Aquaporins in immunity
(Top) Prip channel localization in hemocytes. Panels indicate the distribution of Prip localization in hemocytes in control media (top) and media containing 10 mM H2O2 (I and J). Membrane localization of Prip in an enlarged image is shown (J)
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(Middle Left) We expressed Prip CDS in HEK293 cells and used the TCFB probe to monitor kinetically the increase in fluorescence in cells where exogenous 100 uM H2O2 was added. Top panels are untransfected control cells, where 100uM H2O2 was added, and an increase in TCFB probe fluorescence was monitored over time. The Merge of Brightfield and DsRed channel is shown.
(Middle Right) There was a significant increase in TCFB fluorescence over time in cells expressing Prip versus the vector control HEK cells at 100 uM exogenous H2O2 addition at the end of 15 mins imaging. The cells in all the panels were loaded with a 5mM TCFB probe prior to the addition of H2O2.
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(Bottom Left and Right) A third instar larvae with GFP positive single showing expression of Prip protein in its renal tubules and hemocytes.
Aquaporins (AQP) are integral membrane proteins that primarily facilitate water transport and small solutes across biological membranes. Beyond their classical role in maintaining cellular and organismal water balance, aquaporins have emerged as pivotal players in immune regulation across diverse organisms, including insects and mammals. Recent studies have highlighted their involvement in processes ranging from cell migration to oxidative stress management, making them critical to the broader understanding of immune responses.
In mammals, AQP3 facilitates hydrogen peroxide (Hâ‚‚Oâ‚‚) transport into macrophages, enabling the generation of signaling molecules that mediate inflammatory responses. Similarly, AQP9 contributes to immune regulation by transporting glycerol and other solutes, which influence leukocyte migration and reactive oxygen species (ROS) production. These functions underscore the broader role of aquaporins in modulating immune responses during infection, inflammation, and tissue repair.
In insects, the roles of aquaporins are less well-characterized but are gaining attention due to their relevance in stress response and immune regulation. For example, aquaporins in mosquitoes are vital for maintaining osmotic balance during blood-feeding and are speculated to aid in oxidative stress regulation under pathogen attack.
In our lab, we use Drosophila melanogaster as a model organism to study this aquaporin-mediated immune function. Our focus is on the Drosophila aquaporin Prip (CG7777), which is expressed in hemocytes (fly blood cells/ immune cells), female ovaries, and renal tubules. Our focus is on hemocytes, as they are central to Drosophila's innate immunity, mediating responses such as phagocytosis, signaling molecule production, and tissue repair.
In the 2020 paper published by Chakrabarti and Visweswariah, Prip protein has been identified as a key player in transporting ROS, such as hydrogen peroxide, into hemocytes. ROS are not only critical antimicrobial agents but also function as signaling molecules that activate immune pathways. Research from our lab has shown that Prip-mediated ROS uptake is essential for activating the IMD and Toll pathways, which govern the defense against pathogens and tissue damage in D. melanogaster.
ROS are central to immune regulation, balancing the dual roles of pathogen elimination and tissue protection. In our studies, we discovered that Prip facilitates the uptake of ROS into hemocytes, where these molecules act as signals to upregulate cytokine production. For instance, the production of cytokine-like molecules such as Upd3 depends on Prip-mediated ROS accumulation. Upd3, in turn, activates the JAK/STAT pathway, regulating intestinal stem cell proliferation and repair following injury.
Our research aims to elucidate the molecular mechanisms by which Prip mediates ROS signaling in hemocytes and how this influences broader immune responses. By focusing on the interplay between aquaporins, ROS, and immune pathways, we hope to uncover novel insights into immune regulation during tissue repair and pathogen attack. Given the evolutionary conservation of aquaporins, these findings may also inform studies in higher organisms, including humans and mice, shedding light on how similar pathways maintain immune homeostasis and protect against disease.
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1. Intramacrophage ROS Primes the Innate Immune System via JAK/STAT and Toll Activation.
S. Chakrabarti*, and S. S. Visweswariah.
Cell Reports 2020. https://doi.org/10.1016/j.celrep.2020.108368 PMID: 33176146​​