Complex Carbohydrate Research Center, University of Georgia, Athens, GA 30602, USA.
Dis Model Mech. 2012 Mar;5(2):177-90. doi: 10.1242/dmm.008219. Epub 2011 Nov 1.
The severe pediatric disorder mucolipidosis II (ML-II; also known as I-cell disease) is caused by defects in mannose 6-phosphate (Man-6-P) biosynthesis. Patients with ML-II exhibit multiple developmental defects, including skeletal, craniofacial and joint abnormalities. To date, the molecular mechanisms that underlie these clinical manifestations are poorly understood. Taking advantage of a zebrafish model of ML-II, we previously showed that the cartilage morphogenesis defects in this model are associated with altered chondrocyte differentiation and excessive deposition of type II collagen, indicating that aspects of development that rely on proper extracellular matrix homeostasis are sensitive to decreases in Man-6-P biosynthesis. To further investigate the molecular bases for the cartilage phenotypes, we analyzed the transcript abundance of several genes in chondrocyte-enriched cell populations isolated from wild-type and ML-II zebrafish embryos. Increased levels of cathepsin and matrix metalloproteinase (MMP) transcripts were noted in ML-II cell populations. This increase in transcript abundance corresponded with elevated and sustained activity of several cathepsins (K, L and S) and MMP-13 during early development. Unlike MMP-13, for which higher levels of protein were detected, the sustained activity of cathepsin K at later stages seemed to result from its abnormal processing and activation. Inhibition of cathepsin K activity by pharmacological or genetic means not only reduced the activity of this enzyme but led to a broad reduction in additional protease activity, significant correction of the cartilage morphogenesis phenotype and reduced type II collagen staining in ML-II embryos. Our findings suggest a central role for excessive cathepsin K activity in the developmental aspects of ML-II cartilage pathogenesis and highlight the utility of the zebrafish system to address the biochemical underpinnings of metabolic disease.
严重的儿科疾病黏脂贮积症 II 型（ML-II；也称为 I 细胞病）是由甘露糖 6-磷酸（Man-6-P）生物合成缺陷引起的。ML-II 患者表现出多种发育缺陷，包括骨骼、颅面和关节异常。迄今为止，这些临床表现的分子机制还知之甚少。利用 ML-II 的斑马鱼模型，我们之前表明，该模型中的软骨形态发生缺陷与软骨细胞分化改变和 II 型胶原过度沉积有关，表明依赖于适当细胞外基质稳态的发育方面对 Man-6-P 生物合成的减少很敏感。为了进一步研究软骨表型的分子基础，我们分析了从野生型和 ML-II 斑马鱼胚胎中分离的富含软骨细胞的细胞群体中的几个基因的转录丰度。在 ML-II 细胞群体中观察到组织蛋白酶和基质金属蛋白酶（MMP）转录本水平升高。这种转录本丰度的增加与几种组织蛋白酶（K、L 和 S）和 MMP-13 在早期发育过程中的活性升高和持续升高相对应。与 MMP-13 不同，检测到更高水平的蛋白质，组织蛋白酶 K 的持续活性似乎是由于其异常加工和激活所致。组织蛋白酶 K 活性的药理学或遗传抑制不仅降低了该酶的活性，而且导致其他蛋白酶活性广泛减少，ML-II 胚胎中的软骨形态发生表型显著纠正，并且 II 型胶原染色减少。我们的研究结果表明，过度的组织蛋白酶 K 活性在 ML-II 软骨发病机制的发育方面起着核心作用，并强调了斑马鱼系统在解决代谢疾病的生化基础方面的效用。
