The role of elastic basement membrane pre-stress in Drosophila epithelial morphogenesis

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The shaping of tissues is important for animal development to generate functional organs. Tissue morphogenesis depends on the generation of force and the mechanical properties of cells. Mechanical force depends on the actomyosin cytoskeleton. Recent work also indicates an important role for the extracellular matrix (ECM) in shaping tissues. However, at present, the role of the ECM in shaping tissues is not fully understood. Epithelia are basally supported by a thin ECM layer termed basement membrane. The basement membrane is a dense, elastic, proteinaceous material that can be under mechanical prestress due to elastic deformations. However, how such elastic deformations are generated by an epithelium and how these deformations contribute to tissue morphogenesis is unknown. The Drosophila wing disc is a powerful model system to study tissue morphogenesis. As part of its morphogenesis, the wing disc forms several stereotypic folds, including a fold centred in the prospective hinge region called the H/H fold. Our previously published work showed that the formation of the H/H fold involves the local reduction of ECM beneath the pre-fold cells. Our recent preliminary data acquired by atomic force microscopy (AFM) shows, moreover, that the basement membrane of wing discs is under mechanical prestress, and that mechanical prestress depends on both actomyosin activity and basement membrane components (Collagen IV). The proposed project has four aims: Firstly, we aim to quantify basement membrane prestress in the wing disc by AFM measurements in space and time in correlation with H/H fold formation. Secondly, we aim to understand the origin of mechanical prestress in the basement membrane. In particular, we will test experimentally how changes in i) cell proliferation, ii) hydrostatic pressure, and iii) lateral cellular tension affect mechanical prestress. Concomitantly, we will identify how these alterations modify H/H fold formation.Thirdly, we aim to provide a mechanistic understanding of prestress generation and its influence on epithelial folding by developing a physical theory and computer simulations incorporating mechanical effects of hydrostatic pressure, lateral tension and local reduction of ECM beneath pre-fold cells.Fourth, we will quantify the amount of elastic prestrain in the basement membrane of wing discs during decellularization using imaging and subsequent particle image velocimetry.We expect that this interdisciplinary approach will gain new mechanistic insights into the role of ECM in shaping tissues and, thereby, will contribute to a better understanding of diseases caused by ECM alterations as well as pave the way to the development of efficient tissue engineering protocols.