Bax Inhibitor-1 Research Articles

EPO regulates neuroprotective Transmembrane BAX Inhibitor-1 Motif-containing (TMBIM) family members GRINA and FAIM2 after cerebral ischemia-reperfusion injury

Background and purposeTransmembrane BAX Inhibitor-1 Motif-containing (TMBIM) family members exert inhibitory activities in apoptosis and necroptosis. FAIM2 (TMBIM-2) is neuroprotective against murine focal ischemia and is regulated by erythropoietin (EPO). Similar to FAIM2, GRINA (TMBIM-3) is predominantly expressed in the brain. The role of GRINA in transient brain ischemia, its potential synergistic effects with FAIM2 and its regulation by EPO treatment were assessed. MethodsWe performed transient (30 min) middle cerebral artery occlusion (tMCAo) followed by 72 h of reperfusion in GRINA-deficient (GRINA−/−), FAIM2-deficient (FAIM2−/−), double-deficient (GRINA−/-FAIM2−/−) and wildtype littermates (WT) mice. We administered EPO or saline 0, 24 and 48 h after tMCAo. We subjected primary murine cortical neurons (pMCN) of all mouse strains to oxygen–glucose deprivation (OGD) after GRINA and/or FAIM2 gene transfection. ResultsCompared to wildtype controls GRINA deficiency led to a similar increase in infarct volumes as FAIM2 deficiency (p < .01). We observed the highest neurological deficits and largest infarct sizes in double-deficient mice. EPO administration upregulated GRINA and FAIM2 mRNA levels in wildtype littermates. EPO decreased infarct sizes and abrogated neurological impairments in wildtype controls. GRINA and/or FAIM2 deficient mice showed increased expression levels of cleaved-caspase 3 and of pro-apoptotic BAX mRNA. Further, caspase 8 was upregulated in FAIM2−/− and caspase 9 in GRINA−/− mice. Overexpression of GRINA and FAIM2 in wildtype and in double deficient pMCN decreased cell death rate after OGD. ConclusionsGRINA and FAIM2 are highly expressed in the brain and convey EPO-mediated neuroprotection after ischemic stroke involving different caspases.

The characteristics of the ancient cell death suppressor, TMBIM6, and its related signaling pathways after endoplasmic reticulum stress

Activation of the unfolded protein response in combination with generation of reactive oxygen species, from cytochrome P450 members and NADPH-P450 reductases, are two major consequences of Endoplasmic Reticulum (ER) stress that cause oxidative damage and cell death. Herein, we reviewed the role of Bax Inhibitor-1 (BI-1), an evolutionarily conserved protein encoded by the Transmembrane Bax inhibitor Motif Containing 6 gene, in protection from ER stress. As BI-1 has multimodal properties that can target a wide array of pathophysiological consequences after injury, our main objective was to explore BI-1's protective role in ER stress and its potential signaling pathways.

The Bax inhibitor UvBI-1, a negative regulator of mycelial growth and conidiation, mediates stress response and is critical for pathogenicity of the rice false smut fungus Ustilaginoidea virens

Bax inhibitor-1 (BI-1), an evolutionarily conserved protein, is a suppressor of cell death induced by the proapoptotic protein Bax and is involved in the response to biotic and abiotic stress in animals, plants and yeast. Rice false smut caused by Ustilaginoidea virens is one of the destructive rice diseases worldwide. Although BI-1 proteins are widely distributed across filamentous fungi, few of them are functionally characterized. In this study, we identified a BI-1 protein in U. virens, UvBI-1, which contains a predicted Bax inhibitor-1-like family domain and could suppress the cell death induced by Bax. By co-transformation of the CRISPR/Cas9 construct along with donor DNA fragment containing the hygromycin resistance gene, we successfully generated Uvbi-1 deletion mutants. The UvBI-1 deletion showed an increase in mycelia vegetative growth and conidiation, suggesting this gene acts as a negative regulator of the growth and conidiation. In addition, the Uvbi-1 mutants exhibited higher sensitivity to osmotic and salt stress, hydrogen peroxide stress, and cell wall or membrane stress than the wild-type strain. Furthermore, UvBI-1 deletion was found to cause increased production of secondary metabolites and loss of pathogenicity of U. virens. Taken together, our results demonstrate that UvBI-1 plays a negative role in mycelial growth and conidiation, and is critical for stress tolerance, cell wall integrity, secondary metabolites production and pathogenicity of U. virens. Therefore, this study provides new evidence on the conserved function of BI-1 among fungal organisms and other species.

Transcriptome and Proteome Response of Rhipicephalus annulatus Tick Vector to Babesia bigemina Infection.

A system biology approach was used to gain insight into tick biology and interactions between vector and pathogen. Rhipicephalus annulatus is one of the main vectors of Babesia bigemina which has a massive impact on animal health. It is vital to obtain more information about this relationship, to better understand tick and pathogen biology, pathogen transmission dynamics, and new potential control approaches. In ticks, salivary glands (SGs) play a key role during pathogen infection and transmission. RNA sequencing obtained from uninfected and B. bigemina infected SGs obtained from fed female ticks resulted in 6823 and 6475 unigenes, respectively. From these, 360 unigenes were found to be differentially expressed (p < 0.05). Reversed phase liquid chromatography–mass spectrometry identified a total of 3679 tick proteins. Among them 406 were differently represented in response to Babesia infection. The omics data obtained suggested that Babesia infection lead to a reduction in the levels of mRNA and proteins (n = 237 transcripts, n = 212 proteins) when compared to uninfected controls. Integrated transcriptomics and proteomics datasets suggested a key role for stress response and apoptosis pathways in response to infection. Thus, six genes coding for GP80, death-associated protein kinase (DAPK-1), bax inhibitor-1 related (BI-1), heat shock protein (HSP), heat shock transcription factor (PHSTF), and queuine trna-ribosyltransferase (QtRibosyl) were selected and RNA interference (RNAi) performed. Gene silencing was obtained for all genes except phstf. Knockdown of gp80, dapk-1, and bi-1 led to a significant increase in Babesia infection levels while hsp and QtRibosyl knockdown resulted in a non-significant decrease of infection levels when compared to the respective controls. Gene knockdown did not affect tick survival, but engorged female weight and egg production were affected in the gp80, dapk-1, and QtRibosyl-silenced groups in comparison to controls. These results advanced our understanding of tick–Babesia molecular interactions, and suggested new tick antigens as putative targets for vaccination to control tick infestations and pathogen infection/transmission.

Death of female flower microsporocytes progresses independently of meiosis-like process and can be accelerated by specific transcripts in Asparagus officinalis

Asparagus officinalis (garden asparagus) is a dioecious perennial crop, and the dioecy (i.e., sex) of A. officinalis can affect its productivity. In A. officinalis, flower anthers in female plants fail to accumulate callose around microsporocytes, fail to complete meiosis, and degenerate due to cell death. Although 13 genes have been implicated in the anther development of male and female flowers, it is unclear how these genes regulate the cell death in female flower anthers. The aim of this study was to narrow down factors involved in this process. TUNEL staining and Feulgen staining of female flower microsporocytes suggest that female microsporocytes enter a previously undetected meiosis-like process, and that the cell death occurs independently of this meiosis-like process, excluding the possibility that the cell death is caused by the cessation of meiosis. RNA sequencing with individual floral organs (tepals, pistils and stamens) revealed that several genes possibly regulating the cell death, such as metacaspase genes and a Bax inhibitor-1 gene, are differentially regulated between female and male flower anthers, and that genes involved in callose accumulation are up-regulated only in male flower anthers. These genes are likely involved in regulating the cell death in female flower anthers in A. officinalis.

Dissolved organic matter derived from rape straw pretreated with selenium in soil improves the inhibition of Sclerotinia sclerotiorum growth.

Sclerotinia sclerotiorum (S. sclerotiorum) is a soil-borne pathogen with broad host range. Dissolved organic matter (DOM) plays a vital role in regulating microbial activity in soil. Exogenous selenium (Se) inhibits plant pathogen growth and enhances the capacity of plants to resist disease. DOM from rape straw with Se treated in soil (RSDOMSe) was extracted, and the inhibitory effect on S. sclerotiorum growth was investigated. RSDOMSe inhibited S. sclerotiorum growth, which not only caused severe damage to S. sclerotiorum hyphae but also enhanced soluble protein leakage, thereby improving the growth inhibition ratio by 20.9%. As the action in intercellular, RSDOMSe led to a significant increase in oxalic acid and decrease in CWDE (cell wall-degrading enzyme, which helps pathogens to invade plants) activities, downregulation of Bi1 (BAX inhibitor-1, required for S. sclerotiorum virulence), Ggt1 (γ-glutamyl transpeptidase, regulates the ROS antioxidant system), CWDE2 and CWDE10 gene expression levels, compared with non-Se treated RSDOM (RSDOMN). Eight metabolites upregulated in RSDOMSe were identified by GC-TOF-MS, and among these metabolites, fumaric acid, maleic acid, malonic acid, mucic acid, saccharic acid, succunic acid and phenylacetic acid showed significant inhibition on S. sclerotiorum growth. These findings provide valuable insight into a new approach for developing eco-friendly fungicides.

Bax inhibitor-1 overexpression in prelimbic cortex protects rats against depression-like behavior induced by olfactory bulbectomy and reduces apoptotic and inflammatory signals

ABSTRACTBackground and purpose: Depression is a mental disorder characterized by a pervasive low mood and loss of pleasure or interest in usual activities, and often results in the impairment of learning and memory. Bax inhibitor-1 (BI-1) has been reported to be involved in the pathological mechanisms for neurodegenerative disorders including depression. Here, we aimed to investigate the role of BI-1 in regulating depression-like behavior induced by olfactory bulbectomy (OB) in rats and the possible mechanism.Methods: Adeno-associated virus vectors expressing BI-1 (AAV-BI-1) were bilaterally microinjected into the prelimbic cortex (PFC-PL) to establish a BI-1 overexpression model in the PFC-PL of rats. TUNEL staining was used to evaluate the cellular apoptosis rate in the PFC-PL. Western blot analysis was performed to examine the expressions of apoptotic and inflammatory signals.Results: BI-1 overexpression significantly attenuated the OB-induced behavioral abnormalities, including the decreased hyperactivity in the open field, decreased immobility time in the forced swimming test, as well as the increased sucrose consumption. BI-1 overexpression significantly inhibited cellular apoptosis in the PFC-PL of OB rats. The expressions of tumor necrosis factor (TNF)-α, interleukin (IL)-6, B-cell lymphoma (Bcl)-2 associated X protein (Bax), and caspase-3 in the PFC-PL of OB rats were significantly increased as compared with the sham rats, but the Bcl-2 and IL-10 expressions were decreased, whereas BI-1 overexpression significantly suppressed the changes of these proteins in the PFC-PL of OB rats.Conclusion: These results indicated that BI-1 may play an anti-depression function with concurrent regulation of apoptotic and inflammatory signals.

Abstract TP322: Viral Mediated Gene Delivery of TMBIM6 Protects the Neonatal Brain

Introduction: A wide variety of conditions may contribute to oxidative protein folding processes in the endoplasmic reticulum (ER) which cause ER stress and play a major role in the pathogenesis of neonatal hypoxic ischemic (HI) injury. ER stress triggers the unfolded protein response (UPR) resulting in the activation of pro-apoptotic signaling causing cells to undergo apoptosis. Here were explored the role of Bax Inhibitor-1 (BI-1) protein, an evolutionary protein encoded by the Transmembrane Bax inhibitor Motif Containing 6 (TMBIM6) gene, as a novel modulator of ER stress induced apoptosis after HI brain injury in a neonatal rat pup. BI-1 may modulate the UPR response via a direct physical interaction with the stress sensor receptor, IRE1α, and thus inhibiting it’s pro-apoptotic signaling. Hypothesis: Overexpression of BI-1, using viral-mediated gene delivery of human adenoviral-TMBIM6 (Ad-TMBIM6) vector, will protect the neonatal brain from HI by inhibiting ER stress induced apoptosis. Methods: Ten-day old (P10) unsexed Sprague-Dawley rat pups underwent right common carotid artery ligation, followed by 1.5h of hypoxia. To overexpress BI-1, rat pups were intracerebroventricularly (icv) injected at 48h pre-HI with the human adenoviral vector-TMBIM6 (Ad-TMBIM6). To determine BI-1 confers protection via inhibition of IRE1α, we administered BI-1 siRNA, to silence BI-1, and an IRE1α CRISPR activation plasmid (AP), to counteract BI-1’s effects, at 48h pre-HI. Percent infarct area, immunofluorescent staining and western blot were performed at 72h post HI. Results: Overexpression of BI-1 showed to significantly reduce percent infarcted area, attenuated neuronal degeneration, reduced ER stress induced neuronal apoptosis and improved long-term neurological outcomes. Either silencing BI-1 or over activating IRE1α with a CRISPR AP significantly reversed BI-1’s protective effects. Conclusions: Overexpression of BI-1 attenuated the morphological and neurological consequences post neonatal HI via inhibition of the ER stress sensor, IRE1α. This study showed a novel role for BI-1 and ER stress in the pathophysiology of HI and provided a basis for BI-1 as a potential therapeutic target.

Identification of mutation in TMB1M6 gene in response to heat stress in goats

ABSTRACT BAX inhibitor-1 (BI-1) gene, known as TMB1M6, is evolutionary conserved and regulates cell survival in response to various stress in animals and plants. The present study was carried out to analyse the TMB1M6 gene in relation to stress susceptible and tolerant phenotypes in different goat breeds by high resolution melting (HRM). Phenotyping of individuals were carried out basing on heart rate (HR) and respiration rate (RR) during peak heat stress period and recognized as heat stress-tolerant (HST) and heat stress-susceptible (HSS). Genotyping of TMB1M6 gene was carried out by HRM. HRM indicated four different groups in the analysed samples. Two novel mutations were observed in heat stress-tolerant and stress-susceptible individuals in analysed samples. The nucleotide polymorphisms were identified at base position 55(T>C) in heat stress-tolerant (HST) individual in Barbari goat. Similarly the mutation at base position 96 was observed in heat stress-susceptible (HSS) individual in Jakhrana goat. The nucleotide sequence analysis indicated that the percent similarity of TMBIM 6 gene fragment of goat with cattle, human and mouse was 92.2, 85.2 and 80.9%, respectively. Two SNPs were identified in heat stress susceptible and heat stress tolerant phenotype in goat.

Adenoviral TMBIM6 vector attenuates ER-stress-induced apoptosis in a neonatal hypoxic-ischemic rat model.

ABSTRACTEndoplasmic reticulum (ER) stress is a major pathology encountered after hypoxic-ischemic (HI) injury. Accumulation of unfolded proteins triggers the unfolded protein response (UPR), resulting in the activation of pro-apoptotic cascades that lead to cell death. Here, we identified Bax inhibitor 1 (BI-1), an evolutionarily conserved protein encoded by the transmembrane BAX inhibitor motif-containing 6 (TMBIM6) gene, as a novel modulator of ER-stress-induced apoptosis after HI brain injury in a neonatal rat pup. The main objective of our study was to overexpress BI-1, via viral-mediated gene delivery of human adenoviral-TMBIM6 (Ad-TMBIM6) vector, to investigate its anti-apoptotic effects as well as to elucidate its signaling pathways in an in vivo neonatal HI rat model and in vitro oxygen-glucose deprivation (OGD) model. Ten-day-old unsexed Sprague Dawley rat pups underwent right common carotid artery ligation followed by 1.5 h of hypoxia. Rat pups injected with Ad-TMBIM6 vector, 48 h pre-HI, showed a reduction in relative infarcted area size, attenuated neuronal degeneration and improved long-term neurological outcomes. Furthermore, silencing of BI-1 or further activating the IRE1α branch of the UPR, using a CRISPR activation plasmid, was shown to reverse the protective effects of BI-1. Based on our in vivo and in vitro data, the protective effects of BI-1 are mediated via inhibition of IRE1α signaling and in part via inhibition of the second stress sensor receptor, PERK. Overall, this study showed a novel role for BI-1 and ER stress in the pathophysiology of HI and could provide a basis for BI-1 as a potential therapeutic target.

Arabidopsis Bax inhibitor-1 interacts with enzymes related to very-long-chain fatty acid synthesis.

Bax inhibitor-1 (BI-1) is a widely conserved cell death regulator that confers resistance to environmental stress in plants. Previous studies suggest that Arabidopsis thaliana BI-1 (AtBI-1) modifies sphingolipids by interacting with cytochrome b5 (AtCb5), an electron-transfer protein. To reveal how AtBI-1 regulates sphingolipid synthesis, we screened yeast sphingolipid-deficient mutants and identified yeast ELO2 and ELO3 as novel enzymes that are essential for AtBI-1 function. ELO2 and ELO3 are condensing enzymes that synthesize very-long-chain fatty acids (VLCFAs), major fatty acids in plant sphingolipids. In Arabidopsis, we identified four ELO homologs (AtELO1-AtELO4), localized in the endoplasmic reticulum membrane. Of those AtELOs, AtELO1 and AtELO2 had a characteristic histidine motif and were bound to AtCb5-B. This result suggests that AtBI-1 interacts with AtELO1 and AtELO2 through AtCb5. AtELO2 and AtCb5-B also interact with KCR1, PAS2, and CER10, which are essential for the synthesis of VLCFAs. Therefore, AtELO2 may participate in VLCFA synthesis with AtCb5 in Arabidopsis. In addition, our co-immunoprecipitation/mass spectrometry analysis demonstrated that AtBI-1 forms a complex with AtELO2, KCR1, PAS2, CER10, and AtCb5-D. Furthermore, AtBI-1 contributes to the rapid synthesis of 2-hydroxylated VLCFAs in response to oxidative stress. These results indicate that AtBI-1 regulates VLCFA synthesis by interacting with VLCFA-synthesizing enzymes.

Bax inhibitor 1 is a γ-secretase–independent presenilin-binding protein

Presenilin is the catalytic subunit of γ-secretase, a four-component intramembrane protease responsible for the generation of β-amyloid (Aβ) peptides. Over 200 Alzheimer's disease-related mutations have been identified in presenilin 1 (PS1) and PS2. Here, we report that Bax-inhibitor 1 (BI1), an evolutionarily conserved transmembrane protein, stably associates with PS1. BI1 specifically interacts with PS1 in isolation, but not with PS1 in the context of an assembled γ-secretase. The PS1-BI1 complex exhibits no apparent proteolytic activity, as judged by the inability to produce Aβ40 and Aβ42 from the substrate APP-C99. At an equimolar concentration, BI1 has no impact on the proteolytic activity of γ-secretase; at a 200-fold molar excess, BI1 reduces γ-secretase activity nearly by half. Our biochemical study identified BI1 as a PS1-interacting protein, suggesting additional functions of PS1 beyond its involvement in γ-secretase.

Human cytomegalovirus US21 protein is a viroporin that modulates calcium homeostasis and protects cells against apoptosis

The human cytomegalovirus (HCMV) US12 gene family comprises a set of 10 contiguous genes (US12 to US21) with emerging roles in the regulation of virus cell tropism, virion composition, and immunoevasion. Of all of the US12 gene products, pUS21 shows the highest level of identity with two cellular transmembrane BAX inhibitor motif-containing (TMBIM) proteins: Bax inhibitor-1 and Golgi anti-apoptotic protein, both of which are involved in the regulation of cellular Ca2+ homeostasis and adaptive cell responses to stress conditions. Here, we report the US21 protein to be a viral-encoded ion channel that regulates intracellular Ca2+ homeostasis and protects cells against apoptosis. Indeed, we show pUS21 to be a 7TMD protein expressed with late kinetics that accumulates in ER-derived vesicles. Deletion or inactivation of the US21 gene resulted in reduced HCMV growth, even in fibroblasts, due to reduced gene expression. Ratiometric fluorescence imaging assays revealed that expression of pUS21 reduces the Ca2+ content of intracellular ER stores. An increase in cell resistance to intrinsic apoptosis was then observed as an important cytobiological consequence of the pUS21-mediated alteration of intracellular Ca2+ homeostasis. Moreover, a single point mutation in the putative pore of pUS21 impaired the reduction of ER Ca2+ concentration and attenuated the antiapoptotic activity of pUS21wt, supporting a functional link with its ability to manipulate Ca2+ homeostasis. Together, these results suggest pUS21 of HCMV constitutes a TMBIM-derived viroporin that may contribute to HCMV's overall strategy to counteract apoptosis in infected cells.

Comparative Studies on the Role of Organic Biostimulant in Resistant and Susceptible Cultivars of Rice Grown under Saline Stress - Organic Biostimulant Alleviate Saline Stress in Tolerant and Susceptible Cultivars of Rice

Plants are sessile organisms that experience various abiotic stresses during their lifespan and try to adapt to these environmental stresses by manipulating their physiological, biochemical, cellular, and molecular mechanisms. Salinity is one of the important abiotic stress that affects the metabolism and physiology of plant cells that leads to serious damage to crops and productivity. We investigated the response of two contrasting (salt susceptible and tolerant) cultivars during saline stress by modulating its effect with the application of an important natural biostimulant panchagavya (PG). The results showed that the salinity stress greatly influenced and negatively affects the plant growth, biochemical attributes, and induces the expression of various genes in both cultivars. Furthermore, we assessed the effect of PG alone and by amending with NaCl to alleviate the saline stress which showed a significant enhancement of biochemical and physiological characteristics in both cultivars. Furthermore, we assessed the response of seven autophagy associated gene (ATG1, ATG3, ATG4, ATG6, ATG7, ATG8, and ATG9), BAX Inhibitor -1 (BI-1), Mitogen activated Protein Kinase–1 (MAPK-1), WRKY53, Catalase -1 (CAT-1), Superoxide Dismutase (SOD), and Glutathione Peroxidase (GPX) genes in rice that displayed the differential expression pattern during saline stress in both cultivars. We concluded that saline stress can be manipulated by the application of PG and positively regulate the physiological, biochemical, and gene expression response in salt-susceptible and -tolerant rice cultivars. Furthermore, the current study also suggested that salinity is a mutifactorial and multigenic response. Autophagy and programmed cell death regulated along with salinity and was helpful in adapting the tolerance against the stress condition.

Transcriptomics of Epichloë-Grass Symbioses in Host Vegetative and Reproductive Stages.

Epichloë species are fungal symbionts (endophytes) of cool-season grasses that transmit vertically via inflorescence primordia (IP), ovaries (OV), and ultimately, embryos. Epichloë coenophiala, an endophyte of tall fescue (Schedonorus arundinaceus), provides multiple protective benefits to the grass. We conducted transcriptome analysis of the tall fescue-E. coenophiala symbiosis, comparing IP, OV, vegetative pseudostems (PS), and the lemma and palea (LP) (bracts) of the young floret. Transcriptomes of host OV and PS exhibited almost no significant differences attributable to endophyte presence or absence. Comparison of endophyte gene expression in different plant parts revealed numerous differentially expressed genes (DEGs). The 150 endophyte DEGs significantly higher in PS over OV included genes for alkaloid biosynthesis and sugar or amino acid transport. The 277 endophyte DEGs significantly higher in OV over PS included genes for protein chaperones (including most heat-shock proteins), trehalose synthesis complex, a bax inhibitor-1 protein homolog, the CLC chloride ion channel, catalase, and superoxide dismutase. Similar trends were apparent in the Brachypodium sylvaticum-Epichloë sylvatica symbiosis. Gene expression profiles in tall fescue IP and LP indicated that the endophyte transcriptome shift began early in host floral development. We discuss possible roles of the endophyte DEGs in colonization of reproductive grass tissues.

Uncovering Bax inhibitor-1 dual role in the legume-rhizobia symbiosis in common bean roots.

Bax-inhibitor 1 (BI-1) is a cell death suppressor conserved in all eukaryotes that modulates cell death in response to abiotic stress and pathogen attack in plants. However, little is known about its role in the establishment of symbiotic interactions. Here, we demonstrate the functional relevance of an Arabidopsis thaliana BI-1 homolog (PvBI-1a) to symbiosis between the common bean (Phaseolus vulgaris) and Rhizobium tropici. We show that the changes in expression of PvBI-1a observed during early symbiosis resemble those of some defence response-related proteins. By using gain- and loss-of-function approaches, we demonstrate that the overexpression of PvBI-1a in the roots of common bean increases the number of rhizobial infection events (and therefore the final number of nodules per root), but induces the premature death of nodule cells, affecting their nitrogen fixation efficiency. Nodule morphological alterations are known to be associated with changes in the expression of genes tied to defence, autophagy, and vesicular trafficking. Results obtained in the present work suggest that BI-1 has a dual role in the regulation of programmed cell death during symbiosis, extending our understanding of its critical function in the modulation of host immunity while responding to beneficial microbes.

Wheat Bax Inhibitor-1 interacts with TaFKBP62 and mediates response to heat stress

BackgroundHeat stress is a severe environmental stress that affects plant growth and reduces yield. Bax inhibitor-1 (BI-1) is a cytoprotective protein that is involved in the response to biotic and abiotic stresses. The Arabidopsis (Arabidopsis thaliana) BI-1 mutants atbi1–1 and atbi1–2 are hypersensitive to heat stress, and AtBI-1 overexpression rescues thermotolerance deficiency in atbi1 plants. Nevertheless, the mechanism of BI-1 in plant thermotolerance is still unclear.ResultsWe identified a wheat (Triticum aestivum L.) BI-1 gene, TaBI-1.1, which was highly upregulated in an RNA sequencing (RNA-seq) analysis of heat-treated wheat. The upregulation of TaBI-1.1 under heat stress was further demonstrated by real time quantitative PCR (qRT-PCR) and β-glucuronidase (GUS) staining. Compared with the wild type Col-0, the atbi1–2 mutant is hypersensitive to heat stress, and constitutive expression of TaBI-1.1 in atbi1–2 (35S::TaBI-1.1/ atbi1–2) rescued the deficiency of atbi1–2 under heat stress. Furthermore, we identified TaFKBP62 as a TaBI-1.1-interacting protein that co-localized with TaBI-1.1 on the endoplasmic reticulum (ER) membrane and enhanced heat stress tolerance. Additionally, HSFA2, HSFB1, ROF1, HSP17.4B, HSP17.6A, HSP17.8, HSP70B, and HSP90.1 expression levels were suppressed in atbi1–2 plants under heat stress. In contrast, 35S::TaBI-1.1/atbi1–2 relieved the inhibitory effect of AtBI-1 loss of function.ConclusionsTaBI-1.1 interacted with TaFKBP62 and co-localized with TaFKBP62 on the ER membrane. Both TaBI-1.1 and AtBI-1 regulated the expression of heat-responsive genes and were conserved in plant thermotolerance.

BI1 alleviates cardiac microvascular ischemia-reperfusion injury via modifying mitochondrial fission and inhibiting XO/ROS/F-actin pathways.

Pathogenesis of cardiac microvascular ischemia-reperfusion (IR) injury is associated with excessive mitochondrial fission. However, the upstream mediator of mitochondrial fission remains obscure. Bax inhibitor 1 (BI1) is linked to multiple mitochondrial functions, and there have been no studies investigating the contribution of BI1 on mitochondrial fission in the setting of cardiac microvascular IR injury. This study was undertaken to establish the action of BI1 on the cardiac microvascular reperfusion injury and figure out whether BI1 sustained endothelial viability via inhibiting mitochondrial fission. Our observation indicated that BI1 was downregulated in reperfused hearts and overexpression of BI1 attenuated microvascular IR injury. Mechanistically, reperfusion injury elevated the levels of xanthine oxidase (XO), an effect that was followed by increasedreactive oxygen species (ROS) production. Subsequently, oxidative stress mediated F-actin depolymerization and the latter promoted mitochondrial fission. Aberrant fission caused mitochondrial dysfunction and ultimately activated mitochondrial apoptosis in cardiac microvascular endothelial cells. By comparison, BI1 overexpression repressed XO expression and thus neutralized ROS, interrupting F-actin-mediated mitochondrial fission. The inhibitory effect of BI1 on mitochondrial fission sustained endothelial viability, reversed endothelial barrier integrity, attenuated the microvascular inflammation response, and maintained microcirculation patency. Altogether, we conclude that BI1 is essential in maintaining mitochondrial homeostasis and alleviating cardiac microvascular IR injury. Deregulated BI1 via the XO/ROS/F-actin pathways plays a causative role in the development of cardiac microvascular reperfusion injury.

Bax inhibitor-1 suppresses early brain injury following experimental subarachnoid hemorrhage in rats.

Early brain injury (EBI) following subarachnoid hemorrhage (SAH) is an important cause of high mortality and poor prognosis in SAH. B‑cell lymphoma 2‑associated X protein inhibitor‑1 (BI‑1) is an evolutionarily conserved antiapoptotic protein that is primarily located in the membranes of endoplasmic reticulum (ER). BI‑1 has been studied in certain nervous system‑associated diseases, but the role of this protein in SAH remains unclear. In the present study, the role of BI‑1 in EBI following SAH was investigated in rat models and its associated mechanisms were examined. The SAH rat model was generated by inserting nylon cords into the internal carotid artery from the external carotid artery. Samples were assessed using neurological scores, brain water content measurements, hematoxylin and eosin (H&E) staining, blood‑brain barrier (BBB) permeability, terminal deoxynucleotidyl transferase‑mediated dUTP nick‑end labeling and quantitative polymerase chain reaction assays, and western blot analyses. It was identified that the mRNA and protein levels of BI‑1 decreased markedly and were lowest at 24h after SAH. BI‑1 overexpression and small hairpin RNA (shRNA)‑mediated silencing markedly suppressed or severely exacerbated EBI following SAH, respectively. BI‑1 overexpression in the SAH model improved neurological scores and decreased the brain water content, BBB permeability and levels of apoptosis compared with the control and sham groups following SAH. BI‑1 shRNA in the SAH model demonstrated contrary results. In addition, the mRNA or protein expression levels of ER stress‑associated genes (glucose regulated protein, 78 kDa, C/EBP homologous protein, Serine/threonine‑protein kinase/endoribonuclease IRE1, c‑Jun N terminal kinases and apoptotic signaling kinase‑1) were markedly suppressed or increased following BI‑1 overexpression and shRNA‑mediated silencing, respectively. The present study suggested that BI‑1 serves a neuroprotective role in EBI following SAH by attenuating BBB disruption, brain edema and apoptosis mediated by ER stress.

Abstract 308: Yeast Bax Inhibitor, BXI1, is involved in calcium homeostasis of the ER in Saccharomyces cerevisiae

Abstract Yeast Bax inhibitor-1 (BXI1/YBH3) encodes a protein that belongs to the Bax Inhibitor (TMBIM) family of proteins that all contain a transmembrane BAX inhibitor motif. The crystal structure of a prokaryotic member of the family, BsYetJ, has revealed that the Bax inhibitor proteins are pH sensitive calcium leaks. In mammals, the Bax inhibitor family of proteins has cytoprotective properties that are most evident in paradigms of endoplasmic reticulum (ER) stress. Our published studies have shown that yeast Bxi1p is localized to the endoplasmic reticulum and is involved in the unfolded protein response (UPR) that is triggered by ER stress. BXI1 is thought to act via a mechanism involving altered calcium dynamics. We now show that cells lacking BXI1 accumulate higher levels of calcium in their ER as compared to their wildtype counterparts. We have also over expressed Bxi1p in E.coli and have used a fura-2 based calcium assay to show that the protein facilitates the influx of extracellular calcium into the cell. Our preliminary data with Δbxi1 Δpmr1 double mutants suggest that Bxi1p and Pmr1p, the Golgi-ER calcium pump, function in an antagonistic manner. [Our laboratory is supported by grant NIGMS R15 GM110578, awarded to N. Austriaco.] Citation Format: Nicholas Andrews, Nicholas Mello, Liam McDonough, Joseph Alisch, David Eagan, James Mullin, Alfredo Gonzalez, Nicanor Austriaco. Yeast Bax Inhibitor, BXI1, is involved in calcium homeostasis of the ER in Saccharomyces cerevisiae [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 308.