Lidell Martin E, Moncada Darcy M, Chadee Kris, Hansson Gunnar C
Department of Medical Biochemistry, Göteborg University, Medicinaregatan 9A, S-413 90 Gothenburg, Sweden.
Proc Natl Acad Sci U S A. 2006 Jun 13;103(24):9298-303. doi: 10.1073/pnas.0600623103. Epub 2006 Jun 5.
In order for the protozoan parasite Entamoeba histolytica (E.h.) to cause invasive intestinal and extraintestinal infection, which leads to significant morbidity and mortality, it must disrupt the protective mucus layer by a previously unknown mechanism. We hypothesized that cysteine proteases secreted from the amoeba disrupt the mucin polymeric network, thereby overcoming the protective mucus barrier. The MUC2 mucin is the major structural component of the colonic mucus gel. Heavily O-glycosylated and protease-resistant mucin domains characterize gel-forming mucins. Their N- and C-terminal cysteine-rich domains are involved in mucin polymerization, and these domains are likely to be targeted by proteases because they are less glycosylated, thereby exposing their peptide chains. By treating recombinant cysteine-rich domains of MUC2 with proteases from E.h. trophozoites, we showed that the C-terminal domain was specifically targeted at two sites by cysteine proteases, whereas the N-terminal domain was resistant to proteolysis. The major cleavage site is predicted to depolymerize the MUC2 polymers, thereby disrupting the protective mucus gel. The ability of the cysteine proteases to dissolve mucus gels was confirmed by treating mucins from a MUC2-producing cell line with amoeba proteases. These findings suggest a major role for E.h. cysteine proteases in overcoming the protective mucus barrier in the pathogenesis of invasive amoebiasis. In this report, we identify a specific cleavage mechanism used by an enteric pathogen to disrupt the polymeric nature of the mucin gel.
为了使原生动物寄生虫溶组织内阿米巴（E.h.）引发侵袭性肠道和肠外感染，进而导致显著的发病率和死亡率，它必须通过一种此前未知的机制破坏保护性黏液层。我们推测，阿米巴分泌的半胱氨酸蛋白酶会破坏黏蛋白聚合网络，从而突破保护性黏液屏障。MUC2黏蛋白是结肠黏液凝胶的主要结构成分。高度O-糖基化且具有蛋白酶抗性的黏蛋白结构域是形成凝胶的黏蛋白的特征。它们的N端和C端富含半胱氨酸的结构域参与黏蛋白聚合，并且这些结构域可能成为蛋白酶的作用靶点，因为它们的糖基化程度较低，从而使肽链暴露出来。通过用来自E.h.滋养体的蛋白酶处理MUC2的重组富含半胱氨酸结构域，我们发现C端结构域被半胱氨酸蛋白酶特异性作用于两个位点，而N端结构域对蛋白水解具有抗性。预计主要切割位点会使MUC2聚合物解聚，从而破坏保护性黏液凝胶。通过用阿米巴蛋白酶处理来自产生MUC2的细胞系的黏蛋白，证实了半胱氨酸蛋白酶溶解黏液凝胶的能力。这些发现表明E.h.半胱氨酸蛋白酶在侵袭性阿米巴病发病机制中突破保护性黏液屏障方面起主要作用。在本报告中，我们确定了一种肠道病原体用于破坏黏蛋白凝胶聚合性质的特定切割机制。
