Itoh K, Tang T L, Neel B G, Sokol S Y
Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, MA 02215, USA.
Development. 1995 Dec;121(12):3979-88. doi: 10.1242/dev.121.12.3979.
Shaggy is a downstream component of the wingless and Notch signaling pathways which operate during Drosophila development. To address the role of glycogen synthase kinase 3 beta (GSK3 beta), a mammalian homologue of Shaggy, in vertebrate embryogenesis, it was overexpressed in Xenopus embryos. Microinjection of rat GSK3 beta mRNA into animal ventral blastomeres of 8-cell-stage embryos triggered development of ectopic cement glands with an adjacent anterior neural tissue as evidenced by in situ hybridization with Xotx2, a fore/midbrain marker, and NCAM, a pan-neural marker. In contrast, animal dorsal injection of the same dose of GSK3 beta mRNA caused eye deficiencies, whereas vegetal injections had no pronounced effects on normal development. Using several mutated forms of rat GSK3 beta, we demonstrate that the observed phenotypes are dose-dependent and tightly correlate with GSK3 beta enzymatic activity. Lineage tracing experiments showed that the effects of GSK3 beta are cell autonomous and that ectopic cement glands and eye deficiencies arose directly from cells containing GSK3 beta mRNA. Molecular marker analysis of ectodermal explants overexpressing GSK3 beta has revealed activation of Xotx2 and of cement gland marker XAG-1, but expression of NCAM and XIF-3 was not detected. Phenotypic effects of mRNA encoding a Xenopus homologue of GSK3 beta were identical to those of rat GSK3 beta mRNA. We hypothesize that GSK3 beta mediates the initial steps of neural tissue specification and modulates anteroposterior ectodermal patterning via activation of Otx2 transcription. Our observations implicate GSK3 beta in signaling pathways operating during neural tissue development and during specification of anterior ectodermal cell fates.
Shaggy是无翅和Notch信号通路的下游组分，这两条信号通路在果蝇发育过程中发挥作用。为了研究糖原合酶激酶3β（GSK3β）（Shaggy在哺乳动物中的同源物）在脊椎动物胚胎发生中的作用，在非洲爪蟾胚胎中对其进行了过表达。将大鼠GSK3β mRNA显微注射到8细胞期胚胎的动物腹侧卵裂球中，触发了异位黏腺的发育，其旁伴有前神经组织，这通过与前脑/中脑标志物Xotx2和泛神经标志物NCAM的原位杂交得以证实。相比之下，向动物背侧注射相同剂量的GSK3β mRNA会导致眼睛缺陷，而向植物极注射则对正常发育无明显影响。使用大鼠GSK3β的几种突变形式，我们证明观察到的表型是剂量依赖性的，并且与GSK3β的酶活性紧密相关。谱系追踪实验表明，GSK3β的作用是细胞自主性的，异位黏腺和眼睛缺陷直接源于含有GSK3β mRNA的细胞。对过表达GSK3β的外胚层外植体进行分子标志物分析，发现Xotx2和黏腺标志物XAG-1被激活，但未检测到NCAM和XIF-3的表达。编码非洲爪蟾GSK3β同源物的mRNA的表型效应与大鼠GSK3β mRNA的相同。我们推测，GSK3β介导神经组织特化的初始步骤，并通过激活Otx2转录来调节前后外胚层模式形成。我们的观察结果表明，GSK3β参与了神经组织发育和前外胚层细胞命运特化过程中起作用的信号通路。
