The Function of Nonmuscle Myosin Heavy Chains

在脊椎动物中，三个基因，Myh 9，Myh 10和Myh 14编码非肌肉肌球蛋白II重链（NMHC II）的三种不同亚型。非肌肉肌球蛋白II（NM II）的运动活性位于N-末端球状头域，而肌丝形成位于C-末端杆域。先前的工作已经表明，在小鼠中消融NM II-A导致E6.5的致死性，细胞-细胞粘附缺陷和未能产生有能力的内脏内胚层。为了理解NM II-A在发育过程中的功能，我们使用同源重组产生4种不同的小鼠系：1-我们通过将编码人NMHC II-B的cDNA“敲入”II-A基因座，从而切除II-A并将NMHC II-B置于内源性II-A启动子的控制下，用NM II-B替换NM II-A（Ab*/Ab* 小鼠）。2-我们通过敲入两种嵌合NMHC来替换内源性NMII-A，一种编码与C-末端II-B杆结构域融合的NMHCII-A的N-末端运动结构域（Aab/Aab小鼠），3-另一种编码与II-A的C-末端结构域融合的NMHCII-B的N-末端结构域（阿坝/阿坝小鼠）。4-对于对照小鼠，我们将编码NMHC II-A的cDNA插入II-A位点。用II-B（Ab*/Ab*）替换NM II-A允许内脏内胚层、原肠胚形成、器官发生的正常发育和存活至E9-10。这些小鼠表现出胚胎发育延迟，血管系统和内皮细胞有缺陷。阿坝/阿坝小鼠在与Ab*/Ab* 小鼠相同的年龄死亡，表明N-末端马达结构域在II-A和II-B之间可互换，直到E9-10，尽管马达的动力学性质存在差异。Aab/Aab小鼠存活超过E12，具有正常的脉管系统，但具有发育不全的心脏和覆盖室间隔缺损的主动脉。因此，N-末端II-A马达结构域将发育期从E9-10延长到E13，显示其在早期胚胎发育中的重要性。

In vertebrates three genes, Myh9, Myh10 and Myh14 encode three different isoforms of the nonmuscle myosin II heavy chain (NMHC II). The motor activity of nonmuscle myosin II (NM II) resides in the N-terminal globular head domain and filament formation resides in the C-terminal rod domain. Previous work has shown that ablation of NM II-A in mice results in lethality by E6.5 with defects in cell-cell adhesion and a failure to produce a competent visceral endoderm. To understand the function of NM II-A during development we used homologous recombination to generate 4 different mouse lines:1-we replaced NM II-A with NM II-B by "knocking in" cDNA encoding human NMHC II-B into the II-A locus, thereby ablating II-A and placing NMHC II-B under control of the endogenous II-A promoter (Ab*/Ab* mice). 2-we replaced endogenous NM II-A by knocking in two chimeric NMHCs, one encoding the N-terminal motor domain of NMHC II-A fused to the C-terminal II-B rod domain (Aab/Aab mice), 3-and another encoding the N-terminal domain of NMHC II-B fused to the C-terminal domain of II-A (Aba/Aba mice). 4-For control mice we inserted cDNA encoding NMHC II-A into the II-A locus. Replacing NM II-A with II-B (Ab*/Ab*) allows normal development of the visceral endoderm, gastrulation, organogenesis and survival to E9-10. These mice show a delay in embryonic turning with defects in the vasculature and endocardium. Aba/Aba mice die at the same age as Ab*/Ab* mice indicating that the N-terminal motor domains are interchangeable between II-A and II-B up to E9-10 despite differences in the kinetic properties of the motors. Aab/Aab mice survive beyond E12 with a normal vasculature but with a hypoplastic heart and an aorta overriding a ventricular septal defect. Thus, the N-terminal II-A motor domain has extended the developmental period from E9-10 to E13, displaying its importance for the early embryonic development.