Search

Aquaporin-8 (AQP8) allows the bidirectional transport of water and hydrogen peroxide across biological membranes. Depending on its concentration, H2O2 exerts opposite roles, amplifying growth factor signaling in physiological conditions, but causing severe cell damage when in excess. Thus, H2O2 permeability is likely to be tightly controlled in living cells. Aims: In this study, we investigated whether and how the transport of H2O2 through plasma membrane AQP8 is regulated, particularly during cell stress. Results: We show that diverse cellular stress conditions, including heat, hypoxia, and ER stress, reversibly inhibit the permeability of AQP8 to H2O2 and water. Preventing the accumulation of intracellular reactive oxygen species (ROS) during stress counteracts AQP8 blockade. Once inhibition is established, AQP8-dependent transport can be rescued by reducing agents. Neither H2O2 nor water transport is impaired in stressed cells expressing a mutant AQP8, in which cysteine 53 had been replaced by serine. Cells expressing this mutant are more resistant to stress-, drug-, and radiation-induced growth arrest and death. Innovation and Conclusion: The control of AQP8-mediated H2O2 transport provides a novel mechanism to regulate cell signaling and survival during stress.

H2O2 produced by extracellular NADPH oxidases regulates tyrosine kinase signaling inhibiting phosphatases. How does it cross the membrane to reach its cytosolic targets? Silencing aquaporin-8 (AQP8), but not AQP3 or AQP4, inhibited H2O2 entry into HeLa cells. Re-expression of AQP8 with silencing-resistant vectors rescued H 2O2 transport, whereas a C173A-AQP8 mutant failed to do so. Lowering AQP8 levels affected H2O2 entry into the endoplasmic reticum, but not into mitochondria. AQP8 silencing also inhibited the H2O2 spikes and phosphorylation of downstream proteins induced by epidermal growth factor. These observations lead to the hypothesis that H2O2 does not freely diffuse across the plasma membrane and AQP8 and other H2O2 transporters are potential targets for manipulating key signaling pathways in cancer and degenerative diseases. Antioxid. Redox Signal. 19, 1447-1451. © Mary Ann Liebert, Inc.

Cellular metabolism provides various sources of hydrogen peroxide (H₂O₂) in different organelles and compartments. The suitability of H₂O₂ as an intracellular signaling molecule therefore also depends on its ability to pass cellular membranes. The propensity of the membranous boundary of the endoplasmic reticulum (ER) to let pass H₂O₂ has been discussed controversially. In this essay, we challenge the recent proposal that the ER membrane constitutes a simple barrier for H₂O₂ diffusion and support earlier data showing that (i) ample H₂O₂ permeability of the ER membrane is a prerequisite for signal transduction, (ii) aquaporin channels are crucially involved in the facilitation of H₂O₂ permeation, and (iii) a proper experimental framework not prone to artifacts is necessary to further unravel the role of H₂O₂ permeation in signal transduction and organelle biology.

Limited amounts of reactive oxygen species are necessary for cell survival and signaling, but their excess causes oxidative stress. H2O 2 and other reactive oxygen species are formed as byproducts of several metabolic pathways, possibly including oxidative protein folding in the endoplasmic reticulum. B-to plasma-cell differentiation is characterized by a massive expansion of the endoplasmic reticulum, finalized to sustain abundant immunoglobulin (Ig) synthesis and secretion. The increased production of disulfide-rich Ig might cause oxidative stress that could serve signaling roles in the differentiation and lifespan control of antibody-secreting cells. Here we show that terminal B-cell differentiation entails redox stress, NF-E2-related factor-2 (Nrf2) activation, and reshaping of the antioxidant responses. However, plasma-cell differentiation was not dramatically impaired in peroxiredoxin (Prx)1-, 2-, 3-, and 4-, glutathione peroxidase 1-, and Nrf2-knockout splenocytes, suggesting redundancy and robustness in antioxidant systems. Endoplasmic reticulum (ER)-resident Prx4 increases dramatically during differentiation. In its absence, IgM secretion was not significantly affected, but more high-molecular-weight covalent complexes accumulated intracellularly. Our results suggest that the early intracellular production of H 2O2 facilitates B-cell proliferation and reveal a role for the Nrf2 pathway in the differentiation and function of IgM-secreting cells. © 2010, Mary Ann Liebert, Inc.

Upon antigen stimulation, B lymphocytes differentiate into antibody secreting cells (ASC), most of which undergo apoptosis after a few days of intense Ig production. Differentiation entails expansion of the endoplasmic reticulum (ER) and requires XBP1 but not other elements of the unfolded protein response, like PERK. Moreover, normal and malignant ASC are exquisitely sensitive to proteasome inhibitors, but the underlying mechanisms are poorly understood. Here we analyze the role of C/EBP homologous protein (CHOP), a transcription factor mediating apoptosis in many cell types that experience high levels of ER stress. CHOP is transiently induced early upon B cell stimulation: covalent IgM aggregates form more readily and IgM secretion is slower in chop-/- cells. Despite these subtle changes, ASC differentiation and lifespan are normal in chop-/- mice. Unlike fibroblasts and other cell types, chop-/- ASC are equally or slightly more sensitive to proteasome inhibitors and ER stressors, implying tissue-specific roles for CHOP in differentiation and stress. © 2009 Elsevier Ltd. All rights reserved.

When small B lymphocytes bind their cognate antigens in the context of suitable signals, a dramatic differentiation program is activated that leads to the formation of plasma cells. These are short-lived specialized elements, each capable of secreting several thousands antibodies per second. The massive increase in Ig synthesis and transport entails a dramatic architectural and functional metamorphosis that involves the development of the endoplasmic reticulum (ER) and secretory organelles. Massive Ig secretion poses novel metabolic requirements, particularly for what concerns aminoacid import, ATP synthesis and redox homeostasis. Ig H and L chains enter the ER in the reduced state, to be rapidly oxidised mainly via protein driven relays based on the resident enzymes PDI and Ero1. How do plasma cells cope with the ensuing metabolic and redox stresses? In this essay, we discuss the physiological implications that increased Ig production could have in the control of plasma cell generation, function and lifespan, with emphasis on the potential role of ROS generation in mitochondria and ER. © 2008 Elsevier B.V. All rights reserved.

After few days of intense immunoglobulin (Ig) secretion, most plasma cells undergo apoptosis, thus ending the humoral immune response. We asked whether intrinsic factors link plasma cell lifespan to Ig secretion. Here we show that in the late phases of plasmacytic differentiation, when antibody production becomes maximal, proteasomal activity decreases. The excessive load for the reduced proteolytic capacity correlates with accumulation of poly-ubiquitinated proteins, stabilization of endogenous proteasomal substrates (including Xbp1s, IκBα, and Bax), onset of apoptosis, and sensitization to proteasome inhibitors (PI). These events can be reproduced by expressing Ig-μ chain in nonlymphoid cells. Our results suggest that a developmental program links plasma cell death to protein production, and help explaining the peculiar sensitivity of normal and malignant plasma cells to PI. ©2006 European Molecular Biology Organization.