Notch activity opposes ras-induced differentiation during the second mitotic wave of the developing Drosophila eye

BACKGROUND: EGF receptor acts through Ras and the MAPK cascade to trigger differentiation and maintain survival of most of cell types in the Drosophila retina. Cell types are specified sequentially by separate episodes of EGFR activity. All the cell types differentiate in G1 phase of the cell cycle. Before differentiating, many cells pass through the cell cycle in the "Second Mitotic Wave" in response to Notch activity, but no cell fates are specified during the Second Mitotic Wave. It is not known how fate specification is limited to G1-arrested cells. RESULTS: Competence to differentiate in response to activated RasV12 was diminished during the Second Mitotic Wave accounting for the failure to recruit cell fates from cycling cells. Competence was not restored by blocking cell cycle progression, but was restored by reduced Notch activity. CONCLUSION: Competence to differentiate does not depend on cell cycle progression per se, but on the same receptor activity that also induces cell cycle entry. Dual effects of Notch on the cell cycle and on differentiation help ensure that only G1 phase cells undergo fate specification

The roles of cis-inactivation by Notch ligands and of neuralized during eye and bristle patterning in Drosophila

BACKGROUND: The receptor protein Notch and its ligand Delta are expressed throughout proneural regions yet non-neural precursor cells are defined by Notch activity and neural precursor cells by Notch inactivity. Not even Delta overexpression activates Notch in neural precursor cells. It is possible that future neural cells are protected by cis-inactivation, in which ligands block activation of Notch within the same cell. The Delta-ubiquitin ligase Neuralized has been proposed to antagonize cis-inactivation, favoring Notch activation. Cis-inactivation and role of Neuralized have not yet been studied in tissues where neural precursor cells are resistant to nearby Delta, however, such as the R8 cells of the eye or the bristle precursor cells of the epidermis. RESULTS: Overexpressed ligands could block Notch signal transduction cell-autonomously in non-neural cells of the epidermis and retina, but did not activate Notch nonautonomously in neural cells. High ligand expression levels were required for cis-inactivation, and Serrate was more effective than Delta, although Delta is the ligand normally regulating neural specification. Differences between Serrate and Delta depended on the extracellular domains of the respective proteins. Neuralized was found to act cell nonautonomously in signal-sending cells during eye development, inconsistent with the view that Neuralized antagonizes cis-inactivation in non-neural cells. CONCLUSIONS: Delta and Neuralized contribute cell nonautonomously to Notch signaling in neurogenesis, and the model that Neuralized antagonizes cis-inactivation to permit Notch activity and specification of non-neural cells is refuted. The molecular mechanism rendering Notch insensitive to paracrine activation in neural precursor cells remains uncertain

Evolution of proneural atonal expression during distinct regulatory phases in the developing Drosophila eye

AbstractBackground Receptors of the Notch family affect the determination of many cell types. In the Drosophila eye, Notch antagonises the basic helix–loop–helix (bHLH) protein atonal, which is required for R8 photoreceptor determination. Similar antagonism between Notch and proneural bHLH proteins regulates most neural cell determination, however, it is uncertain whether the mechanisms are similar in all cases. Here, we have analyzed the sensitivity of atonal expression to Notch signalling using a temperature-sensitive Notch allele, by the expression of activated Notch or of the ligand Serrate, and by monitoring expression of the atonal-dependant gene scabrous and of the Notch-dependent Enhancer of split genes.Results The atonal expression pattern evolves from general ‘prepattern’ expression, through transient ‘intermediate groups’ to R8 precursor-specific expression. Successive phases of atonal expression differ in sensitivity to Notch. Prepattern expression of atonal is not inhibited. Inhibition begins at the intermediate group stage, corresponding to the period when atonal gene function is required for its own expression. At the transition to R8 cell-specific expression, Notch is activated in all intermediate group cells except the R8 cell precursor. R8 cells remain sensitive to inhibition in columns 0 and 1, but become less sensitive thereafter; non-R8 cells do not require Notch activity to keep atonal expression inactive. Thus, Notch signaling is coupled to atonal repression for only part of the atonal expression pattern. Accordingly, the Enhancer-of-split mδ protein is expressed reciprocally to atonal at the intermediate group and early R8 stages, but is expressed in other patterns before and after.Conclusions In eye development, inhibition by Notch activity is restricted to specific phases of proneural gene expression, beginning when prepattern decays and is replaced by autoregulation. We suggest that Notch signalling inhibits atonal autoregulation, but not expression by other mechanisms, and that a transition from prepattern to autoregulation is necessary for patterning neural cell determination. Distinct neural tissues might differ in their proneural prepatterns, but use Notch in a similar mechanism

Dimensions of Loss from Mental Illness

This review explores the nature, scope and consequences of loss resulting from mental illness. Losses are described within four key themes: self and identity, work and employment opportunities, relationships, and future-oriented losses. In reflecting upon review findings, several assumptions about loss are illuminated. Findings are situated within the cornerstones of recent mental health reform, specifically a recovery-oriented approach and social inclusion. Particular attention is directed towards notions of risk and responsibility and tensions in realizing the impact of loss within an individualized recovery framework. Implications and recommendations for policy and practice are highlighted

A pathway of signals regulating effector and initiator caspases in the developing Drosophila eye

Regulated cell death and survival play important roles in neural development. Extracellular signals are presumed to regulate seven apparent caspases to determine the final structure of the nervous system. In the eye, the EGF receptor, Notch, and intact primary pigment and cone cells have been implicated in survival or death signals. An antibody raised against a peptide from human caspase 3 was used to investigate how extracellular signals controlled spatial patterning of cell death. The antibody crossreacted specifically with dying Drosophila cells and labelled the activated effector caspase Drice. It was found that the initiator caspase Dronc and the proapoptotic gene head involution defective were important for activation in vivo. Dronc may play roles in dying cells in addition to activating downstream effector caspases. Epistasis experiments ordered EGF receptor, Notch, and primary pigment and cone cells into a single pathway that affected caspase activity in pupal retina through hid and Inhibitor of Apoptosis Proteins. None of these extracellular signals appeared to act by initiating caspase activation independently of hid. Taken together, these findings indicate that in eye development spatial regulation of cell death and survival is integrated through a single intracellular pathway

Author Correction: A cell-free platform for the prenylation of natural products and application to cannabinoid production.

In the original version of this Article, the genotype of the M30 mutant presented in Fig. 3b was given incorrectly as Y288V/A232S, and the M31 mutant was given incorrectly as M1/A232S. The correct genotype of the M30 mutant is Y288A/A232S and for M31 it is Y288V/A232S. In addition, to keep consistency in genotype formatting, the genotype of the M27 mutant should be Y288V/G286S. The errors have been corrected in both the PDF and HTML versions of the Article

Welfare Risks of Repeated Application of On-Farm Killing Methods for Poultry

Council Regulation (EC) no. 1099/2009 on the protection of animals at the time of killing restricts the use of manual cervical dislocation in poultry on farms in the European Union (EU) to birds weighing up to 3 kg and 70 birds per person per day. However, few studies have examined whether repeated application of manual cervical dislocation has welfare implications and whether these are dependent on individual operator skill or susceptibility to fatigue. We investigated the effects of repeated application (100 birds at a fixed killing rate of 1 bird per 2 min) and multiple operators on two methods of killing of broilers, laying hens, and turkeys in commercial settings. We compared the efficacy and welfare impact of repeated application of cervical dislocation and a percussive killer (Cash Poultry Killer, CPK), using 12 male stockworkers on three farms (one farm per bird type). Both methods achieved over 96% kill success at the first attempt. The killing methods were equally effective for each bird type and there was no evidence of reduced performance with time and/or bird number. Both methods of killing caused a rapid loss of reflexes, indicating loss of brain function. There was more variation in reflex durations and post-mortem damage in birds killed by cervical dislocation than that found using CPK. High neck dislocation was associated with improved kill success and more rapid loss of reflexes. The CPK caused damage to multiple brain areas with little variation. Overall, the CPK was associated with faster abolition of reflexes, with fewer birds exhibiting them at all, suggestive of better welfare outcomes. However, technical difficulties with the CPK highlighted the advantages of cervical dislocation, which can be performed immediately with no equipment. At the killing rates tested, we did not find evidence to justify the current EU limit on the number of birds that one operator can kill on–farm by manual cervical dislocation

Dynamic changes in lung microRNA profiles during the development of pulmonary hypertension due to chronic hypoxia and monocrotaline

<b>Objective</b>: MicroRNAs (miRNAs) are small noncoding RNAs that have the capacity to control protein production through binding "seed" sequences within a target mRNA. Each miRNA is capable of potentially controlling hundreds of genes. The regulation of miRNAs in the lung during the development of pulmonary arterial hypertension (PAH) is unknown.<p></p> <b>Methods and Results</b>: We screened lung miRNA profiles in a longitudinal and crossover design during the development of PAH caused by chronic hypoxia or monocrotaline in rats. We identified reduced expression of Dicer, involved in miRNA processing, during the onset of PAH after hypoxia. MiR-22, miR-30, and let-7f were downregulated, whereas miR-322 and miR-451 were upregulated significantly during the development of PAH in both models. Differences were observed between monocrotaline and chronic hypoxia. For example, miR-21 and let-7a were significantly reduced only in monocrotaline-treated rats. MiRNAs that were significantly regulated were validated by quantitative polymerase chain reaction. By using in vitro studies, we demonstrated that hypoxia and growth factors implicated in PAH induced similar changes in miRNA expression. Furthermore, we confirmed miR-21 downregulation in human lung tissue and serum from patients with idiopathic PAH.<p></p> <b>Conclusion</b>: Defined miRNAs are regulated during the development of PAH in rats. Therefore, miRNAs may contribute to the pathogenesis of PAH and represent a novel opportunity for therapeutic intervention.<p></p&gt