PDF(Pubmed)
Abstract：
Visualization of protein dynamics is a crucial step in understanding cellular processes. Chromobodies, fluorescently labelled single-domain antibodies, have emerged as versatile probes for live cell imaging of endogenous proteins. However, how these chromobodies behave in vivo and how accurately they monitor tissue changes remain poorly explored. Here, we generated an endothelial-specific β-catenin chromobody-derived probe and analyzed its expression pattern during cardiovascular development in zebrafish. Using high-resolution confocal imaging, we show that the chromobody signal correlates with the localization of β-catenin in the nucleus and at cell-cell junctions, and thereby can be used to assess endothelial maturation. Loss of Cadherin 5 strongly affects the localization of the chromobody at the cell membrane, confirming the cadherin-based adherens junction role of β-catenin. Furthermore, using a genetic model to block blood flow, we observed that cell junctions are compromised in most endothelial cells but not in the endocardium, highlighting the heterogeneous response of the endothelium to the lack of blood flow. Overall, our data further expand the use of chromobodies for in vivo applications and illustrate their potential to monitor tissue morphogenesis at high resolution.
摘要：
蛋白质动力学的可视化是理解细胞过程的关键步骤。染色体,荧光标记的单结构域抗体，已经成为用于内源性蛋白质的活细胞成像的通用探针。然而,这些显色体如何在体内表现以及它们如何准确地监测组织变化仍未得到充分探索。这里,我们产生了内皮特异性β-catenin显色体来源的探针,并分析了其在斑马鱼心血管发育过程中的表达模式.使用高分辨率共焦成像，我们表明，染色体信号与β-catenin在细胞核和细胞-细胞连接处的定位相关，从而可用于评估内皮成熟。Cadherin5的丢失强烈影响了显色体在细胞膜上的定位，证实了β-连环蛋白的基于钙粘蛋白的粘附连接作用。此外，使用遗传模型来阻断血液流动，我们观察到大多数内皮细胞的细胞连接受损，但在心内膜没有，强调内皮对血流缺乏的异质性反应。总的来说,我们的数据进一步扩大了显色体在体内应用的用途，并说明了它们在高分辨率下监测组织形态发生的潜力。
