Real-time imaging of actin filaments in the zebrafish oocyte and embryo
Real-time imaging of actin filaments in the zebrafish oocyte and embryo
复制标题
斑马鱼卵母细胞和胚胎中肌动蛋白丝的实时成像
作者：
Nukada;Y.;Horie;M.;Fukui;A.;Kotani;T.;Yamashita;M.
细胞质和皮质肌动蛋白丝的动态变化驱动着各种细胞和发育过程。尽管对活细胞中肌动蛋白丝的实时成像技术已经发展起来，但对活生物体中特定细胞中肌动蛋白丝的成像技术仍然有限，特别是对配子形成和早期胚胎发育的分析。在此，我们报道了在无环素b1启动子控制下，表达肌动蛋白丝结合蛋白Moesin C端与绿色荧光蛋白或红色荧光蛋白（GFP/RFP - MoeC）融合的转基因斑马鱼。母细胞表达GFP/RFP - MoeC，标记胚期细胞皮层肌动蛋白骨架。在成人卵巢和睾丸中检测到高水平的GFP/RFP荧光。在卵巢中，GFP/RFP‐MoeC在卵母细胞中表达，而在卵泡细胞中不表达，这使我们能够清楚地看到卵母细胞不同阶段肌动蛋白丝的组织。利用完全成熟的卵母细胞，我们揭示了卵母细胞成熟过程中皮质细胞质中聚集的肌动蛋白柱的动态变化。在生发囊泡破裂（GVBD）前的早期，柱数略有减少，在GVBD时显著减少，GVBD后柱数恢复。我们的转基因鱼可用于分析卵发生和早期胚胎发生过程中肌动蛋白丝的动态。©2015 Wiley期刊公司
Dynamic changes of cytoplasmic and cortical actin filaments drive various cellular and developmental processes. Although real‐time imaging of actin filaments in living cells has been developed, imaging of actin filaments in specific cells of living organisms remains limited, particularly for the analysis of gamete formation and early embryonic development. Here, we report the production of transgenic zebrafish expressing the C‐terminus of Moesin, an actin filament‐binding protein, fused with green fluorescent protein or red fluorescent protein (GFP/RFP‐MoeC), under the control of acyclin B1promoter. GFP/RFP‐MoeC was expressed maternally, which labels the cortical actin cytoskeleton of blastula‐stage cells. High levels of GFP/RFP fluorescence were detected in the adult ovary and testis. In the ovaries, GFP/RFP‐MoeC was expressed in oocytes but not in follicle cells, which allows us to clearly visualize the organization of actin filaments in different stages of the oocyte. Using full‐grown oocytes, we revealed the dynamic changes of actin columns assembled in the cortical cytoplasm during oocyte maturation. The number of columns slightly decreased in the early period before germinal vesicle breakdown (GVBD) and then significantly decreased at GVBD, followed by recovery after GVBD. Our transgenic fish are useful for analyzing the dynamics of actin filaments in oogenesis and early embryogenesis. © 2015 Wiley Periodicals, Inc.