Publication: Caspase-dependent alterations of Ca2+ signaling in the induction of apoptosis by hepatitis B virus X protein.

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The hepatitis B virus In its biological effects, the modulation of cellular Ca2+ signals has been proposed to be involved, but the direct assessment of Ca2+ homeostasis in HBx-transfected cells has not been carried out yet. In this work, we have employed for this purpose aequorin-based recombinant probes specifically targeted to intracellular organelles and microdomains. Using these probes, we observed that overexpression of HBx enhanced agonist-evoked cytosolic Ca2+ signals in HepG2 and HeLa cells, without affecting either the steady state of endoplasmic reticulum Ca2+ concentration or the kinetics of Ca2+ release. Rather, caspase-3-dependent cleavage of the plasma membrane Ca2+ ATPase could be demonstrated, and larger rises were detected in the cytoplasmic rim beneath the plasma membrane. In mitochondria, major morphological (fragmentation and swelling) and functional (reduced Ca2+ uptake) alterations were detected in HBx-expressing cells. As to the cellular consequences, we observed that HBx-induced apoptosis was markedly reduced when the alterations in Ca2+ signaling (eg by loading a Ca2+ chelator or preventing PMCA cleavage) or the downstream effects (eg by inhibiting mitochondrial permeability transition) were prevented. Overall, these results indicate that HBx disrupts intracellular Ca2+ homeostasis, acting on the extrusion mechanisms, and that this effect plays an important role in the control of HBx-related apoptosis.