Neurofibromin controls macropinocytosis and phagocytosis in Dictyostelium.
Neurofibromin controls macropinocytosis and phagocytosis in Dictyostelium.
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作者：
Bloomfield G;Traynor D;Sander SP;Veltman DM;Pachebat JA;Kay RR
细胞利用吞噬作用和巨胞饮作用来内化大量物质，这在吞噬生物中提供生长所需的营养。野生型盘基网骨藻以细菌为食，但几十年来实验室工作一直依赖于也可以在液体培养基上生长的无菌突变体。我们使用正向遗传学来鉴定这种表型的致病基因。该基因编码RasGAP神经纤维蛋白（NF 1）。NF 1的丧失通过增加液体摄取而使无菌生长成为可能。突变体形成超大的巨胞饮体，其通过在胞吞作用位点的更大Ras和PI 3 K活性来促进。相关地，NF 1突变体可以利用吞噬作用摄取比正常颗粒大的颗粒。一个NF 1的记者被招募到新生的macropinosomes，这表明NF 1限制了他们的大小，通过局部抑制Ras信号。我们的研究结果链接NF 1与巨胞饮和吞噬作用的第一次，我们建议，NF 1在早期的吞噬细胞空间调节Ras的活动，从而限制和塑造他们的进食结构。DOI：http://dx.doi.org/10.7554/eLife.04940.001 Dictyosteomaamoeomae是以生活在土壤中的细菌为食的微生物。它们的不寻常之处在于变形虫可以以单细胞形式生存和生长，但当食物稀缺时，许多单个细胞可以聚集在一起形成一个简单的多细胞生物体。为了以细菌为食，变形虫使用一种称为吞噬作用的过程，该过程始于包围细胞的膜向外生长以完全包围细菌。这导致细菌进入称为囊泡的膜隔室中的阿米巴，在那里它们被酶分解成小分子。这些细胞也可以利用一种类似的称为巨胞饮作用的过程来吸收液体和溶解的分子。由于其短暂而相对简单的生活方式，网骨藻经常用于研究吞噬作用，细胞运动和其他在大型生物中发现的过程。例如，动物中的一些免疫细胞利用吞噬作用来捕获和破坏入侵的微生物。大多数使用网骨藻作为模型的研究都使用了带有基因突变的阿米巴，使它们能够在实验室的液体培养物中生长，而不需要以细菌为食。这些突变允许“突变”阿米巴通过巨胞饮作用吸收更多的液体和溶解的营养物质，但不知道这些突变在基因组中的位置。在这里，布卢姆菲尔德等人利用基因组测序揭示了这些突变改变了一个编码神经纤维蛋白的基因。实验表明，神经纤维蛋白的损失增加了阿米巴通过巨胞饮作用摄取的液体量，并且还使得阿米巴在吞噬作用期间摄取比正常颗粒大的颗粒。实验表明，神经纤维蛋白通过抑制另一种称为Ras的蛋白质的活性来控制吞噬作用和巨胞饮作用。神经纤维蛋白存在于动物和许多其他生物体中，因此布卢姆菲尔德等人提出，它是一种古老的蛋白质，在早期单细胞生物中进化，以控制其进食结构的大小和形状。在人类中，编码神经纤维蛋白的基因突变会导致一种严重的疾病-称为1型神经纤维瘤病-在神经系统中形成肿瘤。鉴于肿瘤细胞可以在生长过程中使用吞噬作用和巨胞饮作用来获得营养，了解这种蛋白质在Dictyosteomelamoeastrum中的作用方式可能有助于为未来开发这种人类疾病的治疗方法提供信息。DOI：http://dx.doi.org/10.7554/eLife.04940.002网站
Cells use phagocytosis and macropinocytosis to internalise bulk material, which in phagotrophic organisms supplies the nutrients necessary for growth. Wildtype Dictyostelium amoebae feed on bacteria, but for decades laboratory work has relied on axenic mutants that can also grow on liquid media. We used forward genetics to identify the causative gene underlying this phenotype. This gene encodes the RasGAP Neurofibromin (NF1). Loss of NF1 enables axenic growth by increasing fluid uptake. Mutants form outsized macropinosomes which are promoted by greater Ras and PI3K activity at sites of endocytosis. Relatedly, NF1 mutants can ingest larger-than-normal particles using phagocytosis. An NF1 reporter is recruited to nascent macropinosomes, suggesting that NF1 limits their size by locally inhibiting Ras signalling. Our results link NF1 with macropinocytosis and phagocytosis for the first time, and we propose that NF1 evolved in early phagotrophs to spatially modulate Ras activity, thereby constraining and shaping their feeding structures. DOI: http://dx.doi.org/10.7554/eLife.04940.001 Dictyostelium amoebae are microbes that feed on bacteria living in the soil. They are unusual in that the amoebae can survive and grow in a single-celled form, but when food is scarce, many individual cells can gather together to form a simple multicellular organism. To feed on bacteria, the amoebae use a process called phagocytosis, which starts with the membrane that surrounds the cell growing outwards to completely surround the bacteria. This leads to the bacteria entering the amoeba within a membrane compartment called a vesicle, where they are broken down into small molecules by enzymes. The cells can also take up fluids and dissolved molecules using a similar process called macropinocytosis. With its short and relatively simple lifestyle, Dictyostelium is often used in research to study phagocytosis, cell movement and other processes that are also found in larger organisms. For example, some immune cells in animals use phagocytosis to capture and destroy invading microbes. Most studies using Dictyostelium as a model have used amoebae with genetic mutations that allow them to be grown in liquid cultures in the laboratory without needing to feed on bacteria. The mutations allow the ‘mutant’ amoebae to take up more liquid and dissolved nutrients by macropinocytosis, but it is not known where in the genome these mutations are. Here, Bloomfield et al. used genome sequencing to reveal that these mutations alter a gene that encodes a protein called Neurofibromin. The experiments show that the loss of Neurofibromin increases the amount of fluid taken up by the amoebae through macropinocytosis, and also enables the amoebae to take up larger-than-normal particles during phagocytosis. The experiments suggest that Neurofibromin controls both phagocytosis and macropinocytosis by inhibiting the activity of another protein called Ras. Neurofibromin is found in animals and many other organisms so Bloomfield et al. propose that it is an ancient protein that evolved in early single-celled organisms to control the size and shape of their feeding structures. In humans, mutations in the gene that encodes the Neurofibromin protein can lead to the development of a severe disorder—called Neurofibromatosis type 1—in which tumours form in the nervous system. Given that tumour cells can use phagocytosis and macropinocytosis to gain nutrients as they grow, understanding how this protein works in the Dictyostelium amoebae may help to inform future efforts to develop treatments for this human disease. DOI: http://dx.doi.org/10.7554/eLife.04940.002