Dynamic regulation of Myo-II by Rho kinase and myosin phosphatase organizes contractile Myo-II pulses in both space and time, which is necessary to maintain tissue integrity during morphogenesis.

Mason et al. show that RhoA activity is regulated in space and time by a GEF/GAP module that tunes cell behavior and is required for proper tissue folding and shape during Drosophila morphogenesis.

Summary The propagation of force in epithelial tissues requires that the contractile cytoskeletal machinery be stably connected between cells through E-cadherin-containing adherens junctions. In many epithelial tissues the cells’ contractile network is positioned at a distance from the junction. However, the mechanism(s) that connect the contractile networks to the adherens junctions, and thus mechanically connect neighboring cells, is poorly understood. Here, we identified the role for F-actin turnover in regulating the contractile cytosketal network’s attachment to adherens junctions. Perturbing F-actin turnover via gene depletion or acute drug treatments that slow F-actin turnover destabilized the attachment between the contractile actomyosin network and adherens junctions. Our work identifies a critical role for F-actin turnover in connecting actomyosin to intercellular junctions, defining a dynamic process required for the stability of force balance across intercellular contacts in tissues.

Non-muscle cell contractility is critical for tissues to adopt shape changes. Although, the non-muscle myosin II holoenzyme (myosin) is a molecular motor that powers contraction of actin cytoskeleton networks, recent studies have questioned the importance of myosin motor activity cell and tissue shape changes. Here, combining the biochemical analysis of enzymatic and motile properties for purified myosin mutants with in vivo measurements of apical constriction for the same mutants, we show that in vivo constriction rate scales with myosin motor activity. We show that so-called phosphomimetic mutants of the Drosophila regulatory light chain (RLC) do not mimic the phosphorylated RLC state in vitro. The defect in the myosin motor activity in these mutants is evident in developing Drosophila embryos where tissue recoil following laser ablation is decreased compared to wild-type tissue. Overall, our data highlights that myosin activity is required for rapid cell contraction and tissue folding in developing Drosophila embryos.DOI: http://dx.doi.org/10.7554/eLife.20828.001

In epithelial tissues, cells constantly generate and transmit forces between each other. Forces generated by the actomyosin cytoskeleton regulate tissue shape and structure and also provide signals that influence cells’ decisions to divide, die, or differentiate. Forces are transmitted across epithelia because cells are mechanically linked through junctional complexes and forces can propagate through the cell cytoplasm. Here, we review some of the molecular mechanisms responsible for force generation, with a specific focus on the actomyosin cortex and adherens junctions. We then discuss evidence for how these mechanisms promote cell shape changes and force transmission in tissues.

The formation of a hollow lumen in a formerly solid mass of cells is a key developmental process whose dysregulation leads to diseases of the kidney and other organs. Hydrostatic pressure has been proposed to drive lumen expansion, a view that is supported by experiments in the mouse blastocyst. However, lumens formed in other tissues adopt irregular shapes with cell apical faces that are bowed inward, suggesting that pressure may not be the dominant contributor to lumen shape in all cases. Here we use live-cell imaging to study the physical mechanism of lumen formation in Madin-Darby Canine Kidney cell spheroids, a canonical cell-culture model for lumenogenesis. We find that in this system, lumen shape reflects basic geometrical considerations tied to the establishment of apico-basal polarity. A physical model incorporating both cell geometry and intraluminal pressure can account for our observations as well as cases in which pressure plays a dominant role.

Cryo-electron tomography is the highest resolution tool available for structural analysis of macromolecular complexes within their native cellular environment. At present, data acquisition suffers from low throughput, in part due to the low probability of positioning a cell such that the subcellular structure of interest is on a region of the electron microscopy (EM) grid that is suitable for imaging. Here, we leverage photo-micropatterning of EM grids to optimally position endothelial cells to enable high-throughput imaging of cell-cell contacts. This method increases the distance between contacts and the thicker cell nuclei such that the regions of interest are sufficiently thin for imaging. The resulting pipeline enables the observation of a diverse array of structures at endothelial cell-cell contacts, demonstrating the utility of this technique in enhancing the rate of data acquisition for cellular cryo-electron tomography studies.

The sphingolipids (SLs) of rodent spermatogenic cells (spermatocytes, spermatids) and spermatozoa contain nonhydroxylated and 2-hydroxylated versions of very-long-chain (C26-C32) PUFAs (n-V and h-V, respectively) not present in Sertoli cells (SCs). Here, we investigated the expression of selected fatty acid elongases [elongation of very-long-chain fatty acid protein ()], with a focus on , and a fatty acid 2-hydroxylase () in rat testes with postnatal development and germ cell differentiation. Along with and, was actively transcribed in the adult testis. mRNA levels were high in immature testes and SCs, though the protein was absent. The Elovl4 protein was a germ cell product. All cells under study elongated [H]arachidonate to tetraenoic and pentaenoic C24 PUFA, but only germ cells produced C26-C32 PUFAs. Spermatocytes displayed the highest Elovl4 protein levels and enzymatic activity. mRNA was produced exclusively in germ cells, mostly round spermatids. As a protein, Fa2h was mainly concentrated in late spermatids, in the step of spermiogenesis in which they elongate and their heads change shape. The expression of Elovl4 and Fa2h thus correlate with the abundance of n-Vs and h-Vs in the SLs of rat spermatocytes and spermatids, respectively.