Introduction: Chitinase-like proteins (CLPs) are associated with tissue-remodeling and inflammation but also with several disorders, including fibrosis, atherosclerosis, allergies, and cancer. However, CLP’s role in tumors is far from clear.

Methods: Here, we utilize Drosophila melanogaster and molecular genetics to investigate the function of CLPs (imaginal disc growth factors; Idgf’s) in Ras^V12 dysplastic salivary glands.

Results and discussion: We find one of the Idgf’s members, Idgf3, is transcriptionally induced in a JNK-dependent manner via a positive feedback loop mediated by reactive oxygen species (ROS). Moreover, Idgf3 accumulates in enlarged endosomal vesicles (EnVs) that promote tumor progression by disrupting cytoskeletal organization. The process is mediated via the downstream component, aSpectrin, which localizes to the EnVs. Our data provide new insight into CLP function in tumors and identifies specific targets for tumor control.

Introduction: Tumor-associated macrophages may act to either limit or promote tumor growth, yet the molecular basis for either path is poorly characterized.

Methods: We use a larval Drosophila model that expresses a dominant-active version of the Ras-oncogene (Ras^V12) to study dysplastic growth during early tumor progression. We performed single-cell RNA-sequencing of macrophage-like hemocytes to characterize these cells in tumor- compared to wild-type larvae. Hemocytes included manually extracted tumor-associated- and circulating cells.

Results and discussion: We identified five distinct hemocyte clusters. In addition to Ras^V12 larvae, we included a tumor model where the activation of effector caspases was inhibited, mimicking an apoptosis-resistant setting. Circulating hemocytes from both tumor models differ qualitatively from control wild-type cells—they display an enrichment for genes involved in cell division, which was confirmed using proliferation assays. Split analysis of the tumor models further reveals that proliferation is strongest in the caspase-deficient setting. Similarly, depending on the tumor model, hemocytes that attach to tumors activate different sets of immune effectors—antimicrobial peptides dominate the response against the tumor alone, while caspase inhibition induces a shift toward members of proteolytic cascades. Finally, we provide evidence for transcript transfer between hemocytes and possibly other tissues. Taken together, our data support the usefulness of Drosophila to study the response against tumors at the organismic level.

Pyroptosis has been described in mammalian systems to be a form of programmed cell death that is important in immune function through the subsequent release of cytokines and immune effectors upon cell bursting. This form of cell death has been increasingly well-characterized in mammals and can occur using alternative routes however, across phyla, there has been little evidence for the existence of pyroptosis. Here we provide evidence for an ancient origin of pyroptosis in an in vivo immune scenario in Drosophila melanogaster. Crystal cells, a type of insect blood cell, were recruited to wounds and ruptured subsequently releasing their cytosolic content in a caspase-dependent manner. This inflammatory-based programmed cell death mechanism fits the features of pyroptosis, never before described in an in vivo immune scenario in insects and relies on ancient apoptotic machinery to induce proto-pyroptosis. Further, we unveil key players upstream in the activation of cell death in these cells including the apoptosome which may play an alternative role akin to the inflammasome in proto-pyroptosis. Thus, Drosophila may be a suitable model for studying the functional significance of pyroptosis in the innate immune system.

Insects rely on their innate immune system to successfully mediate complex interactions with their internal microbiota, as well as the microbes present in the environment. Given the variation in microbes across habitats, the challenges to respond to them are likely to result in local adaptations in the immune system. Here we focus upon phagocytosis, a mechanism by which pathogens and foreign particles are engulfed in order to be contained, killed, and processed. We investigated the phenotypic and genetic variation related to phagocytosis in two allopatric populations of the butterfly Pieris napi. Populations were found to differ in their hemocyte composition and overall phagocytic capability, driven by the increased phagocytic propensity of each cell type. Yet, genes annotated to phagocytosis showed no large genomic signal of divergence. However, a gene set enrichment analysis on significantly divergent genes identified loci involved in glutamine metabolism, which recently have been linked to immune cell differentiation in mammals. Together these results suggest that heritable variation in phagocytic capacity arises via a quantitative trait architecture with variation in genes affecting the activation and/or differentiation of phagocytic cells, suggesting them as potential candidate genes underlying these phenotypic differences.

Fibrotic lesions accompany several pathological conditions, including tumors. We show that expression of a dominant-active form of the Ras oncogene in Drosophila salivary glands (SGs) leads to redistribution of components of the basement membrane (BM) and fibrotic lesions. Similar to several types of mammalian fibrosis, the disturbed BM attracts clot components, including insect transglutaminase and phenoloxidase. SG epithelial cells show reduced apicobasal polarity accompanied by a loss of secretory activity. Both the fibrotic lesions and the reduced cell polarity are alleviated by ectopic expression of the antimicrobial peptide drosomycin (Drs), which also restores the secretory activity of the SGs. In addition to extracellular matrix components, both Drs and F-actin localize to fibrotic lesions.

Understanding the tradeoffs that result from successful infection responses is central to understanding how life histories evolve. Gaining such insights, however, can be challenging, as they may be pathogen specific and confounded with experimental design. Here, we investigated whether infection from gram positive or negative bacteria results in different physiological tradeoffs, and whether these infections impact life history later in life (post-diapause development), in the butterfly Pieris napi. During the first 24 h after infection (3, 6, 12, and 24 h), after removing effects due to injection, larvae infected with Micrococcus luteus showed a strong suppression of all non-immunity related processes while several types of immune responses were upregulated. In contrast, this tradeoff between homeostasis and immune response was much less pronounced in Escherichia coli infections. These differences were also visible long after infection, via weight loss and slower development, as well as an increased mortality at higher infection levels during later stages of development. Individuals infected with M. luteus, compared to E. coli, had a higher mortality rate, and a lower pupal weight, developmental rate and adult weight. Further, males exhibited a more negative impact of infection than females. Thus, immune responses come at a cost even when the initial infection has been overcome, and these costs are likely to affect later life history parameters with fitness consequences.

Sensing and responding to changes in the cellular environments are essential for the diverse family of Apicomplexan parasites, which undergo complex life cycles comprised of both extracellular and obligate intracellular stages. Despite evidence of paramount roles for Ca²⁺, the molecular players behind how parasites sense Ca²⁺ and initiate Ca²⁺ signaling cascades have remained enigmatic. In a recent publication, Marquez-Nogueras et al., identify a transient receptor potential (TRP)-like channel in Toxoplasma gondii and show its implication in the crucial processes of parasite invasion and egress from host cells.

Endoparasitoid wasps are important natural enemies of many insect species and are major selective forces on the host immune system. Despite increased interest in insect antiparasitoid immunity, there is sparse information on the evolutionary dynamics of biological pathways and gene regulation involved in host immune defense outside Drosophila species. We de novo assembled transcriptomes from two beetle species and used time-course differential expression analysis to investigate gene expression differences in closely related species Galerucella pusilla and G. calmariensis that are, respectively, resistant and susceptible against parasitoid infection by Asecodes paividava parasitoids. Approximately 271 million and 224 million paired-ended reads were assembled and filtered to form 52,563 and 59,781 transcripts for G. pusilla and G. calmariensis, respectively. In the whole-transcriptome level, an enrichment of functional categories related to energy production, biosynthetic process, and metabolic process was exhibited in both species. The main difference between species appears to be immune response and wound healing process mounted by G. pusilla larvae. Using reciprocal BLAST against the Drosophila melanogaster proteome, 120 and 121 immune-related genes were identified in G. pusilla and G. calmariensis, respectively. More immune genes were differentially expressed in G. pusilla than in G. calmariensis, in particular genes involved in signaling, hematopoiesis, and melanization. In contrast, only one gene was differentially expressed in G. calmariensis. Our study characterizes important genes and pathways involved in different immune functions after parasitoid infection and supports the role of signaling and hematopoiesis genes as key players in host immunity in Galerucella against parasitoid wasps.

Several insect innate immune mechanisms are activated in response to infection by entomopathogenic nematodes (EPNs). In this review, we focus on the coagulation of hemolymph, which acts to stop bleeding after injury and prevent access of pathogens to the body cavity. After providing a general overview of invertebrate coagulation systems, we discuss recent findings in Drosophila melanogaster which demonstrate that clots protect against EPN infections. Detailed analysis at the cellular level provided insight into the kinetics of the secretion of Drosophila coagulation factors, including non-classical modes of secretion. Roughly, clot formation can be divided into a primary phase in which crosslinking of clot components depends on the activity of Drosophila transglutaminase and a secondary, phenoloxidase (PO)-dependent phase, characterized by further hardening and melanization of the clot matrix. These two phases appear to play distinct roles in two commonly used EPN infection models, namely Heterorhabditis bacteriophora and Steinernema carpocapsae. Finally, we discuss the implications of the coevolution between parasites such as EPNs and their hosts for the dynamics of coagulation factor evolution.

Entomopathogenic nematodes (EPNs) have been a useful model for studying wound healing in insects due to their natural mechanism of entering an insect host either through the cuticle or an orifice. While many experiments have shed light on nematode and host behavior, as well as the host immune response, details regarding early nematode entry and proliferative events have been limited. Using high-resolution microscopy, we provide data on the early infection kinetics of Heterorhabditis bacteriophora and its symbiotic bacteria, Photorhabdus luminescens. EPNs appendage themselves to the host and enter through the host cuticle with a drill-like mechanism while leaving their outer sheath behind. EPNs immediately release their symbiotic bacteria in the host which leads to changes in host behavior and septicemia within 6 h while EPNs travel through the host in a predictable manner, congregating in the anterior end of the host. This paper sheds light on the entry and proliferative events of EPN infection, which will further aid in our understanding of wound healing and host immune activation at a high spatiotemporal resolution.