Interplay between mechanical tension and cytoskeletal organization in cell separation at compartment boundaries in Drosophila

在不同命运或功能的细胞之间形成直谱系限制（区室边界）是动物发育形成组织模式的重要策略。在增殖和形态发生活性组织中，室边界的平直性受到导致细胞混合的细胞插入的挑战。与大部分组织中的细胞连接相比，沿着隔室边界的细胞连接沿着的特征在于机械张力的局部增加。计算机模拟表明，这种局部增加细胞键张力的结果在一个偏置的交界处重排在细胞插入，保持直线排列的细胞连接沿着隔室边界。然而，机械张力是如何增加的细胞交界处沿着隔室边界，以及是否需要在机械张力的局部增加偏置交界重排，从而保持典型的直隔室边界形状仍然unknow.The隔室边界分离前，后细胞的果蝇蛹腹部表皮是一个有用的系统来分析隔室边界的机制。这种组织中的细胞可以进行活体成像、遗传操作和力测量。在SPP 1782的当前资助期内，我们已经证明，该隔室边界处的机械张力增加可以分为启动阶段和维持阶段。起始阶段，当前细胞和后细胞之间的第一个连接形成时，与F-肌动蛋白和非肌肉肌球蛋白II的水平升高相关，并且独立于转录调控。在维持阶段，非肌肉肌球蛋白II不再升高，但增加的机械张力取决于转录。这项建议有两个目的。首先，分析机械张力在引发阶段增加的机制。为了实现这一目标，我们将使用遗传学和实时成像相结合的方法分析肌球蛋白II马达蛋白的亚细胞运输和招募。第二，分析在细胞插入期间增加机械张力以偏置细胞重排从而保持直的隔室边界形状的需求。为了实现这一目标，我们将使用光遗传学方法来局部降低细胞连接处沿着隔室边界的机械张力，我们期望我们的结果将提供新的见解的机制，在细胞连接处的机械张力的调节，并通过这种调节影响细胞在组织发育过程中的行为。

The formation of straight lineage restrictions (compartment boundaries) between cells of different fates or functions is an important strategy of animal development to pattern tissues. The straightness of compartment boundaries in proliferating and morphogenetically active tissues is challenged by cell intercalations that lead to cell intermingling. Cell junctions along compartment boundaries are characterized by a local increase of mechanical tension as compared to cell junctions in the bulk of the tissue. Computer simulations indicate that this local increase in cell bond tension results in a bias of junctional rearrangements during cell intercalations that maintains the straight alignment of cell junctions along compartment boundaries. However, how mechanical tension is increased at cell junctions along compartment boundaries and whether the local increase in mechanical tension is required to bias junctional rearrangements and thus to maintain the characteristically straight compartment boundary shape remains unknown.The compartment boundary separating anterior and posterior cells of the Drosophila pupal abdominal epidermis is a useful system to analyze the mechanisms underlying compartment boundaries. Cells in this tissue are amenable to live imaging, genetic manipulation, and force measurements. In the current funding period of the SPP1782, we have shown that increased mechanical tension at this compartment boundary can be separated into an initiation phase and a maintenance phase. The initiation phase, when the first junctions between anterior and posterior cells are formed, correlates with elevated levels of F-actin and non-muscle Myosin II and is independent of transcriptional regulation. During the maintenance phase, non-muscle Myosin II is no longer elevated, but increased mechanical tension depends on transcription. This proposal has two aims. First, to analyze the mechanisms by which mechanical tension is increased during the initiation phase. To achieve this aim, we will analyze the subcellular trafficking and recruitment of Myosin II motor proteins using a combination of genetics and live imaging. Second, to analyze the requirement of increased mechanical tension to bias cell rearrangement during cell intercalations and thus to maintain the straight compartment boundary shape. To achieve this aim, we will use an optogenetic method to locally decrease mechanical tension at cell junctions along the compartment boundary.We expect that our results will provide novel insights into the mechanisms by which mechanical tension at cell junctions is regulated and by which this regulation influences cell behavior during tissue development.