Hashimoto Hisashi, Rebagliati Michael, Ahmad Nadira, Muraoka Osamu, Kurokawa Tadahide, Hibi Masahiko, Suzuki Tohru
National Research Institute of Aquaculture, Nansei, Mie 516-0193, Japan.
Development. 2004 Apr;131(8):1741-53. doi: 10.1242/dev.01070.
We have isolated a novel gene, charon, that encodes a member of the Cerberus/Dan family of secreted factors. In zebrafish, Fugu and flounder, charon is expressed in regions embracing Kupffer's vesicle, which is considered to be the teleost fish equivalent to the region of the mouse definitive node that is required for left-right (L/R) patterning. Misexpression of Charon elicited phenotypes similar to those of mutant embryos defective in Nodal signaling or embryos overexpressing Antivin(Atv)/Lefty1, an inhibitor for Nodal and Activin. Charon also suppressed the dorsalizing activity of all three of the known zebrafish Nodal-related proteins (Cyclops, Squint and Southpaw), indicating that Charon can antagonize Nodal signaling. Because Southpaw functions in the L/R patterning of lateral plate mesoderm and the diencephalon, we asked whether Charon is involved in regulating L/R asymmetry. Inhibition of Charon's function by antisense morpholino oligonucleotides (MOs) led to a loss of L/R polarity, as evidenced by bilateral expression of the left side-specific genes in the lateral plate mesoderm (southpaw, cyclops, atv/lefty1, lefty2 and pitx2) and diencephalon (cyclops, atv/lefty1 and pitx2), and defects in early (heart jogging) and late (heart looping) asymmetric heart development, but did not disturb the notochord development or the atv/lefty1-mediated midline barrier function. MO-mediated inhibition of both Charon and Southpaw led to a reduction in or loss of the expression of the left side-specific genes, suggesting that Southpaw is epistatic to Charon in left-side formation. These data indicate that antagonistic interactions between Charon and Nodal (Southpaw), which take place in regions adjacent to Kupffer's vesicle, play an important role in L/R patterning in zebrafish.
我们分离出了一个新基因——卡戎基因，它编码一种属于刻耳柏洛斯/丹氏家族的分泌因子。在斑马鱼、河豚和比目鱼中，卡戎基因在包含库普弗囊泡的区域表达，库普弗囊泡被认为相当于硬骨鱼中与小鼠确定节点区域相对应的部分，而小鼠的该区域对于左右（L/R）模式形成是必需的。卡戎基因的错误表达引发的表型类似于在Nodal信号传导方面存在缺陷的突变胚胎或过度表达抗维甲酸蛋白（Atv）/Lefty1（一种Nodal和激活素的抑制剂）的胚胎的表型。卡戎基因还抑制了所有三种已知的斑马鱼Nodal相关蛋白（独眼巨人、斜视和左撇子）的背化活性，这表明卡戎基因可以拮抗Nodal信号传导。由于左撇子蛋白在侧板中胚层和间脑的左右模式形成中起作用，我们探究了卡戎基因是否参与调节左右不对称性。反义吗啉代寡核苷酸（MOs）抑制卡戎基因的功能导致左右极性丧失，侧板中胚层（左撇子、独眼巨人、Atv/Lefty1、Lefty2和pitx2）和间脑（独眼巨人、Atv/Lefty1和pitx2）中左侧特异性基因的双侧表达以及早期（心脏摆动）和晚期（心脏环化）不对称心脏发育缺陷证明了这一点，但并未干扰脊索发育或Atv/Lefty1介导的中线屏障功能。MO介导的对卡戎基因和左撇子蛋白的双重抑制导致左侧特异性基因表达减少或丧失，这表明在左侧形成过程中左撇子蛋白对卡戎基因具有上位性。这些数据表明，在与库普弗囊泡相邻的区域发生的卡戎基因与Nodal（左撇子蛋白）之间的拮抗相互作用在斑马鱼的左右模式形成中起重要作用。
