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Animals typically have either compound eyes, or camera-type eyes, both of which have evolved repeatedly in the animal kingdom. Both eye types include two important kinds of cells: photoreceptor cells, which can be excited by light, and non-neuronal support cells (SupCs), which provide essential support to photoreceptors. At the molecular level deeply conserved genes that relate to the differentiation of photoreceptor cells have fueled a discussion on whether or not a shared evolutionary origin might be considered for this cell type. In contrast, only a handful of studies, primarily on the compound eyes of Drosophila melanogaster , have demonstrated molecular similarities in SupCs. D. melanogaster SupCs (Semper cells and primary pigment cells) are specialized eye glia that share several molecular similarities with certain vertebrate eye glia, including Müller glia. This led us to question if there could be conserved molecular signatures of SupCs, even in functionally different eyes such as the image-forming larval camera eyes of the sunburst diving beetle Thermonectus marmoratus . To investigate this possibility, we used an in-depth comparative whole-tissue transcriptomics approach. Specifically, we dissected the larval principal camera eyes into SupC- and retina-containing regions and generated the respective transcriptomes. Our analysis revealed several common features of SupCs including enrichment of genes that are important for glial function (e.g. gap junction proteins such as innexin 3), glycogen production (glycogenin), and energy metabolism (glutamine synthetase 1 and 2). To evaluate similarities, we compared our transcriptomes with those of fly (Semper cells) and vertebrate (Müller glia) eye glia as well as respective retinas. T. marmoratus SupCs were found to have distinct genetic overlap with both fly and vertebrate eye glia. These results suggest that T. marmoratus SupCs are a form of glia, and like photoreceptors, may be deeply conserved.

Lenses are vital components of well-functioning eyes and are crafted through the precise arrangement of proteins to achieve transparency and refractive ability. In addition to optical clarity for minimal scatter and absorption, proper placement of the lens within the eye is equally important for the formation of sharp, focused images on the retina. Maintaining these states is challenging due to dynamic and substantial post-embryonic eye and lens growth. Here, we gain insights into required processes through exploring the optical and visual consequences of silencing a key lens constituent in Thermonectus marmoratus sunburst diving beetle larvae. Using RNAi, we knocked down Lens3, a widely expressed cuticular lens protein during a period of substantial growth of their camera-type principal eyes. We show that lens3 RNAi results in the formation of opacities reminiscent of vertebrate lens ‘cataracts’, causing the projection of blurry and degraded images. Consequences of this are exacerbated in low-light conditions, evidenced by impaired hunting behaviour in this visually guided predator. Notably, lens focal lengths remained unchanged, suggesting that power and overall structure are preserved despite the absence of this major component. Further, we did not detect significant shifts in the in-vivo refractive states of cataract-afflicted larvae. This in stark contrast with findings in vertebrates, in which form-deprivation or the attenuation of image contrast, results in the dysregulation of eye growth, causing refractive errors such as myopia. Our results provide insights into arthropod lens construction and align with previous findings which point towards visual input being inconsequential for maintaining correctly focused eyes in this group. Our findings highlight the utility of T. marmoratus as a tractable model system to probe the aetiology of lens cataracts and refractive errors.

Symbiotic bacteria assist in maintaining homeostasis of the animal immune system. However, the molecular mechanisms that underlie symbiont-mediated host immunity are largely unknown. Tsetse flies (Glossina spp.) house maternally transmitted symbionts that regulate the development and function of their host’s immune system. Herein we demonstrate that the obligate mutualist, Wigglesworthia, up-regulates expression of odorant binding protein six in the gut of intrauterine tsetse larvae. This process is necessary and sufficient to induce systemic expression of the hematopoietic RUNX transcription factor lozenge and the subsequent production of crystal cells, which actuate the melanotic immune response in adult tsetse. Larval Drosophila’s indigenous microbiota, which is acquired from the environment, regulates an orthologous hematopoietic pathway in their host. These findings provide insight into the molecular mechanisms that underlie enteric symbiont-stimulated systemic immune system development, and indicate that these processes are evolutionarily conserved despite the divergent nature of host-symbiont interactions in these model systems. Bacteria live within all animals. While a small number of these microbes can cause disease, most promote the health and wellbeing of their host. Microbes that support their host and benefit from the close association are often referred to as symbionts. Animals can be negatively affected and even become diseased if their symbionts are disrupted. As a result, a more complete understanding of the molecular interactions between animal hosts and their beneficial microbes will lead to better treatments for a number of diseases. Tsetse flies are insects that harbor two bacterial symbionts, which are transferred from pregnant females to their larval offspring. If the offspring mature without these microbes, they fail to develop cells called hemocytes. These cells are normally found in the insect’s equivalent of blood – a fluid called hemolymph – and they comprise an important component of the insect’s immune system. Adult tsetse flies that lack hemocytes are susceptible to certain infections. These findings indicate that the bacterial symbionts induce the production of hemocytes in tsetse fly larvae via an unknown mechanism. Benoit et al. now reveal that the bacterial symbionts trigger tsetse flies to produce a small protein called “odorant binding protein 6”. This protein controls the generation of one specific type of hemocyte called crystal cells in developing larvae. Crystal cells are largely responsible for triggering the production of melanin, a protein involved in killing disease-causing microbes and inhibiting the loss of hemolymph from wound sites in the insect’s exoskeleton. Benoit et al. discovered that bacterial symbionts associated with the larvae of fruit flies also support the development of their host’s immune system. Although these symbionts are acquired from the external environment rather than from the insect’s parent, they too control the production of an odorant binding protein and crystal cells in their larval host. Collectively, these findings confirm that bacterial symbionts are critically important for the development of the immune systems of insects, and they show that this process has been conserved throughout evolution. Future studies are likely to focus on identifying which molecules from the symbionts stimulate their hosts to produce new hemolymph cells. Furthermore, identifying which tissues and cell types in the animal hosts are targets for these molecules will provide a more complete picture of the pathways that lead to the production of new hemolymph cells.

Cryptococcus neoformans is a fungus that is able to survive abnormally high levels of ionizing radiation (IR). The radiolysis of water by IR generates reactive oxygen species (ROS) such as H 2 O 2 and OH - . C . neoformans withstands the damage caused by IR and ROS through antioxidant production and enzyme-catalyzed breakdown of ROS. Given these particular cellular protein needs, questions arise whether transfer ribonucleic acids molecules (tRNAs) undergo unique chemical modifications to maintain their structure, stability, and/or function under such environmental conditions. Here, we investigated the effects of IR and H 2 O 2 exposure on tRNAs in C . neoformans . We experimentally identified the modified nucleosides present in C . neoformans tRNAs and quantified changes in those modifications upon exposure to oxidative conditions. To better understand these modified nucleoside results, we also evaluated tRNA pool composition in response to the oxidative conditions. We found that regardless of environmental conditions, tRNA modifications and transcripts were minimally affected. A rationale for the stability of the tRNA pool and its concomitant profile of modified nucleosides is proposed based on the lack of codon bias throughout the C . neoformans genome and in particular for oxidative response transcripts. Our findings suggest that C . neoformans can rapidly adapt to oxidative environments as mRNA translation/protein synthesis are minimally impacted by codon bias.

Diploptera punctata , also known as the Pacific beetle cockroach, is a viviparous cockroach that gives birth to live offspring and secretes a highly concentrated mixture of glycosylated proteins as a source of nourishment for developing embryos. These proteins are lipocalins that bind to lipids and crystallize in the gut of the embryo. A structure of milk crystals harvested from the embryos showed that the milk-derived crystals were heterogeneous and made of three proteins (called Lili-Mips). We hypothesized that the isoforms of Lili-Mip would display different affinities for fatty acids due to the ability of the pocket to bind multiple acyl chain lengths. We previously reported the structures of Lili-Mip from crystals grown in vivo and recombinantly expressed Lili-Mip2. These structures are similar, and both bind to several fatty acids. This study explores the specificity and affinity of fatty acid binding to recombinantly expressed Lili-Mip 1, 2 & 3. We show that all isoforms can bind to different fatty acids with similar affinities. We also report the thermostability of Lili-Mip is pH dependent, where stability is highest at acidic pH and declines as the pH increases to physiological levels near 7.0. We show that thermostability is an inherent property of the protein, and glycosylation and ligand binding do not change it significantly. Measuring the pH in the embryo’s gut lumen and gut cells suggests that the pH in the gut is acidic and the pH inside the gut cells is closer to neutral pH. In various crystal structures (reported here and previously by us), Phe-98 and Phe-100 occupy multiple conformations in the binding pocket. In our earlier work, we had shown that the loops at the entrance could adapt various conformations to change the size of the binding pocket. Here we show Phe-98 and Phe-100 can reorient to stabilize interactions at the bottom of the cavity–and change the volume of the cavity from 510 Å 3 to 337 Å 3 . Together they facilitate the binding of fatty acids of different acyl chain lengths.

Cyclic AMP (cAMP) signaling pathways are poorly understood in the stercorarian parasite Trypanosoma cruzi . Specifically, the mechanisms driving the activation of TcACs in response to microenvironmental stress are completely unknown. This study unveils the role of TcCARP3 in modulating the content of cAMP through the interaction with several TcACs and putative cAMP effectors in T. cruzi . Particularly, TcCARP3 interacts with TcAC1 in the main developmental stages of this parasite’s life cycle, where both proteins display a dual localization pattern. These results provide new evidence supporting the compartmentalization of cAMP signals in trypanosomes. Moreover, our data unequivocally demonstrates that TcCARP3 is required for essential cellular processes, such as response to osmotic stress, host cell invasion, intracellular replication, and the ability to colonize the hindgut of the triatomine vector. In summary, we found that TcCARP3 is an adenylate cyclase interactor that modulates cAMP signals necessary for the life cycle progression of T. cruzi .

Current insights into the mosquito dehydration response rely on studies that examine specific responses but ultimately fail to provide an encompassing view of mosquito biology. Here, we examined underlying changes in the biology of mosquitoes associated with dehydration. Specifically, we show that dehydration increases blood feeding in the northern house mosquito, Culex pipiens , which was the result of both higher activity and a greater tendency to land on a host. Similar observations were noted for Aedes aegypti and Anopheles quadrimaculatus . RNA-seq and metabolome analyses in C . pipiens following dehydration revealed that factors associated with carbohydrate metabolism are altered, specifically the breakdown of trehalose. Suppression of trehalose breakdown in C . pipiens by RNA interference reduced phenotypes associated with lower hydration levels. Lastly, mesocosm studies for C . pipiens confirmed that dehydrated mosquitoes were more likely to host feed under ecologically relevant conditions. Disease modeling indicates dehydration bouts will likely enhance viral transmission. This dehydration-induced increase in blood feeding is therefore likely to occur regularly and intensify during periods when availability of water is low.

Impact of reproductive processes upon female health has yielded conflicting results; particularly in relation to the role of reproduction-associated stress. We used the viviparous tsetse fly to determine if lactation, birth and involution lead to damage from oxidative stress (OS) that impairs subsequent reproductive cycles. Tsetse females carry an intrauterine larva to full term at each pregnancy cycle, and lactate to nourish them with milk secretions produced by the accessory gland ( = milk gland) organ. Unlike most K-strategists, tsetse females lack an apparent period of reproductive senescence allowing the production of 8-10 progeny over their entire life span. In a lactating female, over 47% of the maternal transcriptome is associated with the generation of milk proteins. The resulting single larval offspring weighs as much as the mother at birth. In studying this process we noted an increase in specific antioxidant enzyme (AOE) transcripts and enzymatic activity at critical times during lactation, birth and involution in the milk gland/fat body organ and the uterus. Suppression of superoxide dismutase (sod) decreased fecundity in subsequent reproductive cycles in young mothers and nearly abolished fecundity in geriatric females. Loss of fecundity was in part due to the inability of the mother to produce adequate milk to support larval growth. Longevity was also impaired after sod knockdown. Generation of OS in virgin females through exogenous treatment with hydrogen peroxide at times corresponding to pregnancy intervals reduced survival, which was exacerbated by sod knockdown. AOE expression may prevent oxidative damage associated with the generation of nutrients by the milk gland, parturition and milk gland breakdown. Our results indicate that prevention of OS is essential for females to meet the growing nutritional demands of juveniles during pregnancy and to repair the damage that occurs at birth. This process is particularly important for females to remain fecund during the latter portion of their lifetime.

Diseases that are transmitted by mosquitoes are a tremendous health and socioeconomic burden with hundreds of millions of people being impacted by mosquito-borne illnesses annually. Many factors have been implicated and extensively studied in disease transmission dynamics, but knowledge regarding how dehydration impacts mosquito physiology, behavior, and resulting mosquito-borne disease transmission remain underdeveloped. The lapse in understanding on how mosquitoes respond to dehydration stress likely obscures our ability to effectively study mosquito physiology, behavior, and vectorial capabilities. The goal of this review is to develop a profile of factors underlying mosquito biology that are altered by dehydration and the implications that are related to disease transmission.

High quality gene models are necessary to expand the molecular and genetic tools available for a target organism, but these are available for only a handful of model organisms that have undergone extensive curation and experimental validation over the course of many years. The majority of gene models present in biological databases today have been identified in draft genome assemblies using automated annotation pipelines that are frequently based on orthologs from distantly related model organisms and usually have minor or major errors. Manual curation is time consuming and often requires substantial expertise, but is instrumental in improving gene model structure and identification. Manual annotation may seem to be a daunting and cost-prohibitive task for small research communities but involving undergraduates in community genome annotation consortiums can be mutually beneficial for both education and improved genomic resources. We outline a workflow for efficient manual annotation driven by a team of primarily undergraduate annotators. This model can be scaled to large teams and includes quality control processes through incremental evaluation. Moreover, it gives students an opportunity to increase their understanding of genome biology and to participate in scientific research in collaboration with peers and senior researchers at multiple institutions.