Connors S A, Trout J, Ekker M, Mullins M C
University of Pennsylvania School of Medicine, Department of Cell and Developmental Biology, 421 Curie Blvd., Philadelphia, PA 19104-6058, USA.
Development. 1999 Jun;126(14):3119-30. doi: 10.1242/dev.126.14.3119.
A highly conserved TGF-&bgr; signaling pathway is involved in the establishment of the dorsoventral axis of the vertebrate embryo. Specifically, Bone Morphogenetic Proteins (Bmps) pattern ventral tissues of the embryo while inhibitors of Bmps, such as Chordin, Noggin and Follistatin, are implicated in dorsal mesodermal and neural development. We investigated the role of Tolloid, a metalloprotease that can cleave Chordin and increase Bmp activity, in patterning the dorsoventral axis of the zebrafish embryo. Injection of tolloid mRNA into six dorsalized mutants rescued only one of these mutants, mini fin. Through chromosomal mapping, linkage and cDNA sequence analysis of several mini fin alleles, we demonstrate that mini fin encodes the tolloid gene. Characterization of the mini fin mutant phenotype reveals that Mini fin/Tolloid activity is required for patterning ventral tissues of the tail: the ventral fin, and the ventroposterior somites and vasculature. Gene expression studies show that mfn mutants exhibit reduced expression of ventrally restricted markers at the end of gastrulation, suggesting that the loss of ventral tail tissues is caused by a dorsalization occurring at the end of gastrulation. Based on the mini fin mutant phenotype and the expression of tolloid, we propose that Mini fin/Tolloid modifes the Bmp activity gradient at the end of gastrulation, when the ventralmost marginal cells of the embryo are in close proximity to the dorsal Chordin-expressing cells. At this time, unimpeded Chordin may diffuse to the most ventral marginal regions and inhibit high Bmp activity levels. In the presence of Mini fin/Tolloid, however, Chordin activity would be negatively modulated through proteolytic cleavage, thereby increasing Bmp signaling activity. This extracellular mechanism is amplified by an autoregulatory loop for bmp gene expression.
一条高度保守的转化生长因子-β（TGF-β）信号通路参与脊椎动物胚胎背腹轴的建立。具体而言，骨形态发生蛋白（Bmps）决定胚胎腹侧组织的模式，而Bmps的抑制剂，如脊索蛋白（Chordin）、头蛋白（Noggin）和卵泡抑素（Follistatin），则与背侧中胚层和神经发育有关。我们研究了Tolloid（一种能切割脊索蛋白并增加Bmp活性的金属蛋白酶）在斑马鱼胚胎背腹轴模式形成中的作用。将tolloid mRNA注射到六个背化突变体中，仅挽救了其中一个突变体——小鳍（mini fin）。通过对几个小鳍等位基因进行染色体定位、连锁分析和cDNA序列分析，我们证明小鳍编码tolloid基因。对小鳍突变体表型的特征分析表明，小鳍/Tolloid活性是尾巴腹侧组织模式形成所必需的：腹鳍、腹后侧体节和脉管系统。基因表达研究表明，小鳍突变体在原肠胚末期腹侧限制性标记物的表达降低，这表明腹侧尾部组织的缺失是由原肠胚末期发生的背化引起的。基于小鳍突变体表型和tolloid的表达，我们提出，在原肠胚末期，当胚胎最腹侧的边缘细胞与表达脊索蛋白的背侧细胞紧密相邻时，小鳍/Tolloid会改变Bmp活性梯度。此时，未受阻碍的脊索蛋白可能扩散到最腹侧的边缘区域并抑制高Bmp活性水平。然而，在存在小鳍/Tolloid的情况下，脊索蛋白的活性会通过蛋白水解切割而受到负调控，从而增加Bmp信号活性。这种细胞外机制通过bmp基因表达的自调节环得到放大。
