Cellular swelling triggers the activation of Cl؊ channels (volume-sensitive outwardly rectifying (VSOR) Cl ؊ channels) in many cell types. Ensuing regulatory volume decrease has been considered the primary function of these channels. However, Cl ؊ channels, which share functional properties with volume-sensitive Cl ؊ channels, have been shown to be involved in other physiological processes, including cell proliferation and apoptosis, raising the question of their physiological roles and the signal transduction pathways involved in their activation. Here we report that exogenously applied H 2 O 2 elicited VSOR Cl ؊ channel activation. Furthermore, activation of these channels was found to be coupled to NAD(P)H oxidase activity. Also, epidermal growth factor, known to increase H 2 O 2 production, activated Cl ؊ channels with properties identical to swelling-sensitive Cl ؊ channels. It is concluded that NAD(P)H oxidasederived H 2 O 2 is the common signal transducing molecule that mediates the activation of these ubiquitously expressed anion channels under a variety of physiological conditions.

Transmembrane BAX inhibitor motif-containing (TMBIM)-6, also known as BAX-inhibitor 1 (BI-1), is an anti-apoptotic protein that belongs to a putative family of highly conserved and poorly characterized genes. Here we report the function of TMBIM3/GRINA in the control of cell death by endoplasmic reticulum (ER) stress. Tmbim3 mRNA levels are strongly upregulated in cellular and animal models of ER stress, controlled by the PERK signaling branch of the unfolded protein response. TMBIM3/GRINA synergies with TMBIM6/BI-1 in the modulation of ER calcium homeostasis and apoptosis, associated with physical interactions with inositol trisphosphate receptors. Loss-of-function studies in D. melanogaster demonstrated that TMBIM3/GRINA and TMBIM6/BI-1 have synergistic activities against ER stress in vivo. Similarly, manipulation of TMBIM3/GRINA levels in zebrafish embryos revealed an essential role in the control of apoptosis during neuronal development and in experimental models of ER stress. These findings suggest the existence of a conserved group of functionally related cell death regulators across species beyond the BCL-2 family of proteins operating at the ER membrane. Cell Death and Differentiation (2012) 19, 1013-1026 doi:10.1038/cdd.2011; published online 13 January 2012Apoptosis is a conserved cell death mechanism essential for normal development and tissue homeostasis in multicellular organisms. In mammals, the BCL-2 family of proteins is a group of crucial upstream regulators of the caspase cascade, comprising both pro-and anti-apoptotic components. 1Although apoptosis is observed in most multicellular organisms, the BCL-2 family of proteins as a whole is poorly conserved in invertebrates including worms, flies, and other species.1,2 In fact, only two BCL-2 homologues are present in flies with controversial roles in programmed cell death. 3,4 A pioneering screening to identify human genes that prevents BAX toxicity in a yeast assay identified transmembrane BAX inhibitor motif-containing (TMBIM)-6, also known as BAX inhibitor-1 (BI-1), as a new mammalian gene that negatively regulates apoptosis (reviewed in Robinson KS et al. and Reimers K et al. 5,6 ). Further studies demonstrated that TMBIM6/BI-1 is a six transmembrane-spanning protein, located at the endoplasmic reticulum (ER) that has a relevant role in preventing apoptosis. 5 Remarkably, bioinformatic analysis defined a putative family of at least six highly conserved orthologs of TMBIM6/BI-1containing the consensus motif UPF0005 with unknown function, a domain encoding for six to seven transmembrane-spanning regions. 7TMBIM family of proteins includes the founder member TMBIM6/BI-1, TMBIM1/RECS1 (responsive to centrifugal force and shear stress gene 1 protein), TMBIM2/LFG (life guard), TMBIM3/GRINA (glutamate receptor ionotropic NMDA protein 1), TMBIM4/GAAP (Golgi anti-apoptoticassociated protein), and TMBIM5/GHTIM (growth hormoneinducible transmembrane protein). The TMBIM family of proteins is highly conserved in mammals, zebrafish, and flies, with homol...

Increased expression of the TRPM4 channel has been reported to be associated with the progression of prostate cancer. However, the molecular mechanism underlying its effect remains unknown. This work found that decreasing TRPM4 levels leads to the reduced proliferation of PC3 cells. This effect was associated with a decrease in total β‐catenin protein levels and its nuclear localization, and a significant reduction in Tcf/Lef transcriptional activity. Moreover, TRPM4 silencing increases the Ser33/Ser37/Thr41 β‐catenin phosphorylated population and reduces the phosphorylation of GSK‐3β at Ser9, suggesting an increase in β‐catenin degradation as the underlying mechanism. Conversely, TRPM4 overexpression in LNCaP cells increases the Ser9 inhibitory phosphorylation of GSK‐3β and the total levels of β‐catenin and its nonphosphorylated form. Finally, PC3 cells with reduced levels of TRPM4 showed a decrease in basal and stimulated phosphoactivation of Akt1, which is likely responsible for the decrease in GSK‐3β activity in these cells. Our results also suggest that the effect of TRPM4 on Akt1 is probably mediated by an alteration in the calcium/calmodulin‐EGFR axis, linking TRPM4 activity with the observed effects in β‐catenin‐related signaling pathways. These results suggest a role for TRPM4 channels in β‐catenin oncogene signaling and underlying mechanisms, highlighting this ion channel as a new potential target for future therapies in prostate cancer.

Organic osmolyte and halide permeability pathways activated in epithelial HeLa cells by cell swelling were studied by radiotracer efflux techniques and single-cell volume measurements. The replacement of extracellular Cl− by anions that are more permeant through the volume-activated Cl− channel, as indicated by electrophysiological measurements, significantly decreased taurine efflux. In the presence of less-permeant anions, an increase in taurine efflux was observed. Simultaneous measurement of the125I, used as a tracer for Cl−, and [3H]taurine efflux showed that the time courses for the two effluxes differed. In Cl−-rich medium the increase in I− efflux was transient, whereas that for taurine was sustained. Osmosensitive Cl− conductance, assessed by measuring changes in cell volume, increased rapidly after hypotonic shock. The influx of taurine was able to counteract Cl− conductance-dependent cell shrinkage but only ∼4 min after triggering cell swelling. This taurine-induced effect was blocked by DIDS. Differences in anion sensitivity, the time course of activation, and sensitivity to DIDS suggest that the main cell swelling-activated permeability pathways for taurine and Cl− are separate.

Cell volume regulation is one of the most fundamental homeostatic mechanisms and essential for normal cellular function. At the same time, however, many physiological mechanisms are associated with regulatory changes in cell size meaning that the set point for cell volume regulation is under physiological control. Thus, cell volume is under a tight and dynamic control and abnormal cell volume regulation will ultimately lead to severe cellular dysfunction, including alterations in cell proliferation and cell death. This review describes the different swelling-activated ion channels that participate as key players in the maintenance of normal steady-state cell volume, with particular emphasis on the intracellular signalling pathways responsible for their regulation during hypotonic stress, cell proliferation and apoptosis.