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...

Disturbance of endoplasmic reticulum (ER) proteostasis is a common feature of amyotrophic lateral sclerosis (ALS). Protein disulfide isomerases (PDIs) are ER foldases identified as possible ALS biomarkers, as well as neuroprotective factors. However, no functional studies have addressed their impact on the disease process. Here, we functionally characterized four ALS-linked mutations recently identified in two major PDI genes, PDIA1 and PDIA3/ERp57. Phenotypic screening in zebrafish revealed that the expression of these PDI variants induce motor defects associated with a disruption of motoneuron connectivity. Similarly, the expression of mutant PDIs impaired dendritic outgrowth in motoneuron cell culture models. Cellular and biochemical studies identified distinct molecular defects underlying the pathogenicity of these PDI mutants. Finally, targeting ERp57 in the nervous system led to severe motor dysfunction in mice associated with a loss of neuromuscular synapses. This study identifies ER proteostasis imbalance as a risk factor for ALS, driving initial stages of the disease.

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.

Adaptation to endoplasmic reticulum (ER) stress depends on the activation of the unfolded protein response (UPR) stress sensor inositol-requiring enzyme 1a (IRE1a), which functions as an endoribonuclease that splices the mRNA of the transcription factor XBP-1 (X-box-binding protein-1). Through a global proteomic approach we identified the BCL-2 family member PUMA as a novel IRE1a interactor. Immunoprecipitation experiments confirmed this interaction and further detected the association of IRE1a with BIM, another BH3-only protein. BIM and PUMA doubleknockout cells failed to maintain sustained XBP-1 mRNA splicing after prolonged ER stress, resulting in early inactivation. Mutation in the BH3 domain of BIM abrogated the physical interaction with IRE1a, inhibiting its effects on XBP-1 mRNA splicing. Unexpectedly, this regulation required BCL-2 and was antagonized by BAD or the BH3 domain mimetic ABT-737. The modulation of IRE1a RNAse activity by BH3-only proteins was recapitulated in a cell-free system suggesting a direct regulation. Moreover, BH3-only proteins controlled XBP-1 mRNA splicing in vivo and affected the ER stress-regulated secretion of antibodies by primary B cells. We conclude that a subset of BCL-2 family members participates in a new UPR-regulatory network, thus assuming apoptosis-unrelated functions.

TRPM4 is critically involved in LPS-induced endothelial cell death. These results demonstrate that either pharmacological inhibition of TRPM4, suppression of TRPM4 expression, or inhibition of TRPM4 activity are able to protect endothelium against LPS injury. These results are useful in sepsis drug design and development of new strategies for sepsis therapy.