How organ-shaping mechanical imbalances are generated is a central question of morphogenesis, with existing paradigms focusing on asymmetric force generation within cells. We show here that organs can be sculpted instead by patterning anisotropic resistance within their extracellular matrix (ECM). Using direct biophysical measurements of elongating Drosophila egg chambers, we document robust mechanical anisotropy in the ECM-based basement membrane (BM) but not in the underlying epithelium. Atomic force microscopy (AFM) on wild-type BM in vivo reveals an anterior–posterior (A–P) symmetric stiffness gradient, which fails to develop in elongation-defective mutants. Genetic manipulation shows that the BM is instructive for tissue elongation and the determinant is relative rather than absolute stiffness, creating differential resistance to isotropic tissue expansion. The stiffness gradient requires morphogen-like signaling to regulate BM incorporation, as well as planar-polarized organization to homogenize it circumferentially. Our results demonstrate how fine mechanical patterning in the ECM can guide cells to shape an organ.DOI: http://dx.doi.org/10.7554/eLife.24958.001

Summary Proper centrosome separation is a prerequisite for establishing and positioning the bipolar spindle. While studies from a diversity of cell types demonstrate that microtubules (MTs) and their associated motors are essential for driving centrosome separation [1], the role of actin in driving centrosome separation remains less clear. Studies in tissue culture cells have led to a model in which actin/myosin based cortical flow is primarily responsible for driving late centrosome separation [2], while other in vivo studies suggest actin plays a more passive role by serving as an attachment site of astral MTs to pull centrosomes apart [3–6]. Here, we demonstrate that prior to nuclear envelope breakdown (NEB) in Drosophila embryos, proper centrosome separation does not require myosin II, but requires dynamic actin rearrangements at the growing edge of the interphase cap. Both Arp2/3- and Formin-mediated actin remodeling are required for separating the centrosome pairs before NEB. The Apc2-Armadillo complex appears to link cap expansion to centrosome separation. In contrast, the mechanisms driving centrosome separation after NEB are independent of the actin cytoskeleton and can compensate for earlier separation defects. Our studies show that the dynamics of actin polymerization drive centrosome separation and this has important implications for centrosome positioning during processes such as cell migration [7, 8], cell polarity maintenance [9, 10] and asymmetric cell division [11, 12].

We present a new method to directly measure and correct the aberrations introduced when imaging through thick biological tissue. A Shack-Hartmann wavefront sensor is used to directly measure the wavefront error induced by a Drosophila embryo. The wavefront measurements are taken by seeding the embryo with fluorescent microspheres used as “artificial guide-stars.” The wavefront error is corrected in ten millisecond steps by applying the inverse to the wavefront error on a micro-electro-mechanical deformable mirror in the image path of the microscope. The results show that this new approach is capable of improving the Strehl ratio by 2 times on average and as high as 10 times when imaging through 100 μm of tissue. The results also show that the isoplanatic half-width is approximately 19 μm resulting in a corrected field of view 38 μm in diameter around the guide-star.

Spatially and temporally dependent optical aberrations induced by the inhomogeneous refractive index of live samples limit the resolution of live dynamic imaging. We introduce an adaptive optical microscope with a direct wavefront sensing method using a Shack-Hartmann wavefront sensor and fluorescent protein guide-stars for live imaging. The results of imaging Drosophila embryos demonstrate its ability to correct aberrations and achieve near diffraction limited images of medial sections of large Drosophila embryos. GFP-polo labeled centrosomes can be observed clearly after correction but cannot be observed before correction. Four dimensional time lapse images are achieved with the correction of dynamic aberrations. These studies also demonstrate that the GFP-tagged centrosome proteins, Polo and Cnn, serve as excellent biological guide-stars for adaptive optics based microscopy.

Organs are sculpted by extracellular as well as cell-intrinsic forces, but how collective cell dynamics are orchestrated in response to environmental cues is poorly understood. Here we apply advanced image analysis to reveal extracellular matrix-responsive cell behaviors that drive elongation of the Drosophila follicle, a model system in which basement membrane stiffness instructs three-dimensional tissue morphogenesis. Through in toto morphometric analyses of wild type and round egg mutants, we find that neither changes in average cell shape nor oriented cell division are required for appropriate organ shape. Instead, a major element is the reorientation of elongated cells at the follicle anterior. Polarized reorientation is regulated by mechanical cues from the basement membrane, which are transduced by the Src tyrosine kinase to alter junctional E-cadherin trafficking. This mechanosensitive cellular behavior represents a conserved mechanism that can elongate edgeless tubular epithelia in a process distinct from those that elongate bounded, planar epithelia.