Margaret Clarke (2011) CIL:12654, Dictyostelium discoideum, cell by organism, eukaryotic cell, amoeboid cell, Eukaryotic Protist. CIL. Dataset

A movie from the time series stack described below (CIL:812) shows an amoeba (Dictyostelium discoideum) trying very hard to ingest a yeast cell (Saccharomyces cerevisiae) that is slightly too large. (The yeast cell is a mutant with an elongated bud. See note 1.) The amoeba is expressing two fluorescent fusion proteins: GFP-PLCdelta2 (note 2), which binds to phosphatidylinositol 4,5-bisphophate and labels the plasma membrane, and mRFP-LimEdelta (note 3), which labels actin filaments. Actin filaments shape the phagocytic cup at its leading edge and ring it in dynamic bands as it extends deep in the cell. The yeast is too long, and eventually the amoeba gives up, as indicated by the disappearance of the actin filaments. The yeast is promptly expelled with remarkable speed and force, and the excess membrane is converted into ruffles. The motile amoeba contacts the yeast anew, spreads along its length until reaching the end, then tries again to engulf it, but with no better luck. The similarities between a cell spreading on a substratum and forming a phagocytic cup are evident. Methods: Living cells were imaged using a Zeiss LSM 510 laser scanning confocal microscope with standard components and a Zeiss Plan-Apochromat 63x 1.4 N.A. DIC objective. Simultaneous fluorescence images (confocal) and DIC images were recorded from a single focal plane. Scans were performed every 3.89 seconds using a scan zoom of 3.5. References to reagents: 1: S. cerevisiae DDY-1306 (Kozminski, K.G., Chen, A.J., Rodal, A.A., Drubin, D.G. Mol. Biol. Cell 11:339-54, 2000). 2: Green Fluorescent Protein fused to the pleckstrin homology domain of phospholipase C delta-1 (Sauffer, T.P., Ahn, S., and Meyer, T., Curr. Biol. 8:343-346, 1998). 3: Monomeric Red Fluorescent Protein fused to a fragment of Dictyostelium LimE (Fischer, M., Haase, I., Simmeth, E., Gerisch, G., and Muller-Taubenberger, A., FEBS Lett. 577:227-232, 2004). The video received an Honorable Mention in the 2009 Olympus Bioscapes International Digital Imaging Competition and also may be viewed here: http://www.olympusbioscapes.com/gallery/2009/.

以下时间序列栈所描述的电影展现了一种变形虫（Dictyostelium discoideum）竭尽全力吞噬一个略微过大的酵母细胞（Saccharomyces cerevisiae）的情景。（该酵母细胞为突变体，具有细长的芽。参见注释1。）该变形虫表达两种荧光融合蛋白：GFP-PLCdelta2（注释2），该蛋白与磷脂酰肌醇4,5-二磷酸结合并标记细胞膜，以及mRFP-LimEdelta（注释3），该蛋白标记肌动蛋白纤维。肌动蛋白纤维在细胞的先端形成吞噬杯，并在深入细胞的过程中以动态带状环绕。由于酵母细胞过长，最终变形虫放弃尝试，这可通过肌动蛋白纤维的消失得到证实。酵母细胞随后迅速且有力地被排出，过剩的膜转化为皱褶。具有移动能力的变形虫重新接触酵母，沿着其长度延伸直至末端，然后再次尝试吞噬，但结果依旧不利。细胞在底物上扩展并形成吞噬杯的相似性显而易见。研究方法：利用蔡司LSM 510激光扫描共聚焦显微镜以及标准组件和蔡司Plan-Apochromat 63x 1.4 N.A. DIC物镜对活细胞进行成像。从单个焦平面同时记录荧光图像（共聚焦）和DIC图像。每3.89秒进行一次扫描，扫描放大倍数为3.5。试剂参考：1：S. cerevisiae DDY-1306（Kozminski, K.G., Chen, A.J., Rodal, A.A., Drubin, D.G. Mol. Biol. Cell 11:339-54, 2000）。2：绿色荧光蛋白与磷脂酶C delta-1的pleckstrin同源结构域融合（Sauffer, T.P., Ahn, S., and Meyer, T., Curr. Biol. 8:343-346, 1998）。3：单体红色荧光蛋白与Dictyostelium LimE片段融合（Fischer, M., Haase, I., Simmeth, E., Gerisch, G., and Muller-Taubenberger, A., FEBS Lett. 577:227-232, 2004）。该视频在2009年奥林巴斯生物景观国际数字成像竞赛中获得荣誉奖，并可在以下链接查看：http://www.olympusbioscapes.com/gallery/2009/。