Functional characterisation of <Emphasis Type="Italic">romeharsha</Emphasis> and <Emphasis Type="Italic">clint1</Emphasis> reaffirms the link between plasma membrane homeostasis,cell size maintenance and tissue homeostasis in developing zebrafish epidermis

In vertebrates, early developing epidermis is a bilayered epithelium consisting of an outer periderm and the underlying basal epidermis. It eventually develops into a multi-layered epithelium. The mechanisms that control the architecture and homeostasis of early developing bilayered epidermis have remained poorly understood. Recently, we have shown that the function of Myosin Vb, an actin based molecular motor, is essential in peridermal cells for maintenance of plasma membrane homeostasis. Furthermore, our analyses of the goosepimples/myosin Vb mutant unravelled a direct link between plasma membrane homeostasis, cell size maintenance and tissue homeostasis in the developing epidermis. However, it remained unclear whether this link is specific to myosin Vb mutant or this is a general principle. Here we have identified two more genetic conditions, romeharsha mutant and clint1 knockdown, in which membrane homeostasis is perturbed, as evident by increased endocytosis and accumulation of lysosomes. As a consequence, peridermal cells exhibit smaller size and increased proliferation. We further show that decreasing endocytosis in romeharsha mutant and clint1 morphants rescues or mitigates the effect on cell size, cell proliferation and morphological phenotype. Our data confirms generality of the principle by reaffirming the causal link between plasma membrane homeostasis, cell size maintenance and tissue homeostasis.     

Ras/Erk MAPK Signaling in Epidermal Homeostasis and Neoplasia

Epidermis provides the cutaneous barrier to the external environment and undergoes a continual process of proliferative self-renewal, with human epidermis undergoing complete turnover approximately 1,000 times in a lifetime. Recent work suggests that this ongoing proliferative replenishment of epidermal cells depends, in part, on continual signals for cell division and survival transmitted by the Ras/Erk MAPK pathway. Such constant cell proliferation, however, requires tight regulation to avoid the uncontrolled tissue expansion characteristic of epidermal neoplasia. Recent studies provide new insight into Ras/Erk MAPK pathway function in the control of normal skin development and homeostasis as well as how its deregulation promotes epidermal tumorigenesis.     

Temperature modulates epidermal cell size in Drosophila melanogaster

Most ectotherms show increased body size at maturity when reared under colder temperatures. In principle, temperature could produce this outcome by influencing growth, proliferation and/or death of epidermal cells. Here we investigated the effects of rearing temperature on the cell size and cell number in the wing blade, the basitarsus of the leg and the cornea of the eye of Drosophila melanogaster from two populations at opposite ends of a South American latitudinal cline. We found that, in both strains of D. melanogaster and in both sexes, a decrease in rearing temperature increases the size of the wings, legs and eyes through an effect on epidermal cell size, with no significant change in cell number. Our results indicate that temperature has a consistent effect on cell size in the Drosophila epidermis and this may also apply to other cell types. In contrast, the evolutionary effects of temperature on the different organs are not consistent. We discuss our findings in the context of growth control in Drosophila.     

Glycogen Distribution in Relation to Epidermal Cell Differentiation During Embryonic Scale Morphogenesis in the Lizard Anolis lineatopus

The differentiation of the epidermis during scale morphogenesis in the lizard Anolis lineatopus has been studied by autoradiographic and immunocytochemical techniques and by electron microscopy, in relation to mitotic activity and to the distribution of glycogen. The flat embryonic epidermis of the early embryo is transformed into symmetric epidermal papillae which progressively become asymmetric and eventually form scales with stratified epidermal and peridermal layers. Papilla asymmetrization and epidermal stratification derive from cell hypertrophy and multiplication in the “basal hypertrophic layer of the forming outer side of scales” (BLOS). Glycogen is scarce or absent during early stages of epidermis development. In the dermis no glycogen is found at any stage of scale morphogenesis. Glycogen particles 25–40 nm in size accumulate in hypertrophic basal cells and peridermal cells during scale development. Conversely cells in the forming inner side of scales do not accumulate glycogen, divide less frequently than in the outer side and do not form a β–keratinized layer. It is suggested that an osmotic effect related to glycogen deposition causes increased hydration of the BLOS, whose cells become swollen and contribute to the asymmetrization of the epidermal papillae. Glycogen decreases in suprabasal differentiating cells and disappears from the BLOS at the stage of complete keratinization of the scale, around the period of hatching. Terminal differentiation in the peridermis and suprabasal epidermal layers takes place by cell flattening and condensation of the nucleus and cytoplasm as typical for apoptotic cells.     

Epidermal Patterning Genes Impose Non-cell Autonomous Cell Size Determination and have Additional Roles in Root Meristem Size Control

The regulation of cellular growth is of vital importance for embryonic and postembryonic patterning. Growth regulation in the epidermis has importance for organ growth rates in roots and shoots, proposing epidermal cells as an interesting model for cellular growth regulation. Here we assessed whether the root epidermis is a suitable model system to address cell size determination. In Arabidopsis thaliana L., root epidermal cells are regularly spaced in neighbouring tricho- (root hair) and atrichoblast (non-hair) cells, showing already distinct cell size regulation in the root meristem. We determined cell sizes in the root meristem and at the onset of cellular elongation, revealing that not only division rates but also cellular shape is distinct in tricho- and atrichoblasts. Intriguingly, epidermal-patterning mutants, failing to define differential vacuolization in neighbouring epidermal cell files, also display non-differential growth. Using these epidermal-patterning mutants, we show that polarized growth behaviour of epidermal tricho- and atrichoblast is interdependent, suggesting non-cell autonomous signals to integrate tissue expansion. Besides the interweaved cell-type-dependent growth mechanism, we reveal an additional role for epidermal patterning genes in root meristem size and organ growth regulation. We conclude that epidermal cells represent a suitable model system to study cell size determination and interdependent tissue growth.     

Patterns of Cell Division,Cell Differentiation and Cell Elongation in Epidermis and Cortex of Arabidopsis pedicels in the Wild Type and in erecta

Plant organ shape and size are established during growth by a predictable, controlled sequence of cell proliferation, differentiation, and elongation. To understand the regulation and coordination of these processes, we studied the temporal behavior of epidermal and cortex cells in Arabidopsis pedicels and used computational modeling to analyze cell behavior in tissues. Pedicels offer multiple advantages for such a study, as their growth is determinate, mostly one dimensional, and epidermis differentiation is uniform along the proximodistal axis. Three developmental stages were distinguished during pedicel growth: a proliferative stage, a stomata differentiation stage, and a cell elongation stage. Throughout the first two stages pedicel growth is exponential, while during the final stage growth becomes linear and depends on flower fertilization. During the first stage, the average cell cycle duration in the cortex and during symmetric divisions of epidermal cells was constant and cells divided at a fairly specific size. We also examined the mutant of ERECTA, a gene with strong influence on pedicel growth. We demonstrate that during the first two stages of pedicel development ERECTA is important for the rate of cell growth along the proximodistal axis and for cell cycle duration in epidermis and cortex. The second function of ERECTA is to prolong the proliferative phase and inhibit premature cell differentiation in the epidermis. Comparison of epidermis development in the wild type and erecta suggests that differentiation is a synchronized event in which the stomata differentiation and the transition of pavement cells from proliferation to expansion are intimately connected.     

Myosin Vb Is Associated with Plasma Membrane Recycling Systems

Myosin Va is associated with discrete vesicle populations in a number of cell types, but little is known of the function of myosin Vb. Yeast two-hybrid screening of a rabbit parietal cell cDNA library with dominant active Rab11a (Rab11aS20V) identified myosin Vb as an interacting protein for Rab11a, a marker for plasma membrane recycling systems. The isolated clone, corresponding to the carboxyl terminal 60 kDa of the myosin Vb tail, interacted with all members of the Rab11 family (Rab11a, Rab11b, and Rab25). GFP-myosin Vb and endogenous myosin Vb immunoreactivity codistributed with Rab11a in HeLa and Madin-Darby canine kidney (MDCK) cells. As with Rab11a in MDCK cells, the myosin Vb immunoreactivity was dispersed with nocodazole treatment and relocated to the apical corners of cells with taxol treatment. A green fluorescent protein (GFP)-myosin Vb tail chimera overexpressed in HeLa cells retarded transferrin recycling and caused accumulation of transferrin and the transferrin receptor in pericentrosomal vesicles. Expression of the myosin Vb tail chimera in polarized MDCK cells stably expressing the polymeric IgA receptor caused accumulation of basolaterally endocytosed polymeric IgA and the polymeric IgA receptor in the pericentrosomal region. The myosin Vb tail had no effects on transferrin trafficking in polarized MDCK cells. The GFP-myosin Va tail did not colocalize with Rab11a and had no effects on recycling system vesicle distribution in either HeLa or MDCK cells. The results indicate myosin Vb is associated with the plasma membrane recycling system in nonpolarized cells and the apical recycling system in polarized cells. The dominant negative effects of the myosin Vb tail chimera indicate that this unconventional myosin is required for transit out of plasma membrane recycling systems.     

Synthesis of Very-Long-Chain Fatty Acids in the Epidermis Controls Plant Organ Growth by Restricting Cell Proliferation

Plant organ growth is controlled by inter-cell-layer communication, which thus determines the overall size of the organism. The epidermal layer interfaces with the environment and participates in both driving and restricting growth via inter-cell-layer communication. However, it remains unknown whether the epidermis can send signals to internal tissue to limit cell proliferation in determinate growth. Very-long-chain fatty acids (VLCFAs) are synthesized in the epidermis and used in the formation of cuticular wax. Here we found that VLCFA synthesis in the epidermis is essential for proper development of Arabidopsis thaliana. Wild-type plants treated with a VLCFA synthesis inhibitor and pasticcino mutants with defects in VLCFA synthesis exhibited overproliferation of cells in the vasculature or in the rib zone of shoot apices. The decrease of VLCFA content increased the expression of IPT3, a key determinant of cytokinin biosynthesis in the vasculature, and, indeed, elevated cytokinin levels. These phenotypes were suppressed in ipt3;5;7 triple mutants, and also by vasculature-specific expression of cytokinin oxidase, which degrades active forms of cytokinin. Our results imply that VLCFA synthesis in the epidermis is required to suppress cytokinin biosynthesis in the vasculature, thus fine-tuning cell division activity in internal tissue, and therefore that shoot growth is controlled by the interaction between the surface (epidermis) and the axis (vasculature) of the plant body.     

Keratinocyte differentiation is regulated by the Rho and ROCK signaling pathway

The epidermis comprises multiple layers of specialized epithelial cells called keratinocytes. As cells are lost from the outermost epidermal layers, they are replaced through terminal differentiation, in which keratinocytes of the basal layer cease proliferating, migrate upwards, and eventually reach the outermost cornified layers. Normal homeostasis of the epidermis requires that the balance between proliferation and differentiation be tightly regulated. The GTP binding protein RhoA plays a fundamental role in the regulation of the actin cytoskeleton and in the adhesion events that are critically important to normal tissue homeostasis. Two central mediators of the signals from RhoA are the ROCK serine/threonine kinases ROCK-I and ROCK-II. We have analyzed ROCK's role in the regulation of epidermal keratinocyte function by using a pharmacological inhibitor and expressing conditionally active or inactive forms of ROCK-II in primary human keratinocytes. We report that blocking ROCK function results in inhibition of keratinocyte terminal differentiation and an increase in cell proliferation. In contrast, activation of ROCK-II in keratinocytes results in cell cycle arrest and an increase in the expression of a number of genes associated with terminal differentiation. Thus, these results indicate that ROCK plays a critical role in regulating the balance between proliferation and differentiation in human keratinocytes.     

Alternative Splicing in Class V Myosins Determines Association with Rab10

Rab proteins influence vesicle trafficking pathways through the assembly of regulatory protein complexes. Previous investigations have documented that Rab11a and Rab8a can interact with the tail region of myosin Vb and regulate distinct trafficking pathways. We have now determined that a related Rab protein, Rab10, can interact with myosin Va, myosin Vb, and myosin Vc. Rab10 localized to a system of tubules and vesicles that have partially overlapping localization with Rab8a. Both Rab8a and Rab10 were mislocalized by the expression of dominant-negative myosin V tails. Interaction with Rab10 was dependent on the presence of the alternatively spliced exon D in myosin Va and myosin Vb and the homologous region in myosin Vc. Yeast two-hybrid assays and fluorescence resonance energy transfer studies confirmed that Rab10 binding to myosin V tails in vivo required the alternatively spliced exon D. In contrast to our previous work, we found that Rab11a can interact with both myosin Va and myosin Vb tails independent of their splice isoform. These results indicate that Rab GTPases regulate diverse endocytic trafficking pathways through recruitment of multiple myosin V isoforms.Eukaryotic cells are comprised of networks of highly organized membranous structures that require the efficient and timely movement of diverse intracellular proteins for proper function. Molecular motors provide the physical force needed to move these materials along microtubules and actin microfilaments. Unconventional myosin motors, such as those belonging to classes V, VI, and VII, have roles in the trafficking and recycling of membrane-bound structures in eukaryotic cells (1) and are recruited to discrete vesicle populations. Myosin VI is involved in clathrin-mediated endocytosis (2), whereas myosin VIIa participates in the proper development of stereocilia of inner ear hair cells and the transport of pigment granules in retinal pigmented epithelial cells (3, 4). Similarly, the three members of vertebrate class V myosins, myosin Va, myosin Vb, and myosin Vc, are required for the proper transport of a wide array of membrane cargoes, such as the melanosomes of pigment cells, synaptic vesicles in neurons, apical recycling endosomes in polarized epithelial cells, and bulk recycling vesicles in non-polarized cells (5).Members of the Rab family of small GTPases regulate many cellular systems, including membrane trafficking (6, 7). Certain Rab proteins associate with and regulate the function of class V myosins. Rab27a, in a complex with the adaptor protein melanophilin/Slac2-a, is required to localize myosin Va to the surface of melanin-filled pigment granules in vertebrates (8-10), whereas Rab27a and Slac2-c/MyRIP associate with both Myosin Va and myosin VIIa (3, 11). Rab11a, in a complex with its adaptor protein Rab11-FIP2, associates with myosin Vb on recycling endosomes (12-14) where the tripartite complex regulates the recycling of a variety of cargoes (15-19). In addition, Rab8a associates with both myosin Vb (20) and myosin Vc (21) as part of the non-clathrin-mediated tubular recycling system (20). Recently, Rab11a has also been shown to associate with myosin Va in the transport of AMPA receptors in dendritic spines (22), contributing to the model of myosin V regulation by multiple Rab proteins.Previous investigations have documented alternative splicing of myosin Va in a tissue-specific manner (23-28). Alternate splicing occurs in a region lying between the coiled-coil region of the neck of the motor and the globular tail region. Three exons in particular are subject to alternative splicing: exons B, D, and F (23-25). Exon F is critical for association with melanophilin/Slac2 and Rab27a (8, 9, 29, 30). Additionally, exon B is required for the interaction of myosin Va with dynein light chain 2 (DLC2) (27, 28). Currently no function for the alternatively spliced exon D has been reported. Similar to myosin Va, myosin Vb contains exons A, B, C, D, and E, whereas no exon F has yet been identified in myosin Vb (Fig. 1A). In addition, exon B in myosin Vb does not resemble the dynein light chain 2 (DLC2) binding region in myosin Va (27, 28), and therefore, it likely does not interact with DLC2. On the other hand, exon D is highly conserved among Myosin Va, myosin Vb, and myosin Vc, suggesting a common function in these molecular motors.Open in a separate windowFIGURE 1.Tissue distribution of human myosin Va and myosin Vb splice isoforms. A, schematic of the alternative exon organization in the tails of myosin Va and myosin Vb. It is known that exons B, D, and F are subject to alternative splicing in myosin Va, whereas there is only evidence that exon D is alternatively spliced in myosin Vb, which does not contain exon F. B, alignment of exon D sequences from mouse and human myosin V''s. myosin Va and myosin Vb both contain exon D (amino acids 1320-1346 of myosin Va and 1315-1340 of myosin Vb), whereas myosin Vc contains an exon D-like region (amino acids 1124-1147 of human myosin Vc) that is not known to be alternatively spliced. Alignment of the exon D regions from all three motors reveals a high degree of homology, especially in the center of the exon. Asterisks indicate amino acid identities. C, PCR-based analysis of human tissue panels reveals the alternative splicing pattern of exon D in myosin Va and myosin Vb. Primers flanking the region encoding exon D for both motors were used to amplify cDNA from human MTC™ panels (Clontech). cDNA amplified from HeLa cell RNA as well as myosin Va and myosin Vb tail constructs were used as positive controls. Variants expressing exon D (upper bands) and lacking exon D (lower bands) were visible. Per., peripheral; Pos., positive.Here we report that Rab10, a protein related to Rab8a and thought to have similar function (31-35), localizes to a system of tubules and vesicles overlapping in distribution with Rab8a in HeLa cells. Utilizing dominant-negative myosin V tail constructs, we show that Rab8a and Rab10 can interact with Myosin Va, myosin Vb, and myosin Vc in vivo. In addition, we have determined that the alternatively spliced exon D in both myosin Va and myosin Vb is required for interaction with Rab10. In contrast to our previous findings, we demonstrate that Rab11a is able to interact with both myosin Va and myosin Vb tails in an exon independent-manner. These results reveal that multiple Rab proteins potentially regulate all three class V myosin motors.     

Myosin Vb is required for trafficking of the cystic fibrosis transmembrane conductance regulator in Rab11a-specific apical recycling endosomes in polarized human airway epithelial cells

Cystic fibrosis transmembrane conductance regulator (CFTR)-mediated Cl(-) secretion across fluid-transporting epithelia is regulated, in part, by modulating the number of CFTR Cl(-) channels in the plasma membrane by adjusting CFTR endocytosis and recycling. However, the mechanisms that regulate CFTR recycling in airway epithelial cells remain unknown, at least in part, because the recycling itineraries of CFTR in these cells are incompletely understood. In a previous study, we demonstrated that CFTR undergoes trafficking in Rab11a-specific apical recycling endosomes in human airway epithelial cells. Myosin Vb is a plus-end-directed, actin-based mechanoenzyme that facilitates protein trafficking in Rab11a-specific recycling vesicles in several cell model systems. There are no published studies examining the role of myosin Vb in airway epithelial cells. Thus, the goal of this study was to determine whether myosin Vb facilitates CFTR recycling in polarized human airway epithelial cells. Endogenous CFTR formed a complex with endogenous myosin Vb and Rab11a. Silencing myosin Vb by RNA-mediated interference decreased the expression of wild-type CFTR and DeltaF508-CFTR in the apical membrane and decreased CFTR-mediated Cl(-) secretion across polarized human airway epithelial cells. A recombinant tail domain fragment of myosin Vb attenuated the plasma membrane expression of CFTR by arresting CFTR recycling. The dominant-negative effect was dependent on the ability of the myosin Vb tail fragment to interact with Rab11a. Taken together, these data indicate that myosin Vb is required for CFTR recycling in Rab11a-specific apical recycling endosomes in polarized human airway epithelial cells.     

Regulation of tomato fruit growth by epidermal cell wall enzymes

Water relations of tomato fruit and the epidermal and pericarp activities of the putative cell wall loosening and tightening enzymes Xyloglucan endotransglycosylase (XET) and peroxidase were investigated, to determine whether tomato fruit growth is principally regulated in the epidermis or pericarp. Analysis of the fruit water relations and observation of the pattern of expansion of tomato fruit slices in vitro , has shown that the pericarp exerts tissue pressure on the epidermis in tomato fruit, suggesting that the rate of growth of tomato fruit is determined by the physical properties of the epidermal cell walls. The epidermal activities of XET and peroxidase were assayed throughout fruit development. Temporal changes in these enzyme activities were found to correspond well with putative cell wall loosening and stiffening during fruit development. XET activity was found to be proportional to the relative expansion rate of the fruit until growth ceased, and a peroxidase activity weakly bound to the epidermal cell wall appeared shortly before cessation of fruit expansion. No equivalent peroxidase activity was detected in pericarp tissue of any age. It is therefore plausible that the expansion of tomato fruit is regulated by the combined action of these enzyme activities in the fruit epidermis.     

盐胁迫对不同品系杨树幼苗生长、细胞超微结构和离子稳态的影响

以欧美杨（Populus canadensis）南杨1号和南杨2号为实验材料,研究了NaCl胁迫对其幼苗生长、细胞超微形态结构和离子分配等的影响。结果发现,低盐（75mmol·L^–1NaCl）胁迫对南杨1号生长的抑制显著高于南杨2号;高盐（150mmol·L^–1NaCl）胁迫对2种品系生长的抑制则差异不显著。低盐胁迫下,南杨1号叶片细胞结构破坏程度明显高于南杨2号;南杨2号根中所有细胞,新生枝条表皮、皮层及木质部细胞,叶片上表皮、栅栏和海绵组织细胞均维持较低的Na＋含量,同时叶片栅栏和海绵组织细胞维持较高的Mg2＋含量,从而表现为向枝条和叶片的Na＋流量显著偏低。维持细胞内的离子稳态可能是南杨2号耐盐性高于南杨1号的重要原因。     

Tensional Homeostasis in Single Fibroblasts

Adherent cells generate forces through acto-myosin contraction to move, change shape, and sense the mechanical properties of their environment. They are thought to maintain defined levels of tension with their surroundings despite mechanical perturbations that could change tension, a concept known as tensional homeostasis. Misregulation of tensional homeostasis has been proposed to drive disorganization of tissues and promote progression of diseases such as cancer. However, whether tensional homeostasis operates at the single cell level is unclear. Here, we directly test the ability of single fibroblast cells to regulate tension when subjected to mechanical displacements in the absence of changes to spread area or substrate elasticity. We use a feedback-controlled atomic force microscope to measure and modulate forces and displacements of individual contracting cells as they spread on a fibronectin-patterned atomic-force microscope cantilever and coverslip. We find that the cells reach a steady-state contraction force and height that is insensitive to stiffness changes as they fill the micropatterned areas. Rather than maintaining a constant tension, the fibroblasts altered their contraction force in response to mechanical displacement in a strain-rate-dependent manner, leading to a new and stable steady-state force and height. This response is influenced by overexpression of the actin crosslinker α-actinin, and rheology measurements reveal that changes in cell elasticity are also strain- rate-dependent. Our finding of tensional buffering, rather than homeostasis, allows cells to transition between different tensional states depending on how they are displaced, permitting distinct responses to slow deformations during tissue growth and rapid deformations associated with injury.     

Motor Force Homeostasis in Skeletal Muscle Contraction

In active biological contractile processes such as skeletal muscle contraction, cellular mitosis, and neuronal growth, an interesting common observation is that multiple motors can perform coordinated and synchronous actions, whereas individual myosin motors appear to randomly attach to and detach from actin filaments. Recent experiment has demonstrated that, during skeletal muscle shortening at a wide range of velocities, individual myosin motors maintain a force of ∼6 pN during a working stroke. To understand how such force-homeostasis can be so precisely regulated in an apparently chaotic system, here we develop a molecular model within a coupled stochastic-elastic theoretical framework. The model reveals that the unique force-stretch relation of myosin motor and the stochastic behavior of actin-myosin binding cause the average number of working motors to increase in linear proportion to the filament load, so that the force on each working motor is regulated at ∼6 pN, in excellent agreement with experiment. This study suggests that it might be a general principle to use catch bonds together with a force-stretch relation similar to that of myosin motors to regulate force homeostasis in many biological processes.     

Tornado1 and tornado2 are required for the specification of radial and circumferential pattern in the Arabidopsis root

The cell layers of the Arabidopsis primary root are arranged in a simple radial pattern. The outermost layer is the lateral root cap and lies outside the epidermis that surrounds the ground tissue. The files of epidermal and lateral root cap cells converge on a ring of initials (lateral root cap/epidermis initial) from which the epidermal and lateral root cap tissues of the seedling are derived, once root growth is initiated after germination. Each initial gives rise to a clone of epidermal cells and a clone of lateral root cap cells. These initial divisions in the epidermal/lateral root cap initial are defective in tornado1 (trn1) and trn2 plants indicating a requirement for TRN1 and TRN2 for initial cell function. Furthermore, lateral root cap cells develop in the epidermal position in trn1 and trn2 roots indicating that TRN1 and TRN2 are required for the maintenance of the radial pattern of cell specification in the root. The death of these ectopic lateral root cap cells in the elongation zone (where lateral root cap cells normally die) results in the development of gaps in the epidermis. These observations indicate that TRN1 and TRN2 are required to maintain the distinction between the lateral root cap and epidermis and suggest that lateral root cap fate is the default state. It also suggests that TRN1 and TRN2 repress lateral root cap fate in cells in the epidermal location. Furthermore, the position-dependent pattern of root hair and non-root hair cell differentiation in the epidermis is defective in trn1 and trn2 mutants. Together these results indicate that TRN1 and TRN2 are required for the maintenance of both the radial pattern of tissue differentiation in the root and for the subsequent circumferential pattern within the epidermis.     

Cell adhesion molecule Echinoid associates with unconventional myosin VI/Jaguar motor to regulate cell morphology during dorsal closure in Drosophila

Echinoid (Ed) is a homophilic immunoglobulin domain-containing cell adhesion molecule (CAM) that localizes to adherens junctions (AJs) and cooperates with Drosophila melanogaster epithelial (DE)-cadherin to mediate cell adhesion. Here we show that Ed takes part in many processes of dorsal closure, a morphogenetic movement driven by coordinated cell shape changes and migration of epidermal cells to cover the underlying amnioserosa. Ed is differentially expressed, appearing in epidermis but not in amnioserosa cells. Ed functions independently from the JNK signaling pathway and is required to regulate cell morphology, and for assembly of actomyosin cable, filopodial protrusion and coordinated cell migration in dorsal-most epidermal cells. The effect of Ed on cell morphology requires the presence of the intracellular domain (Ed^intra). Interestingly, Ed forms homodimers in vivo and Ed^intra monomer directly associates with unconventional myosin VI/Jaguar (Jar) motor protein. We further show that ed genetically interacts with jar to control cell morphology. It has previously been shown that myosin VI is monomeric in vitro and that its dimeric form can associate with and travel processively along actin filaments. Thus, we propose that Ed mediates the dimerization of myosin VI/Jar in vivo which in turn regulates the reorganization and/or contraction of actin filaments to control changes in cell shape. Consistent with this, we found that ectopic ed expression in the amnioserosa induces myosin VI/Jar-dependent apical constriction of this tissue.     

Tensional Homeostasis in Single Fibroblasts

Adherent cells generate forces through acto-myosin contraction to move, change shape, and sense the mechanical properties of their environment. They are thought to maintain defined levels of tension with their surroundings despite mechanical perturbations that could change tension, a concept known as tensional homeostasis. Misregulation of tensional homeostasis has been proposed to drive disorganization of tissues and promote progression of diseases such as cancer. However, whether tensional homeostasis operates at the single cell level is unclear. Here, we directly test the ability of single fibroblast cells to regulate tension when subjected to mechanical displacements in the absence of changes to spread area or substrate elasticity. We use a feedback-controlled atomic force microscope to measure and modulate forces and displacements of individual contracting cells as they spread on a fibronectin-patterned atomic-force microscope cantilever and coverslip. We find that the cells reach a steady-state contraction force and height that is insensitive to stiffness changes as they fill the micropatterned areas. Rather than maintaining a constant tension, the fibroblasts altered their contraction force in response to mechanical displacement in a strain-rate-dependent manner, leading to a new and stable steady-state force and height. This response is influenced by overexpression of the actin crosslinker α-actinin, and rheology measurements reveal that changes in cell elasticity are also strain- rate-dependent. Our finding of tensional buffering, rather than homeostasis, allows cells to transition between different tensional states depending on how they are displaced, permitting distinct responses to slow deformations during tissue growth and rapid deformations associated with injury.