Fukuda Shiro, Kita Shunbun, Obata Yoshinari, Fujishima Yuya, Nagao Hirofumi, Masuda Shigeki, Tanaka Yoshimitsu, Nishizawa Hitoshi, Funahashi Tohru, Takagi Junichi, Maeda Norikazu, Shimomura Iichiro
From the Department of Metabolic Medicine, Graduate School of Medicine, Osaka University, Osaka 565-0871, Japan.
From the Department of Metabolic Medicine, Graduate School of Medicine, Osaka University, Osaka 565-0871, Japan,
J Biol Chem. 2017 May 12;292(19):7840-7849. doi: 10.1074/jbc.M117.780734. Epub 2017 Mar 21.
Adiponectin, an adipocyte-derived circulating protein, accumulates in the heart, vascular endothelium, and skeletal muscles through an interaction with T-cadherin (T-cad), a unique glycosylphosphatidylinositol-anchored cadherin. Recent studies have suggested that this interaction is essential for adiponectin-mediated cardiovascular protection. However, the precise protein-protein interaction between adiponectin and T-cad remains poorly characterized. Using ELISA-based and surface plasmon analyses, we report here that T-cad fused with IgG Fc as a fusion tag by replacing its glycosylphosphatidylinositol-anchor specifically bound both hexameric and larger multimeric adiponectin with a dissociation constant of ∼1.0 nm and without any contribution from other cellular or serum factors. The extracellular T-cad repeats 1 and 2 were critical for the observed adiponectin binding, which is required for classical cadherin-mediated cell-to-cell adhesion. Moreover, the 130-kDa prodomain-bearing T-cad, uniquely expressed on the cell surface among members of the cadherin family and predominantly increased by adiponectin, contributed significantly to adiponectin binding. Inhibition of prodomain-processing by a prohormone convertase inhibitor increased 130-kDa T-cad levels and also enhanced adiponectin binding to endothelial cells both by more preferential cell-surface localization and by higher adiponectin-binding affinity of 130-kDa T-cad relative to 100-kDa T-cad. The preferential cell-surface localization of 130-kDa T-cad relative to 100-kDa T-cad was also observed in normal mice aorta In conclusion, our study shows that a unique key feature of the T-cad prodomain is its involvement in binding of the T-cad repeats 1 and 2 to adiponectin and also demonstrates that adiponectin positively regulates T-cad abundance.
脂联素是一种由脂肪细胞分泌的循环蛋白，通过与T-钙黏蛋白（T-cad）相互作用，在心脏、血管内皮和骨骼肌中积累。T-钙黏蛋白是一种独特的糖基磷脂酰肌醇锚定钙黏蛋白。最近的研究表明，这种相互作用对于脂联素介导的心血管保护至关重要。然而，脂联素与T-钙黏蛋白之间精确的蛋白质-蛋白质相互作用仍不清楚。我们通过基于酶联免疫吸附测定（ELISA）和表面等离子体分析报告，用IgG Fc作为融合标签取代其糖基磷脂酰肌醇锚的T-钙黏蛋白特异性结合六聚体和更大的多聚体脂联素，解离常数约为1.0纳米，且不受其他细胞或血清因子的影响。细胞外T-钙黏蛋白重复序列1和2对于观察到的脂联素结合至关重要，这是经典钙黏蛋白介导的细胞间黏附所必需的。此外，130 kDa含前结构域的T-钙黏蛋白在钙黏蛋白家族成员中独特地表达在细胞表面，且主要由脂联素增加，对脂联素结合有显著贡献。前激素转化酶抑制剂对前结构域加工的抑制增加了130 kDa T-钙黏蛋白水平，并且通过更优先的细胞表面定位以及130 kDa T-钙黏蛋白相对于100 kDa T-钙黏蛋白更高的脂联素结合亲和力，增强了脂联素与内皮细胞的结合。在正常小鼠主动脉中也观察到130 kDa T-钙黏蛋白相对于100 kDa T-钙黏蛋白更优先的细胞表面定位。总之，我们的研究表明，T-钙黏蛋白前结构域的一个独特关键特征是其参与T-钙黏蛋白重复序列1和2与脂联素的结合，并且还证明脂联素正向调节T-钙黏蛋白的丰度。
