Biology subjects

Lemaitre, B.

Publications and source records attributed to Lemaitre, B..

32 records · Page 2Linked to original sources

bioRxiv2021

Cecropins contribute to Drosophila host defense against fungal and Gram-negative bacterial infection

Cecropins are small helical secreted peptides with antimicrobial activity that are widely distributed among insects. Genes encoding Cecropins are strongly induced upon infection, pointing to their role in host-defence. In Drosophila, four Cecropin genes clustered in the genome (CecA1, CecA2, CecB and CecC) are expressed upon infection downstream of the Toll and Imd pathways. In this study, we generated a short deletion{Delta} CecA-C removing the whole Cecropin locus. Using the{Delta} CecA-C deficiency alone or in combination with other antimicrobial peptide (AMP) mutations, we addressed the function of Cecropins in the systemic immune response.{Delta} CecA-C flies were viable and resisted challenge with various microbes as wild-type. However, removing{Delta} CecA-C in flies already lacking ten other AMP genes revealed a role for Cecropins in defence against Gram-negative bacteria and fungi. Measurements of pathogen loads confirm that Cecropins contribute to the control of certain Gram-negative bacteria, notably Enterobacter cloacae and Providencia heimbachae. Collectively, our work provides the first genetic demonstration of a role for Cecropins in insect host defence, and confirms their in vivo activity primarily against Gram-negative bacteria and fungi. Generation of a fly line ({Delta}AMP14) that lacks fourteen immune inducible AMPs provides a powerful tool to address the function of these immune effectors in host-pathogen interactions and beyond.

immunology↗

bioRxiv2021

Drosophila antimicrobial peptides and lysozymes regulate gut microbiota composition and abundance

The gut microbiota affects the physiology and metabolism of animals and its alteration can lead to diseases such as gut dysplasia or metabolic disorders. Several reports have shown that the immune system plays an important role in shaping both bacterial community composition and abundance in Drosophila, and that immune deficit, especially during aging, negatively affects microbiota richness and diversity. However, there has been little study at the effector level to demonstrate how immune pathways regulate the microbiota. A key set of Drosophila immune effectors are the antimicrobial peptides (AMPs), which confer defense upon systemic infection. AMPs and lysozymes, a group of digestive enzymes with antimicrobial properties, are expressed in the gut and are good candidates for microbiota regulation. Here, we take advantage of the model organism Drosophila melanogaster to investigate the role of AMPs and lysozymes in regulation of gut microbiota structure and diversity. Using flies lacking AMPs and newly generated lysozyme mutants, we colonized gnotobiotic flies with a defined set of commensal bacteria and analyzed changes in microbiota composition and abundance in vertical transmission and aging contexts through 16S rRNA gene amplicon sequencing. Our study shows that AMPs and, to a lesser extent, lysozymes are necessary to regulate the total and relative abundance of bacteria in the gut microbiota. We also decouple the direct function of AMPs from the IMD signaling pathway that regulates AMPs but also many other processes, more narrowly defining the role of these effectors in the microbial dysbiosis observed in IMD-deficient flies upon aging. ImportanceThis study advances current knowledge in the field of host-microbe interactions by demonstrating that the two families of immune effectors, antimicrobial peptides and lysozymes, actively regulate the gut microbiota composition and abundance. Consequences of the loss of these antimicrobial peptides and lysozymes are exacerbated during aging, and their loss contributes to increased microbiota abundance and shifted composition in old flies. This work shows that immune effectors, typically associated with resistance to pathogenic infections, also help shape the beneficial gut community, consistent with the idea that host-symbiont interactions use the same language typically associated with pathogenesis.

microbiology↗

bioRxiv2021

Extensive duplication and convergent sequence evolution of an antimicrobial peptide gene

Antimicrobial peptides (AMPs) are host-encoded antibiotics that combat invading pathogens. These genes commonly encode multiple products as post-translationally cleaved polypeptides. Recent studies have highlighted roles for AMPs in neurological contexts suggesting functions for these defence molecules beyond infection. During our immune study characterizing the antimicrobial peptide gene Baramicin, we recovered multiple Baramicin paralogs in Drosophila melanogaster and other species, united by their N-terminal IM24 domain. Not all paralogs were immune-induced. Here, through careful dissection of the Baramicin familys evolutionary history, we find that these non-immune paralogs result from repeated events of duplication and subsequent truncation of the coding sequence from an immune-inducible ancestor. These truncations leave only the IM24 domain as the prominent gene product. Surprisingly, using mutation and targeted gene silencing we demonstrate that two such genes are adapted for function in neural contexts in D. melanogaster. We also show enrichment in the head for independent Baramicin genes in other species. The Baramicin evolutionary history reveals that the IM24 Baramicin domain is not strictly useful in an immune context. We thus provide a case study for how an AMP-encoding gene might play dual roles in both immune and non-immune processes via its multiple peptide products. We reflect on these findings to highlight a blind spot in the way researchers approach AMP research in in vivo contexts. Significance statementAntimicrobial peptides are immune proteins recently implicated in neurological roles. To date little attention has been paid to the contributions of different gene products in this function. Here we show that an antimicrobial peptide gene encodes multiple products with either immune-specific or neurological roles.

evolutionary biology↗

bioRxiv2021

Dual proteomics of Drosophila melanogaster hemolymph infected with the heritable endosymbiont Spiroplasma poulsonii

Insects are frequently infected with heritable bacterial endosymbionts. Endosymbionts have a dramatic impact on their host physiology and evolution. Their tissue distribution is variable with some species being housed intracellularly, some extracellularly and some having a mixed lifestyle. The impact of extracellular endosymbionts on the biofluids they colonize (e.g. insect hemolymph) is however difficult to appreciate because biofluids composition can depend on the contribution of numerous tissues. Here we address the question of Drosophila hemolymph proteome response to the infection with the endosymbiont Spiroplasma poulsonii. S. poulsonii inhabits the fly hemolymph and gets vertically transmitted over generations by hijacking the oogenesis in females. Using dual proteomics on infected hemolymph, we uncovered a low level and chronic activation of the Toll immune pathway by S. poulsonii that was previously undetected by transcriptomics-based approaches. Using Drosophila genetics, we also identified candidate proteins putatively involved in controlling S. poulsonii growth. Last, we also provide a deep proteome of S. poulsonii, which, in combination with previously published transcriptomics data, improves our understanding of the post-transcriptional regulations operating in this bacterium. Summary statementWe report the changes in Drosophila melanogaster hemolymph proteome upon infection with the heritable bacterial endosymbiont Spiroplasma poulsonii.

microbiology↗

bioRxiv2020

The Drosophila Baramicin polypeptide gene protects against fungal infection

The fruit fly Drososphila melanogaster combats microbial infection by producing a battery of effector peptides that are secreted into the haemolymph. Technical difficulties prevented the investigation of these short effector genes until the recent advent of the CRISPR/CAS era. As a consequence, many putative immune effectors remain to be characterized and exactly how each of these effectors contributes to survival is not well characterized. Here we describe a novel Drosophila antifungal peptide gene that we name Baramicin A. We show that BaraA encodes a precursor protein cleaved into multiple peptides via furin cleavage sites. BaraA is strongly immune-induced in the fat body downstream of the Toll pathway, but also exhibits expression in the nervous system. Importantly, we show that flies lacking BaraA are viable but susceptible to the enomopathogenic fungus Beauveria bassiana. Consistent with BaraA being directly antimicrobial, overexpression of BaraA promotes resistance to fungi and the IM10-like peptides produced by BaraA synergistically inhibit growth of fungi in vitro when combined with a membrane-disrupting antifungal. Surprisingly, BaraA males but not females display an erect wing phenotype upon infection. Collectively, we identify a new antifungal immune effector downstream of Toll signalling, improving our knowledge of the Drosophila antimicrobial response.

immunology↗

bioRxiv2020

Rapid molecular evolution of Spiroplasma symbionts of Drosophila

Spiroplasma are a group of Mollicutes whose members include plant pathogens, insect pathogens, and endosymbionts of animals. Spiroplasma phenotypes have been repeatedly observed to be spontaneously lost in Drosophila cultures, and several studies have documented a high genomic turnover in Spiroplasma symbionts and plant pathogens. These observations suggest that Spiroplasma evolves quickly in comparison to other insect symbionts. Here, we systematically assess evolutionary rates and patterns of Spiroplasma poulsonii, a natural symbiont of Drosophila. We analysed genomic evolution of sHy within flies, and sMel within in vitro culture over several years. We observed that S. poulsonii substitution rates are among the highest reported for any bacteria, and around two orders of magnitude higher compared with other inherited arthropod endosymbionts. The absence of mismatch repair loci mutS and mutL is conserved across Spiroplasma and likely contributes to elevated substitution rates. Further, the closely related strains sMel and sHy (>99.5% sequence identity in shared loci) show extensive structural genomic differences, which potentially indicates a higher degree of host adaptation in sHy, a protective symbiont of Drosophila hydei. Finally, comparison across diverse Spiroplasma lineages confirms previous reports of dynamic evolution of toxins, and identifies loci similar to the male-killing toxin Spaid in several Spiroplasma lineages and other endosymbionts. Overall, our results highlight the peculiar nature of Spiroplasma genome evolution, which may explain unusual features of its evolutionary ecology.

evolutionary biology↗

bioRxiv2020

Comparative RNA-Seq Analyses of Drosophila Plasmatocytes Reveal Gene Specific Signatures In Response To Clean Injury And Septic Injury

Drosophila melanogasters blood cells (hemocytes) play essential roles in wound healing and are involved in clearing microbial infections. Here, we report the transcriptional changes of larval plasmatocytes after clean injury or infection with the Gram-negative bacterium Escherichia coli or the Gram-positive bacterium Staphylococcus aureus compared to hemocytes recovered from unchallenged larvae via RNA-Sequencing. This study reveals 676 differentially expressed genes (DEGs) in hemocytes from clean injury samples compared to unchallenged samples, and 235 and 184 DEGs in E. coli and S. aureus samples respectively compared to clean injury samples. The clean injury samples showed enriched DEGs for immunity, clotting, cytoskeleton, cell migration, hemocyte differentiation, and indicated a metabolic reprogramming to aerobic glycolysis, a well-defined metabolic adaptation observed in mammalian macrophages. Microbial infections trigger significant transcription of immune genes, with significant differences between the E. coli and S. aureus samples suggesting that hemocytes have the ability to engage various programs upon infection. Collectively, our data bring new insights on Drosophila hemocyte function and open the route to post-genomic functional analysis of the cellular immune response.

immunology↗

bioRxiv2019

Metabolic adjustment of Drosophila hemocyte number and sessility by an adipokine

In animals, growth is regulated by the complex interplay between paracrine and endocrine signals. When food is scarce, tissues compete for nutrients, leading to critical resource allocation and prioritization. Little is known about how the immune system maturation is coordinated with the growth of other tissues. Here, we describe a signaling mechanism that regulates the number of hemocytes (blood cells) according to the nutritional state of the Drosophila larva. Specifically, we found that the adipokine NimB5 is produced in the fat body upon nutrient scarcity downstream of metabolic sensors. NimB5 is then secreted and bind to hemocytes to down-regulate their proliferation and adhesion. Blocking this signaling loop results in conditional lethality when larvae are raised on a poor diet, due to excessive hemocyte numbers and insufficient energy storage. Similar regulatory mechanisms shaping the immune system in response to nutrient availability are likely to be widespread in animals.\n\nAuthor summaryDrosophila larval hemocytes (blood cells) are found in two compartments: the lymph gland considered as a reservoir, and the peripheral compartment. Peripheral hemocytes form sessile patches attached to the internal surface of the larval body wall or are found freely circulating in the hemolymph. Little is known about the signals that regulate hemocytes proliferation and localization in the peripheral compartment. In this study, we have identified a new gene, NimrodB5, coding for the NimB5 protein, which is secreted by the fat body and binds to hemocytes. NimB5 inhibits hemocyte proliferation while promoting sessility, leading to an increased number of circulating hemocytes and adhesion defects in NimB5 mutant. We show that nimrodB5 expression by the fat body is controlled by metabolic cues to adjust hemocyte number to the physiological state of the larvae.\n\nInterestingly, deregulation of NimB5 causes lethality when larvae are raised on a poor diet due to a defect in regulating hemocytes proliferation. In conclusion, we have identified a new adipokine that optimizes hemocytes number to the physiological state of larvae. Our study also reveals a major role of the fat body in peripheral hematopoiesis regulation and outline how it can be costly to maintain a basal immune defense.

immunology↗

bioRxiv2019

Shavenbaby protein isoforms orchestrate the self-renewal versus differentiation of Drosophila intestinal stem cells

Signaling pathways are key regulators of adult stem cell homeostasis and underlying mechanisms are often deregulated in cancers. Recent studies of epithelial tumors have involved OvoL/Svb transcription factors, which produce isoforms with antagonistic activities. Here we show that Svb, the unique OvoL factor in Drosophila, directly integrates multiple signaling inputs to coordinate the behavior of adult intestinal stem cell lineage. Under steady state, Svb mediates Wnt and EGFR signaling to ensure stem cell renewal and progenitor survival. This requires the post-translational processing of Svb into a transcriptional activator by Polished rice (Pri) regulatory peptides, under the regulation of ecdysone signaling. In response to PDM1, Svb expression is specifically maintained in enterocytes where it acts as a transcriptional repressor sufficient to override mitogenic signals and impose differentiation. Altogether, these results demonstrate that the OvoL/Svb transcriptional switch controls the balance between stem cell survival, self-renewal and differentiation.

developmental biology↗

bioRxiv2019

Blood cells of adult Drosophila do not expand, but control survival after bacterial infection by induction of Drosocin around their reservoir at the respiratory epithelia

Drosophila melanogaster has been an excellent model for innate immunity, but the role and regulation of adult blood cells and organismal immunity have remained incompletely understood. Here we address these questions in a comprehensive investigation of the blood cell system in adult Drosophila. As a central finding, we reveal the largest reservoir of blood cells (hemocytes) at the respiratory epithelia (tracheal air sacs) and fat body of the thorax and head. We show that most hemocytes of adult Drosophila are phagocytic macrophages (plasmatocytes), derived by more than 60% from the embryonic lineage that parallels vertebrate tissue macrophages. Surprisingly, in contrast to hemocytes at the larval stage, we find no capacity of the adult blood cell system to expand. Instead, we demonstrate its central role in relaying an innate immune response to tissues surrounding the blood cell reservoir: Hemocytes, through Imd signaling and the Jak/Stat pathway ligand Upd3, act as sentinels of bacterial infection that induce expression of the antimicrobial peptide gene Drosocin in the respiratory epithelia and colocalizing domains of the fat body. We demonstrate that endogenous Drosocin expression in these tissues promotes animal survival after bacterial infection. Our work identifies the first molecular step in a new relay of organismal immunity, establishing adult Drosophila as model to dissect mechanisms of inter-organ immunity.

immunology↗

bioRxiv2019

The antimicrobial peptide Defensin cooperates with Tumour Necrosis Factor to drive tumour cell death in Drosophila

Antimicrobial peptides (AMPs) are small cationic molecules best known as mediators of the innate defence against microbial infection. While in vitro and ex vivo evidence suggest AMPs capacity to kill cancer cells, in vivo demonstration of an anti-tumour role of endogenous AMPs is lacking. Using a Drosophila model of tumourigenesis, we demonstrate a role for the AMP Defensin in the control of tumour progression. Our results reveal that Tumour Necrosis Factor mediates exposure of phosphatidylserine (PS), which makes tumour cells selectively sensitive to the action of Defensin remotely secreted from tracheal and fat tissues. Defensin binds tumour cells in PS-enriched areas, provoking cell death and tumour regression. Altogether, our results provide the first in vivo demonstration for a role of an endogenous AMP as an anti-cancer agent, as well as a mechanism that explains tumour cell sensitivity to the action of AMPs.

cancer biology↗

bioRxiv2018

Synergy and remarkable specificity of antimicrobial peptides in vivo using a systematic knockout approach

Antimicrobial peptides (AMPs) are host-encoded antibiotics that combat invading microorganisms. These short, cationic peptides have been implicated in many biological processes, primarily involving innate immunity. In vitro studies have shown AMPs kill bacteria and fungi at physiological concentrations, but little validation has been done in vivo. We utilised CRISPR gene editing to delete all known immune inducible AMPs of Drosophila, namely: 4 Attacins, 4 Cecropins, 2 Diptericins, Drosocin, Drosomycin, Metchnikowin and Defensin. Using individual and multiple knockouts, including flies lacking all 14 AMP genes, we characterize the in vivo function of individual and groups of AMPs against diverse bacterial and fungal pathogens. We found that Drosophila AMPs act primarily against Gram-negative bacteria and fungi, acting either additively or synergistically. We also describe remarkable specificity wherein certain AMPs contribute the bulk of microbicidal activity against specific pathogens, providing functional demonstrations of highly specific AMP-pathogen interactions in an in vivo setting.

immunology↗

bioRxiv2018

Two Nimrod receptors, NimC1 and Eater, synergistically contribute to bacterial phagocytosis in Drosophila melanogaster

Eater and NimC1 are transmembrane receptors of the Drosophila Nimrod family, specifically expressed in hemocytes, the insect blood cells. Previous ex vivo and in vivo RNAi studies have pointed to their role in the phagocytosis of bacteria. Here, we have created a novel null mutant in NimC1 to re-evaluate the role of NimC1, alone or in combination with Eater, in the cellular immune response. We show that NimC1 functions as an adhesion molecule ex vivo, but in contrast to Eater is not required for hemocyte sessility in vivo. Ex vivo phagocytosis assays and electron microscopy experiments confirmed that Eater is the main phagocytic receptor for Gram-positive, but not Gram-negative bacteria, and contributes to microbe tethering to hemocytes. Surprisingly, the NimC1 deletion did not impair phagocytosis of bacteria, nor their adhesion to the hemocytes. However, phagocytosis of both types of bacteria was almost abolished in NimC11;eater1 hemocytes. This indicates that both receptors contribute synergistically to the phagocytosis of bacteria, but that Eater can bypass the requirement for NimC1. Finally, we uncovered that NimC1, but not Eater, is essential for uptake of latex beads and zymosan particles. We conclude that Eater and NimC1 are the two main receptors for phagocytosis of bacteria in Drosophila, and that each receptor likely plays distinct roles in microbial uptake.

immunology↗

bioRxiv2018

More than black and white: complex relationships involving serine proteases regulate the Toll pathway and the melanization response in Drosophila

The melanization response is a rapid and important defense mechanism in arthropods. Melanin is produced around wound sites and invading microorganisms by phenoloxidases (POs), which need to be activated by the sequential activation of an extracellular serine protease (SP) cascade. Drosophila melanogaster has been a useful genetic model for dissecting insect immune signaling, but understanding these proteolytic cascades has been complicated by the large number of SP genes, possibly with redundant function. Taking advantage of recently-generated null and compound mutants, we re-investigated the role of SPs involved in the melanization response in D. melanogaster and discovered phenotypes previously concealed in single mutant analysis. We found that two of them, Hayan and Sp7, can activate the melanization response in two different manners: Hayan is required in the local blackening of wound sites, while Sp7 regulates an alternate melanization reaction responsible for the clearance of septic infections with Staphylococcus aureus. We present evidence that both Sp7 and Hayan regulate the Toll NF-kB pathway. Sp7 is regulated by canonical Toll signaling downstream of PGRP-SA, ModSP, and Grass, leading to control of septic infections via a Sp7-dependent melanization response. Additionally, we found that Hayan and the Toll-regulating SP Psh are the result of a recent gene duplication. Using genetic manipulations, we reveal the hidden role for Hayan, alongside Psh, in propagating Toll signaling downstream of pattern recognition receptors. Thus, we describe the existence of two pathways leading to the melanization response and reveal previously unknown dynamics in the activation of the Toll pathway.

immunology↗