Yang Cheng

The intestinal epithelium, predominantly composed of enterocytes (ECs), integrates dietary cues to regulate intestinal stem cell (ISC) activity and maintain intestinal homeostasis. However, the underlying mechanisms remain incompletely understood. Here, we demonstrate the functions of essential nutrient-sensing mediators Rag GTPases and mTORC1 on Drosophila ISC activity. Inhibition of Rag GTPases in ECs triggers Mitf activation, which subsequently induces Upd3 expression to non-cell autonomously increase the ISC proliferation and differentiation through stimulating the JAK-STAT pathway. Inactivation of mTORC1 in ECs also activates the Upd3-JAK-STAT axis and increases ISC activity, but through a Mitf-independent mechanism. Moreover, in contrast to mTORC1 inactivation blocking ISC proliferation and differentiation, depletion of Rag GTPases in ISCs shows no obvious cell autonomous effect on ISC activity. Consistent with inhibition of the nutrient-responsive pathway in ECs promoting ISC proliferation, long-term nutrient starvation enhances ISC activity and Upd3 expression. Together, our findings reveal a nutrient-sensing regulatory network that controls ISC proliferation involving the Rag GTPases-Mitf axis and mTORC1 activity.

Grass carp (Ctenopharyngodon idellus) is an important aquaculture species in China that is affected by serious diseases, especially hemorrhagic disease caused by grass carp reovirus (GCRV). Grass carp have previously shown age-dependent susceptibility to GCRV, however, the mechanism by which this occurs remains poorly understood. Therefore, we performed transcriptome and metabolome sequencing on five-month-old (FMO) and three-year-old (TYO) grass carp to identify the potential mechanism. Viral challenge experiments showed that FMO fish were susceptible, whereas TYO fish were resistant to GCRV. RNA-seq showed that the genes involved in immune response, antigen presentation, and phagocytosis were significantly upregulated in TYO fish before the GCRV infection and at the early stage of infection. Metabolome sequencing showed that most metabolites were upregulated in TYO fish and downregulated in FMO fish after virus infection. Intragroup analysis showed that arachidonic acid metabolism was the most significantly upregulated pathway in TYO fish, whereas choline metabolism in cancer and glycerophospholispid metabolism were significantly downregulated in FMO fish after virus infection. Intergroup comparison revealed that metabolites from carbohydrate, amino acid, glycerophospholipid, and nucleotide metabolism were upregulated in TYO fish when compared with FMO fish. Moreover, the significantly differentially expressed metabolites showed antiviral effects both in vivo and in vitro. Based on these results, we concluded that the immune system and host biosynthesis and metabolism, can explain the age-dependent viral susceptibility in grass carp.

Aging and age-related diseases occur in almost all organisms. Recently, it was discovered that the inhibition of target of rapamycin complex 1 (TORC1), a conserved complex that mediates nutrient status and cell metabolism, can extend an individual's lifespan and inhibit age-related diseases in many model organisms. However, the mechanism whereby TORC1 affects aging remains elusive. Here, we use a loss-of-function mutation in nprl2, a component of GATOR1 that mediates amino acid levels and inhibits TORC1 activity, to investigate the effect of increased TORC1 activity on the occurrence of age-related digestive dysfunction in Drosophila. We found that the nprl2 mutation decreased Drosophila lifespan. Furthermore, the nprl2 mutant had a distended crop, with food accumulation at an early age. Interestingly, the inappropriate food distribution and digestion along with decreased crop contraction in nprl2 mutant can be rescued by decreasing TORC1 activity. In addition, nprl2-mutant flies exhibited age-related phenotypes in the midgut, including short gut length, a high rate of intestinal stem cell proliferation, and metabolic dysfunction, which could be rescued by inhibiting TORC1 activity. Our findings showed that the gastrointestinal tract aging process is accelerated in nprl2-mutant flies, owing to high TORC1 activity, which suggested that TORC1 promotes digestive tract senescence.

Mosquitoes act as vectors of numerous pathogens that cause human diseases. Dengue virus (DENV) transmitted by mosquito, Aedes aegypti, is responsible for dengue fever epidemics worldwide with a serious impact on human health. Currently, disease control mainly relies on vector targeted intervention strategies. Therefore, it is imperative to understand the molecular mechanisms underlying the innate immune response of mosquitoes against pathogens. In the present study, the expression profiles of immunity-related genes in the midgut responding to DENV infection by feeding were analyzed by transcriptome and quantitative real-time PCR. The level of Antimicrobial peptides (AMPs) increased seven days post-infection (d.p.i.), which could be induced by the Toll immune pathway. The expression of reactive oxygen species (ROS) genes, including antioxidant genes, such as HPX7, HPX8A, HPX8B, HPX8C were induced at one d.p.i. and peaked again at ten d.p.i. in the midgut. Interestingly, down-regulation of the antioxidant gene HPX8C by RNA interference led to reduction in the virus titer in the mosquito, probably due to the elevated levels of ROS. Application of a ROS inhibitor and scavenger molecules further established the role of oxygen free radicals in the modulation of the immune response to DENV infection. Overall, our comparative transcriptome analyses provide valuable information about the regulation of immunity related genes in the transmission vector in response to DENV infection. It further allows us to identify novel molecular mechanisms underlying the host-virus interaction, which might aid in the development of novel strategies to control mosquito-borne diseases. HPX8C is a heme-containing peroxidase, which can move reactive oxygen species (ROS) damage to the organism by reducing H₂O₂ to H₂O. Previously, the peroxidase gene has been shown to modulate midgut immunity and regulate anti-malarial response in mosquitoes. In this study, the classical immune signaling pathways, Toll and IMD genes might be late responses against the viruses. HPX8C was demonstrated here to play a role in antiviral immunity against DENV infection in Ae. Aegypti mosquitoes. HPX8C expression was induced by DENV infection and continued to increase with an elevated virus titer. In HPX8C-depleted mosquitoes, the ROS level was found to be increased with a corresponding decrease in the DENV and ZIKV virus titer. Therefore, it was speculated that HPX8C mediated immune responses against the DENV in the mosquito in the late stage of viral infection, which could be controlled by Toll pathway.

Parasitoid wasps depend on a variety of maternal virulence factors to ensure successful parasitism. Encapsulation response carried out by host hemocytes is one of the major host immune responses toward limiting endoparasitoid wasp offspring production. We found that VRF1, a metalloprotease homolog venom protein identified from the endoparasitoid wasp, Microplitis mediator, could modulate egg encapsulation in its host, the cotton bollworm, Helicoverpa armigera. Here, we show that the VRF1 proenzyme is cleaved after parasitism, and that the C-terminal fragment containing the catalytic domain enters host hemocytes 6 h post-parasitism. Furthermore, using yeast two-hybrid and pull-down assays, VRF1 is shown to interact with the H. armigera NF-κB factor, Dorsal. We also show that overexpressed of VRF1 in an H. armigera cell line cleaved Dorsal in vivo. Taken together, our results have revealed a novel mechanism by which a component of endoparasitoid wasp venom interferes with the Toll signaling pathway in the host hemocytes.

Serine protease cascade‐mediated melanization is an important innate immune response in insects and crustaceans, which involves the proteolytic activation of prophenoloxidase (PPO). In this study, we investigated the role of Aedes aegypti PPO3 in antifungal immune defence. We expressed and purified recombinant PPO3 (rPPO3) in Escherichia coli and demonstrated that rPPO3 was activated by ethanol and, to a lesser extent, by cetylpyridinium chloride. In the presence of Cu ²⁺ , rPPO3 exhibited enzyme activity. Immunoblot results revealed that the rPPO3 was cleaved by the haemolymph from immune‐challenged mosquitoes or purified Ostrinia furnacalis serine protease 105 in vitro . The cleaved rPPO3 converted dopamine to toxic intermediates that killed fungal conidia of Beauveria bassiana in vitro . In mosquitoes challenged with Be. bassiana , cleavage of rPPO3 produced a 50 kDa phenoloxidase (PO) fragment. Further analysis revealed that the survival rate of mosquitoes with fungal infection increased significantly following injection of rPPO3 into the haemocoel. Taken together, our results suggest that proteolytic cleavage of the mosquito PPO3 plays an important role in the antifungal immune response. This has led to a better understanding of the mechanism of PPO activation in the mosquito and the role of melanization in the antifungal immune response.