Publications (69) View all
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Article: Tyrosine kinase signal modulation: a matter of H2O2 membrane permeability?
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ABSTRACT: 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 H2O2 transport, while a C173A-AQP8 mutant failed to do so. Lowering AQP8 levels affected H2O2 entry into the endoplasmic reticulum, but not into mitochondria. AQP8-silencing also inhibited the H2O2 spikes and phosphorylation of downstream proteins induced by EGF. 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.Antioxidants & Redox Signaling 03/2013; · 8.20 Impact Factor -
Article: Plasma cells require autophagy for sustainable immunoglobulin production.
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ABSTRACT: The role of autophagy in plasma cells is unknown. Here we found notable autophagic activity in both differentiating and long-lived plasma cells and investigated its function through the use of mice with conditional deficiency in the essential autophagic molecule Atg5 in B cells. Atg5(-/-) differentiating plasma cells had a larger endoplasmic reticulum (ER) and more ER stress signaling than did their wild-type counterparts, which led to higher expression of the transcriptional repressor Blimp-1 and immunoglobulins and more antibody secretion. The enhanced immunoglobulin synthesis was associated with less intracellular ATP and more death of mutant plasma cells, which identified an unsuspected autophagy-dependent cytoprotective trade-off between immunoglobulin synthesis and viability. In vivo, mice with conditional deficiency in Atg5 in B cells had defective antibody responses, complete selection in the bone marrow for plasma cells that escaped Atg5 deletion and fewer antigen-specific long-lived bone marrow plasma cells than did wild-type mice, despite having normal germinal center responses. Thus, autophagy is specifically required for plasma cell homeostasis and long-lived humoral immunity.Nature Immunology 01/2013; · 26.01 Impact Factor -
Article: Ero1-PDI interactions, the response to redox flux and the implications for disulfide bond formation in the mammalian endoplasmic reticulum.
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ABSTRACT: The protein folding machinery of the endoplasmic reticulum (ER) ensures that proteins entering the eukaryotic secretory pathway acquire appropriate post-translational modifications and reach a stably folded state. An important component of this protein folding process is the supply of disulfide bonds. These are introduced into client proteins by ER resident oxidoreductases, including ER oxidoreductin 1 (Ero1). Ero1 is usually considered to function in a linear pathway, by 'donating' a disulfide bond to protein disulfide isomerase (PDI) and receiving electrons that are passed on to the terminal electron acceptor molecular oxygen. PDI engages with a range of clients as the direct catalyst of disulfide bond formation, isomerization or reduction. In this paper, we will consider the interactions of Ero1 with PDI family proteins and chaperones, highlighting the effect that redox flux has on Ero1 partnerships. In addition, we will discuss whether higher order protein complexes play a role in Ero1 function.Philosophical Transactions of The Royal Society B Biological Sciences 01/2013; 368(1617):20110403. · 6.40 Impact Factor -
Article: Iron increases the susceptibility of multiple myeloma cells to bortezomib.
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ABSTRACT: Background Multiple myeloma is a malignant still incurable plasma cell disorder. Pharmacological treatment based on proteasome inhibition has improved patient outcome, however bortezomib-resistance remains a major clinical problem. Inhibition of proteasome functionality affects cellular iron homeostasis and iron is a potent inducer of reactive oxygen species and cell death, unless safely stored in ferritin. We explored the potential role of iron in bortezomib-resistance. Design and Methods We analyzed iron proteins, oxidative status and cell viability in 7 multiple myeloma cell lines and in plasma cells from 5 patients. Cells were treated with increasing bortezomib concentrations with or without iron supplementation. We reduced ferritin levels by both shRNA technology and by drug induced iron starvation. Results Multiple myeloma cell lines are characterized by distinct ferritin levels, which directly correlate with bortezomib resistance. We observed that iron supplementation upon bortezomib promotes proteins oxidation and cell death and that iron toxicity inversely correlates with basal ferritin levels. Bortezomib prevents ferritin up-regulation in response to iron, thus limiting the ability to buffer reactive oxygen species. In accordance, reduction of basal ferritin levels increases both bortezomib sensitivity and iron toxicity. In patients cells we confirmed that bortezomib prevents ferritin increase, that iron supplementation upon bortezomib increases cell death and that ferritin reduction overcomes bortezomib resistance. Conclusions Bortezomib affects iron homeostasis sensitizing cells to oxidative damage. Modulation of iron status is a strategy worth to be explored to improve the efficacy of proteasome inhibition therapies.Haematologica 12/2012; · 6.42 Impact Factor -
Article: Pivotal Advance: Protein synthesis modulates responsiveness of differentiating and malignant plasma cells to proteasome inhibitors.
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ABSTRACT: A previously unsuspected, considerable proportion of newly synthesized polypeptides are hydrolyzed rapidly by proteasomes, possibly competing with endogenous substrates and altering proteostasis. In view of the anti-cancer effects of PIs, we set out to achieve a quantitative assessment of proteasome workload in cells hallmarked by different PI sensitivity, namely, a panel of MM cells, and in a dynamic model of plasma cell differentiation, a process that confers exquisite PI sensitivity. Our results suggest that protein synthesis is a key determinant of proteasomal proteolytic burden and PI sensitivity. In different MM cells and in differentiating plasma cells, the average proteolytic work accomplished per proteasome ranges over different orders of magnitude, an unexpected degree of variability, with increased workload invariably associated to increased PI sensitivity. The unfavorable load-versus-capacity balance found in highly PI-sensitive MM lines is accounted for by a decreased total number of immunoproteasomes/cell coupled to enhanced generation of RDPs. Moreover, indicative of cause-effect relationships, attenuating general protein synthesis by the otherwise toxic agent CHX reduces PI sensitivity in activated B and in MM cells. Our data support the view that in plasma cells protein synthesis contributes to determine PI sensitivity by saturating the proteasomal degradative capacity. Quantitating protein synthesis and proteasome workload may thus prove crucial to design novel negative proteostasis regulators against cancer.Journal of leukocyte biology 06/2012; · 4.99 Impact Factor
