Hubrecht Institute, KNAW and University Medical Centre Utrecht, Uppsalalaan 8, 3584CT Utrecht, The Netherlands.
FASEB J. 2012 Oct;26(10):4092-101. doi: 10.1096/fj.11-202663. Epub 2012 Jul 11.
To systematically identify novel gene functions essential for osteogenesis and skeletal mineralization, we performed a forward genetic mutagenesis screen in zebrafish and isolated a mutant that showed delayed skeletal mineralization. Analysis of the mutant phenotype in an osterix:nuclear-GFP transgenic background demonstrated that mutants contain osterix-expressing osteoblasts comparable to wild-type embryos. Positional cloning revealed a premature stop mutation in the macrophage-stimulating protein (msp) gene, predicted to result in a biologically inactive protein. Analysis of the embryonic expression pattern for the receptor for Msp, Ron, shows specific expression in the corpuscles of Stannius, a teleost-specific organ that produces stanniocalcin, a pivotal hormone in fish calcium homeostasis. Knockdown of Ron resulted in identical phenotypes as observed in msp mutants. Msp mutant embryos could be rescued by excess calcium. Consistent with a role for Msp/Ron in calcium homeostasis, calcium-regulating factors, such as pth1, pth2, stc1l, and trpv5/6 were significantly affected in msp mutant larvae. While Msp and Ron have previously been shown to play a critical role in a wide variety of biological processes, we introduce here the Msp/Ron signaling axis as a previously unappreciated player in calcium homeostasis and embryonic skeletal mineralization.
为了系统地鉴定新的基因功能，这些基因对成骨和骨骼矿化是必不可少的，我们在斑马鱼中进行了正向遗传诱变筛选，并分离出一种表现出骨骼矿化延迟的突变体。在骨形态发生蛋白（Osterix）：核-GFP 转基因背景下分析突变体表型，表明突变体含有与野生型胚胎相当的表达 Osterix 的成骨细胞。定位克隆显示巨噬细胞刺激蛋白（Msp）基因发生过早终止突变，预计会导致生物活性蛋白缺失。Msp 受体 Ron 的胚胎表达模式分析表明，其在 Stannius 小体中特异性表达，Stannius 小体是硬骨鱼特有的器官，能产生参与鱼类钙稳态的关键激素——鱼降钙素。Ron 的敲低导致与 msp 突变体中观察到的表型完全相同。Msp 突变体胚胎可通过过量钙来拯救。Msp 和 Ron 先前被证明在多种生物学过程中发挥关键作用，我们在这里提出 Msp/Ron 信号轴作为钙稳态和胚胎骨骼矿化中一个以前未被认识的调节因子。
