes plasma membrane localization of LEDA-1/PIANP that is a pre-requisite of functional receptor-ligand interactions in vitro and in vivo.
肝内皮分化相关蛋白-1（LEDA-1/PIANP）是一种 I 型跨膜蛋白，最初由我们鉴定为肝窦内皮细胞中一种假定的连接蛋白。其他人已经表明，LEDA-1/PIANP 通过反式结合并激活免疫抑制受体 PILRα，这一过程需要 LEDA-1/PIANP 的唾液酸化。在这里，我们表明 LEDA-1/PIANP 受到 O-糖基化和唾液酸化的影响，这在脑组织以及表达 LEDA-1 的细胞系中通过使用抗 LEDA-1/PIANP C 端抗体得到证实。此外，通过在细胞外结构域的不同位置插入 His 标签对 LEDA-1/PIANP 加工的分析表明，在蛋白质成熟过程中会发生多个步骤的蛋白水解切割。aa59 和 aa83 之间的蛋白水解切割发生在蛋白质分拣到质膜之前。缺失 aa75-79 并用 Furin 抑制剂 I 抑制证实 LEDA-1/PIANP 由类 Furin 蛋白原转化酶加工。总之，这些发现表明，类 Furin 蛋白原转化酶依赖性加工先于 LEDA-1/PIANP 质膜定位，这是体外和体内功能性受体-配体相互作用的先决条件。
