Autotaxin/Lysophospholipase D-mediated Lysophosphatidic Acid Signaling Is Required to Form Distinctive Large Lysosomes in the Visceral Endoderm Cells of the Mouse Yolk Sac
Autotaxin/Lysophospholipase D-mediated Lysophosphatidic Acid Signaling Is Required to Form Distinctive Large Lysosomes in the Visceral Endoderm Cells of the Mouse Yolk Sac
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DOI：
10.1074/jbc.m109.012716
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发表时间：
2009-11-27
影响因子：
4.8
通讯作者：
Masu, Masayuki
中科院分区：
文献类型：
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作者：
Koike, Seiichi;Keino-Masu, Kazuko;Masu, Masayuki
Autotaxin 是一种由 Enpp2 基因编码的溶血磷脂酶 D，是一种在细胞外空间产生溶血磷脂酸的外切酶。溶血磷脂酸作用于特定的 G 蛋白偶联受体，从而调节细胞生长、迁移和存活。先前的研究表明，Enpp2(-/-)小鼠胚胎由于卵黄囊的血管生成缺陷而在大约胚胎日(E) 9.5时死亡。然而，Enpp2(-/-)胚胎中发生哪些细胞缺陷以及哪些细胞内信号通路参与表型表现仍然未知。在这里，我们证明 Enpp2 是在卵黄囊内脏内胚层细胞中形成独特的大溶酶体所必需的。从E7.5到E9.5，Enpp2 mRNA在内脏内胚层细胞中大量表达。在Enpp2(-/-)小鼠胚胎中，内脏内胚层细胞中的溶酶体破碎。通过使用全胚胎培养系统结合细胞内信号分子的特异性药理学抑制剂，我们发现溶血磷脂酸受体和 Rho-Rho 相关卷曲螺旋蛋白激酶 (ROCK)-LIM 激酶途径是形成大溶酶体所必需的。此外，Rho、ROCK 或 LIM 激酶的显性失活形式的电穿孔也会导致野生型内脏内胚层细胞中溶酶体的尺寸减小。在Enpp2(-/-)内脏内胚层细胞中，丝切蛋白磷酸化和肌动蛋白聚合的稳态水平降低。此外，肌动蛋白抑制剂细胞松弛素 B 和 jasplakinolide 对肌动蛋白周转动力学的扰动导致溶酶体形成缺陷。这些结果表明，需要通过自分泌运动蛋白的作用在细胞外产生溶血磷脂酸来组成性激活 Rho-ROCK-LIM 激酶途径，以维持内脏内胚层细胞中溶酶体的大尺寸。
Autotaxin, a lysophospholipase D encoded by the Enpp2 gene, is an exoenzyme that produces lysophosphatidic acid in the extracellular space. Lysophosphatidic acid acts on specific G protein-coupled receptors, thereby regulating cell growth, migration, and survival. Previous studies have revealed that Enpp2(-/-) mouse embryos die at about embryonic day (E) 9.5 because of angiogenic defects in the yolk sac. However, what cellular defects occur in Enpp2(-/-) embryos and what intracellular signaling pathways are involved in the phenotype manifestation remain unknown. Here, we show that Enpp2 is required to form distinctive large lysosomes in the yolk sac visceral endoderm cells. From E7.5 to E9.5, Enpp2 mRNA is abundantly expressed in the visceral endoderm cells. In Enpp2(-/-) mouse embryos, lysosomes in the visceral endoderm cells are fragmented. By using a whole embryo culture system combined with specific pharmacological inhibitors for intracellular signaling molecules, we show that lysophosphatidic acid receptors and the Rho-Rho-associated coiled-coil containing protein kinase (ROCK)-LIM kinase pathway are required to form large lysosomes. In addition, electroporation of dominant negative forms of Rho, ROCK, or LIM kinase also leads to the size reduction of lysosomes in wild-type visceral endoderm cells. In Enpp2(-/-) visceral endoderm cells, the steady-state levels of cofilin phosphorylation and actin polymerization are reduced. In addition, perturbations of actin turnover dynamics by actin inhibitors cytochalasin B and jasplakinolide result in the defect in lysosome formation. These results suggest that constitutive activation of the Rho-ROCK-LIM kinase pathway by extracellular production of lysophosphatidic acid by the action of autotaxin is required to maintain the large size of lysosomes in visceral endoderm cells.