Novel micropatterning technique reveals dependence of cell-substrate adhesion and migration of social amoebas on parental strain, development, and fluorescent markers
Novel micropatterning technique reveals dependence of cell-substrate adhesion and migration of social amoebas on parental strain, development, and fluorescent markers
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DOI：
10.1371/journal.pone.0236171
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发表时间：
2020-07-23
期刊：
影响因子：
3.7
通讯作者：
Tarantola, Marco
中科院分区：
文献类型：
--
作者：
Karmakar, Richa;Schich, Christoph;Tarantola, Marco
社会性阿米巴Dictyosteelium discoideum是一种常用于研究趋化性的模式生物，其细胞-基质粘附是非特异性的，不涉及粘着斑复合物。因此，难以制备其中粘附的网骨藻细胞被限制在指定的微观区域的微图案化基底。在这里，我们提出了一个微图案化技术的Dictyosteoblastoma细胞，依赖于涂层基板与一个类似的1 μ m厚的层聚乙二醇（PEG）凝胶。我们发现，当平板上的基板与PEG-凝胶和玻璃的窄平行条纹，网骨藻细胞几乎排他性的坚持和迁移沿着玻璃条纹，从而提供了一个模型系统，研究一维迁移的变形细胞。令人惊讶的是，我们发现营养和发达的细胞之间，两个不同的纯实验室菌株的网骨藻，AX 2和AX 4之间的粘附到PEG-凝胶和玻璃条纹的实质性差异。甚至更令人惊讶的是，我们发现，PEG-凝胶和玻璃条纹之间的Dictyosteoblasts细胞的分布显着影响的细胞骨架的几个荧光蛋白标记物的表达。我们进行基于原子力显微镜的单细胞力光谱测量，证实了PEG-凝胶基质的粘附力在营养细胞和发育细胞、AX 2和AX 4细胞以及具有和不具有荧光标记的细胞之间可以是显著不同的。因此，父母背景的选择，发展的程度，和荧光蛋白标记物的表达都可以有一个深刻的影响细胞基板的粘附，并应考虑比较迁移的细胞时，设计微图案基板。
Cell-substrate adhesion of the social amoeba Dictyostelium discoideum, a model organism often used for the study of chemotaxis, is non-specific and does not involve focal adhesion complexes. Therefore, micropatterned substrates where adherent Dictyostelium cells are constrained to designated microscopic regions are difficult to make. Here we present a micropatterning technique for Dictyostelium cells that relies on coating the substrate with an similar to 1 mu m thick layer of polyethylene glycol (PEG) gel. We show that, when plated on a substrate with narrow parallel stripes of PEG-gel and glass, Dictyostelium cells nearly exclusive adhere to and migrate along the glass stripes, thus providing a model system to study one-dimensional migration of amoeboid cells. Surprisingly, we find substantial differences in the adhesion to PEG-gel and glass stripes between vegetative and developed cells and between two different axenic laboratory strains of Dictyostelium, AX2 and AX4. Even more surprisingly, we find that the distribution of Dictyostelium cells between PEG-gel and glass stripes is significantly affected by the expression of several fluorescent protein markers of the cytoskeleton. We carry out atomic force microscopy based single cell force spectroscopy measurements that confirm that the force of adhesion to PEG-gel substrate can be significantly different between vegetative and developed cells, AX2 and AX4 cells, and cells with and without fluorescent markers. Thus, the choice of parental background, the degree of development, and the expression of fluorescent protein markers can all have a profound effect on cell-substrate adhesion and should be considered when comparing migration of cells and when designing micropatterned substrates.