Rozario Tania, Mead Paul E, DeSimone Douglas W
Department of Cell Biology and The Morphogenesis and Regenerative Medicine Institute, University of Virginia, School of Medicine, Charlottesville, VA 22908, USA.
Department of Pathology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
Mech Dev. 2014 Aug;133:203-17. doi: 10.1016/j.mod.2014.07.004. Epub 2014 Aug 28.
The kindlin/fermitin family includes three proteins involved in regulating integrin ligand-binding activity and adhesion. Loss-of-function mutations in kindlins1 and 3 have been implicated in Kindler Syndrome and Leukocyte Adhesion Deficiency III (LAD-III) respectively, whereas kindlin2 null mice are embryonic lethal. Post translational regulation of cell-cell and cell-ECM adhesion has long been presumed to be important for morphogenesis, however, few specific examples of activation-dependent changes in adhesion molecule function in normal development have been reported. In this study, antisense morpholinos were used to reduce expression of individual kindlins in Xenopus laevis embryos in order to investigate their roles in early development. Kindlin1 knockdown resulted in developmental delays, gross malformations of the gut and eventual lethality by tadpole stages. Kindlin2 morphant embryos displayed late stage defects in vascular maintenance and angiogenic branching consistent with kindlin2 loss of function in the mouse. Antisense morpholinos were also used to deplete maternal kindlin2 protein in oocytes and eggs. Embryos lacking maternal kindlin2 arrested at early cleavage stages due to failures in cytokinesis. Kindlin3 morphant phenotypes included defects in epidermal ciliary beating and partial paralysis at tailbud stages but these embryos recovered eventually as morpholino levels decayed. These results indicate a remarkably diverse range of kindlin functions in vertebrate development.
黏着斑蛋白/fermitin家族包含三种参与调节整合素配体结合活性和黏附作用的蛋白质。黏着斑蛋白1和3的功能丧失突变分别与Kindler综合征和白细胞黏附缺陷III型(LAD-III)有关,而黏着斑蛋白2基因敲除的小鼠胚胎致死。长期以来,人们一直认为细胞间和细胞与细胞外基质黏附的翻译后调控对形态发生很重要,然而,在正常发育过程中,关于黏附分子功能依赖激活的变化的具体例子报道很少。在本研究中,使用反义吗啉代寡核苷酸来降低非洲爪蟾胚胎中单个黏着斑蛋白的表达,以研究它们在早期发育中的作用。敲低黏着斑蛋白1导致发育延迟、肠道严重畸形,并在蝌蚪阶段最终死亡。黏着斑蛋白2的吗啉代寡核苷酸处理的胚胎在血管维持和血管生成分支方面表现出晚期缺陷,这与小鼠中黏着斑蛋白2的功能丧失一致。反义吗啉代寡核苷酸也被用于耗尽卵母细胞和卵子中的母源黏着斑蛋白2蛋白。缺乏母源黏着斑蛋白2的胚胎由于胞质分裂失败而在早期卵裂阶段停滞。黏着斑蛋白3的吗啉代寡核苷酸处理的表型包括表皮纤毛跳动缺陷和尾芽阶段的部分麻痹,但随着吗啉代寡核苷酸水平的下降,这些胚胎最终恢复。这些结果表明黏着斑蛋白在脊椎动物发育中具有非常多样的功能。