Hnia Karim, Notarnicola Cécile, de Santa Barbara Pascal, Hugon Gérald, Rivier François, Laoudj-Chenivesse Dalila, Mornet Dominique
INSERM ERI 25 Muscle et Pathologies, Université Montpellier 1, EA 4202, CHU Arnaud de Villeneuve, Montpellier, France.
PLoS One. 2008;3(12):e3854. doi: 10.1371/journal.pone.0003854. Epub 2008 Dec 5.
The potential role and function of gastrokine-1 (GNK1) in smooth muscle cells is investigated in this work by first establishing a preparative protocol to obtain this native protein from freshly dissected chicken gizzard. Some unexpected biochemical properties of gastrokine-1 were deduced by producing specific polyclonal antibody against the purified protein. We focused on the F-actin interaction with gastrokine-1 and the potential role and function in smooth muscle contractile properties.
GNK1 is thought to provide mucosal protection in the superficial gastric epithelium. However, the actual role of gastrokine-1 with regards to its known decreased expression in gastric cancer is still unknown. Recently, trefoil factors (TFF) were reported to have important roles in gastric epithelial regeneration and cell turnover, and could be involved in GNK1 interactions. The aim of this study was to evaluate the role and function of GNK1 in smooth muscle cells.
METHODOLOGY/PRINCIPAL FINDINGS: From fresh chicken gizzard smooth muscle, an original purification procedure was used to purify a heat soluble 20 kDa protein that was sequenced and found to correspond to the gastrokine-1 protein sequence containing one BRICHOS domain and at least two or possibly three transmembrane regions. The purified protein was used to produce polyclonal antibody and highlighted the smooth muscle cell distribution and F-actin association of GNK1 through a few different methods.
CONCLUSION/SIGNIFICANCE: Altogether our data illustrate a broader distribution of gastrokine-1 in smooth muscle than only in the gastrointestinal epithelium, and the specific interaction with F-actin highlights and suggests a new role and function of GNK1 within smooth muscle cells. A potential role via TFF interaction in cell-cell adhesion and assembly of actin stress fibres is discussed.
本研究通过首先建立一种从新鲜解剖的鸡胗中获取这种天然蛋白质的制备方案，来研究胃动素-1（GNK1）在平滑肌细胞中的潜在作用和功能。通过针对纯化蛋白产生特异性多克隆抗体，推断出胃动素-1的一些意外生化特性。我们重点研究了F-肌动蛋白与胃动素-1的相互作用以及在平滑肌收缩特性中的潜在作用和功能。
GNK1被认为在胃浅表上皮中提供黏膜保护。然而，胃动素-1在胃癌中已知表达降低，其实际作用仍不清楚。最近，三叶因子（TFF）被报道在胃上皮再生和细胞更新中起重要作用，并且可能参与GNK1的相互作用。本研究的目的是评估GNK1在平滑肌细胞中的作用和功能。
方法/主要发现：从新鲜鸡胗平滑肌中，采用一种原始纯化程序纯化出一种热溶性20 kDa蛋白质，经测序发现其与包含一个BRICHOS结构域和至少两个或可能三个跨膜区域的胃动素-1蛋白质序列相对应。纯化后的蛋白质用于制备多克隆抗体，并通过几种不同方法突出显示了GNK1在平滑肌细胞中的分布以及与F-肌动蛋白的关联。
结论/意义：我们的数据总体表明，胃动素-1在平滑肌中的分布比仅在胃肠道上皮中更广泛，并且与F-肌动蛋白的特异性相互作用突出并暗示了GNK1在平滑肌细胞中的新作用和功能。讨论了通过TFF相互作用在细胞-细胞黏附和肌动蛋白应力纤维组装中的潜在作用。
