A Dual Function for Prickle in Regulating Frizzled Stability during Feedback-Dependent Amplification of Planar Polarity

The core planar polarity pathway coordinates epithelial cell polarity during animal development, and loss of its activity gives rise to a range of defects, from aberrant morphogenetic cell movements to failure to correctly orient structures, such as hairs and cilia. The core pathway functions via a mechanism involving segregation of its protein components to opposite cells ends, where they form asymmetric intracellular complexes that couple cell-cell polarity. This segregation is a self-organizing process driven by feedback interactions between the core proteins themselves. Despite intense efforts, the molecular pathways underlying feedback have proven difficult to elucidate using conventional genetic approaches. Here we investigate core protein function during planar polarization of the Drosophila wing by combining quantitative measurements of protein dynamics with loss-of-function genetics, mosaic analysis, and temporal control of gene expression. Focusing on the key core protein Frizzled, we show that its stable junctional localization is promoted by the core proteins Strabismus, Dishevelled, Prickle, and Diego. In particular, we show that the stabilizing function of Prickle on Frizzled requires Prickle activity in neighboring cells. Conversely, Prickle in the same cell has a destabilizing effect on Frizzled. This destabilizing activity is dependent on the presence of Dishevelled and blocked in the absence of Dynamin and Rab5 activity, suggesting an endocytic mechanism. Overall, our approach reveals for the first time essential in vivo stabilizing and destabilizing interactions of the core proteins required for self-organization of planar polarity

Uptake of the Necrotic Serpin in Drosophila melanogaster via the Lipophorin Receptor-1

The humoral response to fungal and Gram-positive infections is regulated by the serpin-family inhibitor, Necrotic. Following immune-challenge, a proteolytic cascade is activated which signals through the Toll receptor. Toll activation results in a range of antibiotic peptides being synthesised in the fat-body and exported to the haemolymph. As with mammalian serpins, Necrotic turnover in Drosophila is rapid. This serpin is synthesised in the fat-body, but its site of degradation has been unclear. By “freezing” endocytosis with a temperature sensitive Dynamin mutation, we demonstrate that Necrotic is removed from the haemolymph in two groups of giant cells: the garland and pericardial athrocytes. Necrotic uptake responds rapidly to infection, being visibly increased after 30 mins and peaking at 6–8 hours. Co-localisation of anti-Nec with anti-AP50, Rab5, and Rab7 antibodies establishes that the serpin is processed through multi-vesicular bodies and delivered to the lysosome, where it co-localises with the ubiquitin-binding protein, HRS. Nec does not co-localise with Rab11, indicating that the serpin is not re-exported from athrocytes. Instead, mutations which block late endosome/lysosome fusion (dor, hk, and car) cause accumulation of Necrotic-positive endosomes, even in the absence of infection. Knockdown of the 6 Drosophila orthologues of the mammalian LDL receptor family with dsRNA identifies LpR1 as an enhancer of the immune response. Uptake of Necrotic from the haemolymph is blocked by a chromosomal deletion of LpR1. In conclusion, we identify the cells and the receptor molecule responsible for the uptake and degradation of the Necrotic serpin in Drosophila melanogaster. The scavenging of serpin/proteinase complexes may be a critical step in the regulation of proteolytic cascades

Implementation of A Condor Pool At The University Of Huddersfield That Conforms To A Green IT Policy

The University of Huddersfield has a large number of computers laboratories on campus that are used to capacity during timetabled session but out of these sessions the machine can be unused for long periods of time. Whilst these machines have power management software installed to reduce the universities electricity bill. The idle machines could be used to perform complex calculations and simulations to benefit the research community through cycles stealing techniques and High Throughput Computing (HTC) middlewares. In order to provide a suitable HTC service an investigation was undertaken into what middlewares are available and how they compared against each other. This study also looked at what is green IT and how the chosen HTC middleware has been adapted to conform. The investigation also involved looking into publication from other universities to see how it is used. A survey was conducted into how useful a Condor HTC grid could fit in with other universities High Performance Computing (HPC) clusters and how beneficial Condor is. The survey also looked at how Condor is funded and how it is administered. Overall the results show that Condor is an extremely useful and low cost HTC solution. Condor has been deployed within Canalside East within the University of Huddersfield as a test bed with plans to expand the Condor pool across campus. To help Condor fit within the green IT policy the compute nodes were configured to allow the machines to go into a low power state when required. To be able to prevent the possibility of having a large job queue with very few nodes online a number of scripts were created that would collect the information required to remotely wake machines up using Wake on LAN (WoL). The scripts will wake machine when a number of jobs are idle and there are machine available that are offline. In order to make Condor able to run programs that have been developed for Windows and Linux a dual Condor client system has be implemented. This has been achieved by using the standard Windows client and a virtualized Linux client with Condor on called Pools of Virtual Boxes (PoVB). These clients run as a Windows service that can be remotely switched on and off when required remotely within the same script that can wake machines when required

Implementing a Condor pool using a Green-IT policy

High Throughput Computing (HTC) systems are designed to utilise available resources on a network of idle machines in an institution or organization by cycle stealing. It provides an additional ‘free’ resource from the existing computing and networking infrastructure for modelling and simulation requiring a large number of small jobs, such as applications from biology, chemistry, physics, and digital signal processing. At the University of Huddersfield, there are thousands of idle laboratory machines that could be used to run serial/parallel jobs by cycle stealing. Our HTC system, implemented in Condor [1], is part of the Queensgate Campus Grid (QGG) [2] that consists of a number of dedicated departmental and university computer clusters. Condor is an excellent HTC tool that excels in cycle stealing and job scheduling on idle machines. However, only idle powered machines can be used from a networked pool. Many organizations deploy power saving mechanisms to try to reduce energy consumption in their systems, and power down idle resources, using rigid and inflexible power management policies. The University of Huddersfield Computing Services use the Energy Star EZ GPO power saving tool that runs as a Windows service and detects how long the computer has been idle. Then it allows the computer to first turn off the screen and then go into hibernation. Our research and development work is focused on implementing a HTC system using Condor to work within a “green IT” policy of a higher education institutions that conform to green IT challenges for a multi-platform, multi-discipline user/ resource base. This system will allow Condor to turn on machines that may have gone to sleep due to lack of usage when there is a large queue of pending jobs. The decision to utilise dormant resources will be made on a variety of factors such as job priority, job requirements, user priority, time of day, flocking options, queue conditions etc. Good practice scheduling policies would need to be devised that would work within this “green IT” pool

The Drosophila serpins: multiple functions in immunity and morphogenesis

Members of the serpin superfamily of proteins have been found in all living organisms, although rarely in bacteria or fungi. They have been extensively studied in mammals, where many rapid physiological responses are regulated by inhibitory serpins. In addition to the inhibitory serpins, a large group of noninhibitory proteins with a conserved serpin fold have also been identified in mammals. These noninhibitory proteins have a wide range of functions, from storage proteins to molecular chaperones, hormone transporters, and tumor suppressors. In contrast, until recently, very little was known about insect serpins in general, or Drosophila serpins in particular. In the last decade, however, there has been an increasing interest in the serpin biology of insects. It is becoming clear that, like in mammals, a similar wide range of physiological responses are regulated in insects and that noninhibitory serpin-fold proteins also play key roles in insect biology. Drosophila is also an important model organism that can be used to study human pathologies (among which serpinopathies or other protein conformational diseases) and mechanisms of regulation of proteolytic cascades in health or to develop strategies for control of insect pests and disease vectors. As most of our knowledge on insect serpins comes from studies on the Drosophila immune response, we survey here the Drosophila serpin literature and describe the laboratory techniques that have been developed to study serpin-regulated responses in this model genetic organism

Genetic and phenotypic analysis of the genes of the elbow-no-ocelli region of chromosome 2L of Dvosophila melanogaster

The elbow locus is found to be two genes elA and elB, each of which has a distinct phenotype when mutant. Mutations of the elA gene have a strong phenotype where the wing is markedly disrupted. Mutations of elB are weak, mainly affecting the alula and the wing bristles. The two genes are dominant enhancers of each other. Homozygous deletion of the complete elbow region results in lethality. Situated between the elbow genes is the pupal gene and a locus which when deleted causes a crippled leg phenotype. This locus may be a control region for elbow. Immediately adjacent on the proximal side of elA is the no-ocelli locus. The phenotypes of noc alleles vary from extreme, where the ocelli and associated bristles are absent, to weak where these structures are disrupted. The various noc phenotypes are associated with genetically distinct gene regions, mutations of which act as enhancers of each other. Alleles of el and noc show partial failure of complementation, heterozygotes having weak el or weak noc phenotypes. Alleles of both these genes interact with the antimorphic noc allele Sco