Balancing competing effects of tissue growth and cytoskeletal regulation during Drosophila wing disc development.

How a developing organ robustly coordinates the cellular mechanics and growth to reach a final size and shape remains poorly understood. Through iterations between experiments and model simulations that include a mechanistic description of interkinetic nuclear migration, we show that the local curvature, height, and nuclear positioning of cells in the Drosophila wing imaginal disc are defined by the concurrent patterning of actomyosin contractility, cell-ECM adhesion, ECM stiffness, and interfacial membrane tension. We show that increasing cell proliferation via different growth-promoting pathways results in two distinct phenotypes. Triggering proliferation through insulin signaling increases basal curvature, but an increase in growth through Dpp signaling and Myc causes tissue flattening. These distinct phenotypic outcomes arise from differences in how each growth pathway regulates the cellular cytoskeleton, including contractility and cell-ECM adhesion. The coupled regulation of proliferation and cytoskeletal regulators is a general strategy to meet the multiple context-dependent criteria defining tissue morphogenesis.

Vestigial-dependent induction contributes to robust patterning but is not essential for wing-fate recruitment in Drosophila.

Cell recruitment is a process by which a differentiated cell induces neighboring cells to adopt its same cell fate. In Drosophila, cells expressing the protein encoded by the wing selector gene, vestigial (vg), drive a feed-forward recruitment signal that expands the Vg pattern as a wave front. However, previous studies on Vg pattern formation do not reveal these dynamics. Here, we use live imaging to show that multiple cells at the periphery of the wing disc simultaneously activate a fluorescent reporter of the recruitment signal, suggesting that cells may be recruited without the need for their contact neighbors be recruited in advance. In support of this observation, when Vg expression is inhibited either at the dorsal-ventral boundary or away from it, the activation of the recruitment signal still occurs at a distance, suggesting that Vg expression is not absolutely required to send or propagate the recruitment signal. However, the strength and extent of the recruitment signal is clearly compromised. We conclude that a feed-forward, contact-dependent cell recruitment process is not essential for Vg patterning, but it is necessary for robustness. Overall, our findings reveal a previously unidentified role of cell recruitment as a robustness-conferring cell differentiation mechanism.

Inducing your neighbors to become like you: cell recruitment in developmental patterning and growth.

Cell differentiation, proliferation, and morphogenesis are generally driven by instructive signals that are sent and interpreted by adjacent tissues, a process known as induction. Cell recruitment is a particular case of induction in which differentiated cells produce a signal that drives adjacent cells to differentiate into the same type as the inducers. Once recruited, these new cells may become inducers to continue the recruitment process, closing a feed-forward loop that propagates the growth of a specific cell-type population. So far, little attention has been given to cell recruitment as a developmental mechanism. Here, we review the components of cell recruitment and discuss its contribution to development in three different examples: the Drosophila wing, the vertebrate inner ear, and the mammalian thyroid gland. Finally, we posit some open questions about the role of cell recruitment in organ patterning and growth.

Cystathionine ß-synthase Deficiency Impairs Vision in the Fruit Fly, Drosophila melanogaster.

PURPOSE: Deficiency in Cystathionine ß-synthase (CBS) leads to an abnormal accumulation of homocysteine and results in classical homocystinuria, a multi-systemic disorder that affects connective tissue, muscles, the central nervous system, and the eyes. However, the genetic players and mechanisms underlying vision alterations in patients with homocystinuria are little understood. MATERIALS AND METHODS: The fruit fly, Drosophila melanogaster, is a useful system to investigate the genetic basis of several human diseases, but no study to date has used Drosophila as model of homocystinuria. Here, we use Drosophila genetic tools to down-regulate CBS expression and evaluate its behavioral response to light. RESULTS: We show that CBS-deficient flies do not display the normal stereotypical behavior of attraction towards a luminous source, known as phototaxis. This behavior cannot be attributed to a motor or olfactory deficiency, but it is most likely related to a lower visual acuity. CBS-deficient flies are overall smaller, but smaller eyes do not explain their lack of phototactic response. CONCLUSIONS: The vision phenotype of CBS knock-down flies is consistent with severe myopia in homocystinuria patients. We propose to use Drosophila as a model to investigate ocular manifestations underlying homocystinuria.

A dynamic cell recruitment process drives growth of the Drosophila wing by overscaling the vestigial expression pattern.

Organs mainly attain their size by cell growth and proliferation, but sometimes also grow through recruitment of undifferentiated cells. Here we investigate the participation of cell recruitment in establishing the pattern of Vestigial (Vg), the product of the wing selector gene in Drosophila. We find that the Vg pattern overscales along the dorsal-ventral (DV) axis of the wing imaginal disc, i.e., it expands faster than the DV length of the pouch. The overscaling of the Vg pattern cannot be explained by differential proliferation, apoptosis, or oriented-cell divisions, but can be recapitulated by a mathematical model that explicitly considers cell recruitment. When impairing cell recruitment genetically, we find that the Vg pattern almost perfectly scales and adult wings are approximately 20% smaller. Conversely, impairing cell proliferation results in very small wings, suggesting that cell recruitment and cell proliferation additively contribute to organ growth in this system. Furthermore, using fluorescent reporter tools, we provide direct evidence that cell recruitment is initiated between early and mid third-instar larval development. Altogether, our work quantitatively shows when, how, and by how much cell recruitment shapes the Vg pattern and drives growth of the Drosophila wing.

Lung Beractant Increases Free Cytosolic Levels of Ca2+ in Human Lung Fibroblasts.

Beractant, a natural surfactant, induces an antifibrogenic phenotype and apoptosis in normal human lung fibroblasts (NHLF). As intracellular Ca2+ signalling has been related to programmed cell death, we aimed to assess the effect of beractant on intracellular Ca2+ concentration ([Ca2+]i) in NHLF in vitro. Cultured NHLF were loaded with Fura-2 AM (3 µM) and Ca2+ signals were recorded by microfluorimetric techniques. Beractant causes a concentration-dependent increase in [Ca2+]i with a EC50 of 0.82 µg/ml. The application of beractant, at a concentration of 500 µg/ml, which has been shown to exert an apoptotic effect in human fibroblasts, elicited different patterns of Ca2+ signals in NHLF: a) a single Ca2+ spike which could be followed by b) Ca2+ oscillations, c) a sustained Ca2+ plateau or d) a sustained plateau overlapped by Ca2+ oscillations. The amplitude and pattern of Ca2+ transients evoked by beractant were dependent on the resting [Ca2+]i. Pharmacological manipulation revealed that beractant activates a Ca2+ signal through Ca2+ release from intracellular stores mediated by phospholipase Cß (PLCß), Ca2+ release from inositol 1,4,5-trisphosphate receptors (IP3Rs) and Ca2+ influx via a store-operated pathway. Moreover, beractant-induced Ca2+ release was abolished by preventing membrane depolarization upon removal of extracellular Na+ and Ca2+. Finally, the inhibition of store-operated channels prevented beractant-induced NHLF apoptosis and downregulation of α1(I) procollagen expression. Therefore, beractant utilizes SOCE to exert its pro-apoptotic and antifibrinogenic effect on NHLF.