The angiotensin-converting enzyme (ACE) gene family of Anopheles gambiae

Background Members of the M2 family of peptidases, related to mammalian angiotensin converting enzyme (ACE), play important roles in regulating a number of physiological processes. As more invertebrate genomes are sequenced, there is increasing evidence of a variety of M2 peptidase genes, even within a single species. The function of these ACE-like proteins is largely unknown. Sequencing of the A. gambiae genome has revealed a number of ACE-like genes but probable errors in the Ensembl annotation have left the number of ACE-like genes, and their structure, unclear. Results TBLASTN and sequence analysis of cDNAs revealed that the A. gambiae genome contains nine genes (AnoACE genes) which code for proteins with similarity to mammalian ACE. Eight of these genes code for putative single domain enzymes similar to other insect ACEs described so far. AnoACE9, however, has several features in common with mammalian somatic ACE such as a two domain structure and a hydrophobic C terminus. Four of the AnoACE genes (2, 3, 7 and 9) were shown to be expressed at a variety of developmental stages. Expression of AnoACE3, AnoACE7 and AnoACE9 is induced by a blood meal, with AnoACE7 showing the largest (approximately 10-fold) induction. Conclusion Genes coding for two-domain ACEs have arisen several times during the course of evolution suggesting a common selective advantage to having an ACE with two active-sites in tandem in a single protein. AnoACE7 belongs to a sub-group of insect ACEs which are likely to be membrane-bound and which have an unusual, conserved gene structure

lemmingA encodes the Apc11 subunit of the APC/C in Drosophila melanogaster that forms a ternary complex with the E2-C type ubiquitin conjugating enzyme, Vihar and Morula/Apc2

<p>Abstract</p> <p>Background</p> <p>Ubiquitin-dependent protein degradation is a critical step in key cell cycle events, such as metaphase-anaphase transition and mitotic exit. The anaphase promoting complex/cyclosome (APC/C) plays a pivotal role in these transitions by recognizing and marking regulatory proteins for proteasomal degradation. Its overall structure and function has been elucidated mostly in yeasts and mammalian cell lines. The APC/C is, however, a multisubunit assembly with at least 13 subunits and their function and interaction within the complex is still relatively uncharacterized, particularly in metazoan systems. Here, <it>lemming </it>(<it>lmg</it>) mutants were used to study the APC/C subunit, Apc11, and its interaction partners in <it>Drosophila melanogaster</it>.</p> <p>Results</p> <p>The <it>lmg </it>gene was initially identified through a pharate adult lethal P element insertion mutation expressing developmental abnormalities and widespread apoptosis in larval imaginal discs and pupal abdominal histoblasts. Larval neuroblasts were observed to arrest mitosis in a metaphase-like state with highly condensed, scattered chromosomes and frequent polyploidy. These neuroblasts contain high levels of both cyclin A and cyclin B. The <it>lmg </it>gene was cloned by virtue of the <it>lmg⁰³⁴²⁴</it>P element insertion which is located in the 5' untranslated region. The <it>lemming </it>locus is transcribed to give a 2.0 kb mRNA that contains two ORFs, <it>lmgA </it>and <it>lmgB</it>. The <it>lmgA </it>ORF codes for a putative protein with more than 80% sequence homology to the APC11 subunit of the human APC/C. The 85 amino acid protein also contains a RING-finger motif characteristic of known APC11 subunits. The <it>lmgA </it>ORF alone was sufficient to rescue the lethal and mitotic phenotypes of the <it>lmg¹³⁸</it>null allele and to complement the temperature sensitive lethal phenotype of the <it>APC11-myc9 </it>budding yeast mutant. The LmgA protein interacts with Mr/Apc2, and they together form a binding site for Vihar, the E2-C type ubiquitin conjugating enzyme. Despite being conserved among <it>Drosophila </it>species, the LmgB protein is not required for viability or fertility.</p> <p>Conclusions</p> <p>Our work provides insight into the subunit structure of the <it>Drosophila </it>APC/C with implications for its function. Based on the presented data, we suggest that the Lmg/Apc11 subunit recruits the E2-C type ubiquitin conjugating enzyme, Vihar, to the APC/C together with Mr/Apc2 by forming a ternary complex.</p

The Drosophila insulin receptor independently modulates lifespan and locomotor senescence

The Insulin/IGF-like signalling (IIS) pathway plays an evolutionarily conserved role in ageing. In model organisms reduced IIS extends lifespan and ameliorates some forms of functional senescence. However, little is known about IIS in nervous system ageing and behavioural senescence. To investigate this role in Drosophila melanogaster, we measured the effect of reduced IIS on senescence of two locomotor behaviours, negative geotaxis and exploratory walking. Two long-lived fly models with systemic IIS reductions (daGAL4/UAS-InRDN (ubiquitous expression of a dominant negative insulin receptor) and d2GAL/UAS-rpr (ablation of insulin-like peptide producing cells)) showed an amelioration of negative geotaxis senescence similar to that previously reported for the long-lived IIS mutant chico. In contrast, exploratory walking in daGAL4/UAS-InRDN and d2GAL/UAS-rpr flies declined with age similarly to controls. To determine the contribution of IIS in the nervous system to these altered senescence patterns and lifespan, the InRDN was targeted to neurons (elavGAL4/UAS-InRDN), which resulted in extension of lifespan in females, normal negative geotaxis senescence in males and females, and detrimental effects on age-specific exploratory walking behaviour in males and females. These data indicate that the Drosophila insulin receptor independently modulates lifespan and age-specific function of different types of locomotor behaviour. The data suggest that ameliorated negative geotaxis senescence of long-lived flies with systemic IIS reductions is due to ageing related effects of reduced IIS outside the nervous system. The lifespan extension and coincident detrimental or neutral effects on locomotor function with a neuron specific reduction (elavGAL4/UAS-InRDN) indicates that reduced IIS is not beneficial to the neural circuitry underlying the behaviours despite increasing lifespan

Development of a real-time qPCR assay for quantification of covert baculovirus infections in a major African crop pest

Many pathogens and parasites are present in host individuals and populations without any obvious signs of disease. This is particularly true for baculoviruses infecting lepidopteran hosts, where studies have shown that covert persistent viral infections are almost ubiquitous in many species. To date, the infection intensity of covert viruses has rarely been quantified. In this study, we investigated the dynamics of a covert baculovirus infection within the lepidopteran crop pest Spodoptera exempta. A real-time quantitative polymerase chain reaction (qPCR) procedure using a 5' nuclease hydrolysis (TaqMan) probe was developed for specific detection and quantification of Spodoptera exempta nucleopolyhedrovirus (SpexNPV). The qPCR assay indicated that covert baculovirus dynamics varied considerably over the course of the host life-cycle, with infection load peaking in early larval instars and being lowest in adults and final-instar larvae. Adult dissections indicated that, contrary to expectation, viral load aggregation was highest in the head, wings and legs, and lowest in the thorax and abdomen. The data presented here have broad implications relating to our understanding of transmission patterns of baculoviruses and the role of covert infections in host-pathogen dynamics

Loss of angiotensin-converting enzyme-related (ACER) peptidase disrupts behavioural and metabolic responses to diet in Drosophila melanogaster

Drosophila Acer (Angiotensin-converting enzyme-related) encodes a member of the angiotensin-converting enzyme (ACE) family of metallopeptidases that in mammals play roles in the endocrine regulation of blood homeostasis. ACE is also expressed in adipose tissue where it is thought to play a role in metabolic regulation. Drosophila Acer is expressed in the adult fat body of the head and abdomen and is secreted into the haemolymph. Acer null mutants have previously been found to have reduced night time sleep and greater sleep fragmentation. Acer may thus be part of a signalling system linking metabolism with sleep. To further understand the role of Acer in response to diet, we measured sleep and other nutrient-responsive phenotypes in Acer null flies under different dietary conditions. We show that loss of Acer disrupts the normal response of sleep to changes in nutrition. Other nutrient sensitive phenotypes, including survival and glycogen storage, were also altered in the Acer mutant but lipid storage was not. Although the physiological substrate of the Acer peptidase has not been identified, an alteration of the normal nutrient dependent control of Drosophila insulin-like peptide 5 protein in the Acer mutant suggests insulin/IGF-like signalling as a candidate pathway modulated by Acer in the nutrient-dependent control of sleep, survival and metabolism

Drosophila type XV/XVIII collagen, Mp, is involved in Wingless distribution

Multiplexin (Mp) is the Drosophila orthologue of vertebrate collagens XV and XVIII. Like them, Mp is widely distributed in the basement membranes of the developing embryos, including those of neuroblasts in the central and peripheral nervous systems, visceral muscles of the gut, and contractile cardioblasts. Here we report the identification of mutant larvae bearing piggyBac transposon insertions that exhibit decrease Mp production associated with abdominal cuticular and wing margin defects, malformation of sensory organs and impaired sensitivity to physical stimuli. Additional findings include the abnormal ultrastructure of fatbody associated with abnormal collagen IV deposition, and reduced Wingless deposition. Collectively, these findings are consistent with the notion that Mp is required for the proper formation and/or maintenance of basement membrane, and that Mp may be involved in establishing the Wingless signaling gradients in the Drosophila embryo

Ecdysteroids affect Drosophila ovarian stem cell niche formation and early germline differentiation

Ecdysteroids affect Drosophila ovarian stem cell niche formation and early germline differentiation. The steroid hormone ecdysone regulates germline development and stem cell niche establishment in the Drosophila ovary by modulating TGF-β signalling and cell adhesion

A Buoyancy-Based Screen of Drosophila Larvae for Fat-Storage Mutants Reveals a Role for Sir2 in Coupling Fat Storage to Nutrient Availability

Obesity has a strong genetic component, but few of the genes that predispose to obesity are known. Genetic screens in invertebrates have the potential to identify genes and pathways that regulate the levels of stored fat, many of which are likely to be conserved in humans. To facilitate such screens, we have developed a simple buoyancy-based screening method for identifying mutant Drosophila larvae with increased levels of stored fat. Using this approach, we have identified 66 genes that when mutated increase organismal fat levels. Among these was a sirtuin family member, Sir2. Sirtuins regulate the storage and metabolism of carbohydrates and lipids by deacetylating key regulatory proteins. However, since mammalian sirtuins function in many tissues in different ways, it has been difficult to define their role in energy homeostasis accurately under normal feeding conditions. We show that knockdown of Sir2 in the larval fat body results in increased fat levels. Moreover, using genetic mosaics, we demonstrate that Sir2 restricts fat accumulation in individual cells of the fat body in a cell-autonomous manner. Consistent with this function, changes in the expression of metabolic enzymes in Sir2 mutants point to a shift away from catabolism. Surprisingly, although Sir2 is typically upregulated under conditions of starvation, Sir2 mutant larvae survive better than wild type under conditions of amino-acid starvation as long as sugars are provided. Our findings point to a Sir2-mediated pathway that activates a catabolic response to amino-acid starvation irrespective of the sugar content of the diet