Major Vault Protein Research Articles

The major vault protein integrates adhesion-driven signals to regulate collagen remodeling

DDR1 interacts with fibrillar collagen and can affect β1 integrin-dependent signaling, but the mechanism that mediates functional interactions between these two different receptors is not defined. We searched for molecules that link DDR1 and β1 integrin-dependent signaling in response to collagen binding. The activation of DDR1 by binding to fibrillar collagen reduced by 5-fold, β1 integrin-dependent ERK phosphorylation that leads to MMP1 expression. In contrast, pharmacological inhibition of DDR1 or culturing cells on fibronectin restored ERK phosphorylation and MMP1 expression mediated by the β1 integrin. A phospho-site screen indicated that collagen-induced DDR1 activation inhibited β1 integrin-dependent ERK signaling by regulating autophosphorylation of focal adhesion kinase (FAK). Immunoprecipitation, mass spectrometry, and protein-protein interaction mapping showed that while DDR1 and FAK do not interact directly, the major vault protein (MVP) binds DDR1 and FAK depending on the substrate. MVP associated with DDR1 in cells expressing β1 integrin that were cultured on collagen. Knockdown of MVP restored ERK activation and MMP1 expression in DDR1-expressing cells cultured on collagen. Immunostaining of invasive cancers in human colon showed colocalization of DDR1 with MVP. These data indicate that MVP interactions with DDR1 and FAK contribute to the regulation of β1 integrin-dependent signaling pathways that drive collagen degradation.

Major Vault Protein Promotes Kidney Inflammation and Macrophage Recruitment in a Murine Model of AKI

Major Vault Protein Promotes Kidney Inflammation and Macrophage Recruitment in a Murine Model of AKI

Major Vault Protein/Lung Resistance-Related Protein: A Novel Biomarker for Inflammation and Acute Infections.

Vault particles are large cytoplasmic ribonucleoprotein particles that participate in inflammation. The aim of this study was to assess the diagnostic and prognostic value of major vault protein (MVP) in patients with inflammation, in order to determine whether MVP could be used as a biomarker for infection or inflammation. We also aimed to compare the diagnostic impact of MVP compared to other conventional measurements, such as CRP or white blood cell (WBC) counts. CRP and MVP levels were measured in 111 sera samples from 85 patients with inflammation admitted to a tertiary-care hospital and 26 healthy individuals during an 18-month period (2019-2020), using nephelometry and a custom MVP sandwich ELISA assay, respectively. In addition, WBC counts were measured using a commercial assay. MVP levels were found to be elevated in patients with inflammation compared to healthy individuals (p < 0.0001). Moreover, MVP levels were higher in patients with inflammation due to an infectious etiology compared to those with non-infectious etiology (p = 0.0006). MVP levels significantly decreased during the first four days of infection in response to antibiotic treatment, while CRP levels showed a less-sensitive decline. An ROC curve analysis demonstrated that MVP and CRP have similarly high diagnostic accuracy, with AUCs of 0.955 and 0.995, respectively, followed by WBCs with an AUC of 0.805. The ROC curves demonstrated that MVP has the potential to serve as a diagnostic biomarker for inflammation and infection. Additionally, MVP levels may reflect the efficacy of antibiotic treatment.

Single-cell sensor analyses reveal signaling programs enabling Ras-G12C drug resistance.

Clinical resistance to rat sarcoma virus (Ras)-G12C inhibitors is a challenge. A subpopulation of cancer cells has been shown to undergo genomic and transcriptional alterations to facilitate drug resistance but the immediate adaptive effects on Ras signaling in response to these drugs at the single-cell level is not well understood. Here, we used Ras biosensors to profile the activity and signaling environment of endogenous Ras at the single-cell level. We found that a subpopulation of KRas-G12C cells treated with Ras-G12C-guanosine-diphosphate inhibitors underwent adaptive signaling and metabolic changes driven by wild-type Ras at the Golgi and mutant KRas at the mitochondria, respectively. Our Ras biosensors identified major vault protein as a mediator of Ras activation through its scaffolding of Ras signaling pathway components and metabolite channels. Overall, methods including ours that facilitate direct analysis on the single-cell level can report the adaptations that subpopulations of cells adopt in response to cancer therapies, thus providing insight into drug resistance.

MVP enhances FGF21-induced ferroptosis in hepatocellular carcinoma by increasing lipid peroxidation through regulation of NOX4.

Ferroptosis is a novel, iron-dependent regulatory cell death mainly caused by an imbalance between the production and degradation of intracellular reactive oxygen species (ROS). Recently, ferroptosis induction has been considered a potential therapeutic approach for hepatocellular carcinoma (HCC). Fibroblast growth factor 21 (FGF21) is a new modulator of ferroptosis; however, the regulatory role of FGF21 in HCC ferroptosis has not been investigated. In this study, we explored the role of FGF21 and its underlying molecular mechanism in the ferroptotic death of HCC cells. We identified Major vault protein (MVP) as a target of FGF21 and revealed that knockdown of MVP inhibited the lipid peroxidation levels of HCC cells by decreasing NADPH oxidase 4 (NOX4, a major source of ROS) transcription, thereby attenuating the effect of FGF21-mediated ferroptosis. On the other hand, MVP overexpression showed the opposite results. Mechanistically, MVP binds to IRF1 and thus interferes with the interaction between IRF1 and the YAP1 promoter, leading to an increase in NOX4 transcription. Importantly, forced expression of IRF1 or downregulation of YAP1 partially reversed the effect of MVP overexpression on HCC ferroptosis. Furthermore, the results in xenograft tumor models suggested that overexpression of MVP can efficiently increase the level of lipid peroxidation invivo. Taken together, these results provide new insights into the regulatory mechanism of ferroptosis in HCC.

Major vault protein directly enhances adaptive immunity induced by Influenza A virus or indirectly through innate immunity

As we previously revealed, major vault protein (MVP) is a virus-induced host factor, and its expression is crucial for innate immune responses. Nevertheless, the function of MVP in adaptive immunity is poorly known. Here, we demonstrate that Mvp knockout mice had attenuated antibody responses and reduced survival after rechallenge with homologous influenza A virus (IAV) relative to wild-type mice. Analysis of B cell populations showed that MVP promoted germinal center (GC) responses to develop optimal antiviral humoral immunity. Although MVP-deficient T cells and dendritic cells (DCs) were not intrinsically damaged, MVP promoted activating effector T cells and T follicular helper responses and regulated specific DC subsets. These findings suggest that MVP directs an effective adaptive immune response against IAV by directly engaging in GC reactions or indirectly augmenting cellular immunity via innate immune pathways.

Major vault protein is part of an extracellular cement material in the Atlantic salmon louse (Lepeophtheirussalmonis)

Major vault protein (MVP) is the main component of the vault complex, which is a highly conserved ribonucleoprotein complex found in most eukaryotic organisms. MVP or vaults have previously been found to be overexpressed in multidrug-resistant cancer cells and implicated in various cellular processes such as cell signaling and innate immunity. The precise function of MVP is, however, poorly understood and its expression and probable function in lower eukaryotes are not well characterized. In this study, we report that the Atlantic salmon louse expresses three full-length MVP paralogues (LsMVP1-3). Furthermore, we extended our search and identified MVP orthologues in several other ecdysozoan species. LsMVPs were shown to be expressed in various tissues at both transcript and protein levels. In addition, evidence for LsMVP to assemble into vaults was demonstrated by performing differential centrifugation. LsMVP was found to be highly expressed in cement, an extracellular material produced by a pair of cement glands in the adult female salmon louse. Cement is important for the formation of egg strings that serve as protective coats for developing embryos. Our results imply a possible novel function of LsMVP as a secretory cement protein. LsMVP may play a role in structural or reproductive functions, although this has to be further investigated.

Structural Insights into the Roles of PARP4 and NAD + in the Human Vault Cage.

Vault is a massive ribonucleoprotein complex found across Eukaryota. The major vault protein (MVP) oligomerizes into an ovular cage, which contains several minor vault components (MVCs) and is thought to transport transiently bound "cargo" molecules. Vertebrate vaults house a poly (ADP-ribose) polymerase (known as PARP4 in humans), which is the only MVC with known enzymatic activity. Despite being discovered decades ago, the molecular basis for PARP4's interaction with MVP remains unclear. In this study, we determined the structure of the human vault cage in complex with PARP4 and its enzymatic substrate NAD + . The structures reveal atomic-level details of the protein-binding interface, as well as unexpected NAD + -binding pockets within the interior of the vault cage. In addition, proteomics data show that human vaults purified from wild-type and PARP4-depleted cells interact with distinct subsets of proteins. Our results thereby support a model in which PARP4's specific incorporation into the vault cage helps to regulate vault's selection of cargo and its subcellular localization. Further, PARP4's proximity to MVP's NAD + -binding sites could support its enzymatic function within the vault.

An Efficient Method for Vault Nanoparticle Conjugation with Finely Adjustable Amounts of Antibodies and Small Molecules.

Vaults are eukaryotic ribonucleoproteins consisting of 78 copies of the major vault protein (MVP), which assemble into a nanoparticle with an about 60 nm volume-based size, enclosing other proteins and RNAs. Regardless of their physiological role(s), vaults represent ideal, natural hollow nanoparticles, which are produced by the assembly of the sole MVP. Here, we have expressed in Komagataella phaffi and purified an MVP variant carrying a C-terminal Z peptide (vault-Z), which can tightly bind an antibody's Fc portion, in view of targeted delivery. Via surface plasmon resonance analysis, we could determine a 2.5 nM affinity to the monoclonal antibody Trastuzumab (Tz)/vault-Z 1:1 interaction. Then, we characterized the in-solution interaction via co-incubation, ultracentrifugation, and analysis of the pelleted proteins. This showed virtually irreversible binding up to an at least 10:1 Tz/vault-Z ratio. As a proof of concept, we labeled the Fc portion of Tz with a fluorophore and conjugated it with the nanoparticle, along with either Tz or Cetuximab, another monoclonal antibody. Thus, we could demonstrate antibody-dependent, selective uptake by the SKBR3 and MDA-MB 231 breast cancer cell lines. These investigations provide a novel, flexible technological platform that significantly extends vault-Z's applications, in that it can be stably conjugated with finely adjusted amounts of antibodies as well as of other molecules, such as fluorophores, cell-targeting peptides, or drugs, using the Fc portion as a scaffold.

Senescent Macrophages Release Inflammatory Cytokines and RNA-Loaded Extracellular Vesicles to Circumvent Fibroblast Senescence.

Senescent cells, which accumulate with age, exhibit a pro-inflammatory senescence-associated secretory phenotype (SASP) that includes the secretion of cytokines, lipids, and extracellular vesicles (EVs). Here, we established an in vitro model of senescence induced by Raf-1 oncogene in RAW 264.7 murine macrophages (MΦ) and compared them to senescent MΦ found in mouse lung tumors or primary macrophages treated with hydrogen peroxide. The transcriptomic analysis of senescent MΦ revealed an important inflammatory signature regulated by NFkB. We observed an increased secretion of EVs in senescent MΦ, and these EVs presented an enrichment for ribosomal proteins, major vault protein, pro-inflammatory miRNAs, including miR-21a, miR-155, and miR-132, and several mRNAs. The secretion of senescent MΦ allowed senescent murine embryonic fibroblasts to restart cell proliferation. This antisenescence function of the macrophage secretome may explain their pro-tumorigenic activity and suggest that senolytic treatment to eliminate senescent MΦ could potentially prevent these deleterious effects.

The prognostic value and immunological role of MVP in pan-cancer study.

Major Vault Protein (MVP) has emerged as a potential prognostic and immunological biomarker in various cancer types. This pan-cancer study aimed to investigate expression of MVP and its correlation with clinical outcomes and immune infiltration across diverse cancer types. We conducted an analysis of extensive transcriptomic and clinical data from publicly available databases. Our findings unveiled a significant association between MVP expression and cancer progression, with higher expression levels predicting poorer overall survival in multiple cancer types. Importantly, MVP expression demonstrated a close relationship with immune infiltration in the tumor microenvironment, showing that higher expression levels were associated with increased immune cell infiltration. We further validated expression of MVP and function in cancer cell lines A549 and AGS. These compelling results suggest that MVP holds promise as a valuable biomarker for prognostic assessment and the development of immunotherapeutic strategies across various cancer types. Consequently, targeting MVP may offer a compelling therapeutic approach in the treatment of human cancers.

Human Vault RNAs: Exploring Their Potential Role in Cellular Metabolism.

Non-coding RNAs have been described as crucial regulators of gene expression and guards of cellular homeostasis. Some recent papers focused on vault RNAs, one of the classes of non-coding RNA, and their role in cell proliferation, tumorigenesis, apoptosis, cancer response to therapy, and autophagy, which makes them potential therapy targets in oncology. In the human genome, four vault RNA paralogues can be distinguished. They are associated with vault complexes, considered the largest ribonucleoprotein complexes. The protein part of these complexes consists of a major vault protein (MVP) and two minor vault proteins (vPARP and TEP1). The name of the complex, as well as vault RNA, comes from the hollow barrel-shaped structure that resembles a vault. Their sequence and structure are highly evolutionarily conserved and show many similarities in comparison with different species, but vault RNAs have various roles. Vaults were discovered in 1986, and their functions remained unclear for many years. Although not much is known about their contribution to cell metabolism, it has become clear that vault RNAs are involved in various processes and pathways associated with cancer progression and modulating cell functioning in normal and pathological stages. In this review, we discuss known functions of human vault RNAs in the context of cellular metabolism, emphasizing processes related to cancer and cancer therapy efficacy.

Abstract 1663: Single-cell signaling analysis reveals that major vault protein facilitates RasG12C inhibitor resistance

Abstract Covalent inhibitors of RasG12C are the first clinically-approved drugs for targeting mutant Ras-driven cancers. However, the rapid development of clinical resistance to current Ras G12C inhibitors is common. It has been demonstrated that a subpopulation of cancer cells facilitate drug resistance but the mechanistic basis of this heterogenous response is not well understood. Here, we utilized recently developed Ras sensors to profile the environment of active Ras and to measure the activity of endogenous Ras to pair structure (Ras signalosome) to function (Ras activity), respectively, at a single-cell level. With this approach, we found that a subpopulation of KRasG12C cells treated with RasG12C-GDP inhibitors underwent oncogenic signaling and metabolic changes driven by WT Ras at the golgi and mutant Ras at mitochondria, respectively. Our Ras sensors identified Major Vault Protein (MVP) as a mediator of Ras activation at both subcellular locations by scaffolding Ras signaling pathway components and metabolite channels. We found that recently developed RasG12C-GTP inhibitors also led to MVP-mediated WT Ras signaling at the golgi, demonstrating that this a general mechanism of RasG12C inhibitor resistance. Overall, single-cell analyses enabled the discovery of a RasG12C inhibitor-resistant subpopulation driven by MVP, providing insight into the complex and heterogenous rewiring that confers drug resistance in cancer. Citation Format: Jason Z. Zhang, Shao-En Ong, David Baker, Dustin J. Maly. Single-cell signaling analysis reveals that major vault protein facilitates RasG12C inhibitor resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1663.

Dissecting 16p11.2 hemi-deletion to study sex-specific striatal phenotypes of neurodevelopmental disorders.

Neurodevelopmental disorders (NDDs) are polygenic in nature and copy number variants (CNVs) are ideal candidates to study the nature of this polygenic risk. The disruption of striatal circuits is considered a central mechanism in NDDs. The 16p11.2 hemi-deletion (16p11.2 del/+) is one of the most common CNVs associated with NDD, and 16p11.2 del/+ mice show sex-specific striatum-related behavioral phenotypes. However, the critical genes among the 27 genes in the 16p11.2 region that underlie these phenotypes remain unknown. Previously, we applied a novel strategy to identify candidate genes associated with the sex-specific phenotypes of 16p11.2 del/+ mice and highlighted three genes within the deleted region: thousand and one amino acid protein kinase 2 (Taok2), seizure-related 6 homolog-like 2 (Sez6l2), and major vault protein (Mvp). Using CRISPR/Cas9, we generated mice carrying null mutations inTaok2, Sez6l2, and Mvp (3 gene hemi-deletion (3g del/+)). Hemi-deletion of these 3 genes recapitulates sex-specific behavioral alterations in striatum-dependent behavioral tasks observed in 16p11.2 del/+ mice, specifically male-specific hyperactivity and impaired motivation for reward seeking. Moreover, RNAseq analysis revealed that 3g del/+ mice exhibit gene expression changes in the striatum similar to 16p11.2 del/+ mice exclusively in males. Subsequent analysis identified translation dysregulation and/or extracellular signal-regulated kinase signaling as plausible molecular mechanisms underlying male-specific, striatum-dependent behavioral alterations. Interestingly, ribosomal profiling supported the notion of translation dysregulation in both 3g del/+ and 16p11.2 del/+ male mice. However, mice carrying a 4-gene deletion (with an additional deletion of Mapk3) exhibited fewer phenotypic similarities with 16p11.2 del/+ mice. Together, the mutation of 3 genes within the 16p11.2 region phenocopies striatal sex-specific phenotypes of 16p11.2 del/+ mice. These results support the importance of a polygenic approach to study NDDs and underscore that the effects of the large genetic deletions result from complex interactions between multiple candidate genes.

Major vault protein regulates tumor-associated macrophage polarization through interaction with signal transducer and activator of transcription 6.

Tumor-associated macrophages (TAMs) are critical in the tumor microenvironment (TME) of hepatocellular carcinoma (HCC). Major vault protein (MVP) mediates multidrug resistance, cell growth and development, and viral immunity. However, the relationship between MVP and TAMs polarization has not been clarified in HCC. We found that MVP significantly increased M2-TAMs infiltration levels in tumor tissues of HCC patients. MVP promoted HCC proliferation, metastasis, and invasion by regulating M2 polarization in vivo and in vitro. Mechanistically, MVP associated with signal transducer and activator of transcription 6 (STAT6) and enhanced STAT6 phosphorylation. STAT6 translocated from the cytosol to the nucleus and regulated M2 macrophage-associated gene transcription. These findings suggest that MVP modulates the macrophage M2 transcriptional program, revealing its potential role in the TAMs of TME.

Label-free quantitative proteomic profiling reveals differential plasma protein expression in patients with obesity after treatment with liraglutide.

Treatment and management of obesity is clinically challenging. The inclusion of GLP-1 receptor agonists (GLP1RA) in the medical management of obesity has proven to be efficacious. However, mechanisms underlying the molecular changes arising from GLP1RA treatment in patients with obesity remain to be elucidated. A single-center, prospective study was undertaken to evaluate the changes in the plasma proteins after liraglutide 3 mg therapy in twenty patients (M/F: 7/13) with obesity (mean BMI 40.65 ± 3.7 kg/m2). Anthropometric and laboratory parameters were measured, and blood samples were collected at two time points: baseline, before initiating treatment (pretreatment group, PT), and after three months of receiving the full dose liraglutide 3 mg (posttreatment group, PoT). An untargeted label-free LC MSMS mass spectrometric approach combined with bioinformatics and network pathway analysis was used to determine changes in the proteomic profiles. The mean age of the study participants was 36.0 ± 11.1 years. A statistically significant change was observed in weight, BMI and HbA1c levels between the PT and PoT groups (paired t-test, P < 0.001). A significant dysregulation was noted in the abundances of 151 proteins (31 up and 120 downregulated) between the two groups. The potential biomarkers were evaluated using receiver operating characteristic (ROC) curves. The top ten proteins (area under the curve (AUC) of 0.999 (95% CI)) were identified as potential biomarkers between PT and PoT groups and included Cystatin-B, major vault protein, and plastin-3, which were upregulated, whereas multimerin-2, large ribosomal P2, and proline-rich acidic protein 1 were downregulated in the PoT group compared with the PT group. The top network pathway identified using ingenuity pathway analysis (IPA), centered around dysregulation of MAPK, AKT, and PKc signaling pathways and related to cell-to-cell signaling and interaction, cellular assembly and organization, cellular compromise and a score of 46 with 25 focus proteins. Through label-free quantitative proteomic analysis, our study revealed significant dysregulation of plasma proteins after liraglutide 3 mg treatment in patients with obesity. The alterations in the proteomic profile between the PT and PoT groups demonstrated a decrease in levels of proteins involved in inflammation and oxidative stress pathways. On the other hand proteins involved in the glycolytic and lipolytic metabolic pathways as well as those participating in cytoskeletal and endothelial reorganization were observed to be increased. Understanding actions of liraglutide at a molecular and proteomic levels provides a holistic look into how liraglutide impacts metabolism, induces weight loss and improves overall metabolic health.

Structure, Dynamics and Functional Implications of the Eukaryotic Vault Complex.

Vault ribonucleoprotein particles are naturally designed nanocages, widely found in the eukaryotic kingdom. Vaults consist of 78 copies of the major vault protein (MVP) that are organized in 2 symmetrical cup-shaped halves, of an approximate size of 70x40x40 nm, leaving a huge internal cavity which accommodates the vault poly(ADP-ribose) polymerase (vPARP), the telomerase-associated protein-1 (TEP1) and some small untranslated RNAs. Diverse hypotheses have been developed on possible functions of vaults, based on their unique capsular structure, their rapid movements and the distinct subcellular localization of the particles, implicating transport of cargo, but they are all pending confirmation. Vault particles also possess many attributes that can be exploited in nanobiotechnology, particularly in the creation of vehicles for the delivery of multiple molecular cargoes. Here we review what is known about the structure and dynamics of the vault complex and discuss a possible mechanism for the vault opening process. The recent findings in the characterization of the vaults in cells and in its natural microenvironment will be also discussed.

Unraveling the role of Major Vault Protein as a novel immune-related biomarker that promotes the proliferation and migration in pancreatic adenocarcinoma.

Pancreatic adenocarcinoma (PAAD) is a formidable challenge in oncology research, with a complex pathogenesis that requires to be explored. Major Vault Protein (MVP) is the principal structural component of the vault complex, and its expression level is remarkably upregulated in various cancers. Extensive investigations have been conducted to explore the role of MVP in specific cancer contexts, yet the potential molecular mechanisms and biological functions of MVP in PAAD still remain considerably elusive. This study aims to explore the role of MVP as a novel immune-related biomarker in the pathogenesis and clinical treatment of PAAD. Gene expression data and clinical information were collected from TCGA, GTEx and GEO databases. Survival, prognostic and functional enrichment analysis were employed with R software. Immunological correlation analysis was performed using TIMER2.0, TIDE scores, TISIDB and TISCH. Epigenetic analysis was implemented by MethSurv, CPTAC, UALCAN, and cBioPortal. Drug analysis was conducted using Enrichr and CellMiner. Moreover, cellular experiments, like RNA interference, qRT-PCR, Western blot, cell cycle analysis, cell apoptosis analysis, colony formation assay, transwell assay, and wound healing assay, were performed for verifying the functional properties of MVP in the PAAD progression. We demonstrated an abnormally upregulated expression of MVP in PAAD tissues, which notably correlated with an adverse prognosis in PAAD patients. Functional analysis suggested the conceivable involvement of MVP in immune modulation, and immunotherapy. Additionally, we identified genetic alterations, reduced promoter methylation, and heightened phosphorylation in MVP. We also clarified Suloctidil and Tetradioxin as the most notable potential drugs targeting MVP in PAAD. Moreover, our experimental observations consistently highlighted the significant impact of MVP deficiency on impeding PAAD cell proliferation, inhibiting cell migration, and accelerating cell apoptosis. Interestingly, a potential link between MVP and ERK or AKT pathways was displayed, which opens new avenues for further exploration of the molecular mechanisms of MVP-targeted therapies in PAAD. This study systematically describes MVP as an immune-related biomarker with remarkable potential for predicting the prognosis, tumor progression and immunotherapeutic efficacy in PAAD.

Dissecting the autism-associated 16p11.2 locus identifies multiple drivers in neuroanatomical phenotypes and unveils a male-specific role for the major vault protein

BackgroundUsing mouse genetic studies and systematic assessments of brain neuroanatomical phenotypes, we set out to identify which of the 30 genes causes brain defects at the autism-associated 16p11.2 locus.ResultsWe show that multiple genes mapping to this region interact to regulate brain anatomy, with female mice exhibiting far fewer brain neuroanatomical phenotypes. In male mice, among the 13 genes associated with neuroanatomical defects (Mvp, Ppp4c, Zg16, Taok2, Slx1b, Maz, Fam57b, Bola2, Tbx6, Qprt, Spn, Hirip3, and Doc2a), Mvp is the top driver implicated in phenotypes pertaining to brain, cortex, hippocampus, ventricles, and corpus callosum sizes. The major vault protein (MVP), the main component of the vault organelle, is a conserved protein found in eukaryotic cells, yet its function is not understood. Here, we find MVP expression highly specific to the limbic system and show that Mvp regulates neuronal morphology, postnatally and specifically in males. We also recapitulate a previously reported genetic interaction and show that Mvp+/−;Mapk3+/− mice exhibit behavioral deficits, notably decreased anxiety-like traits detected in the elevated plus maze and open field paradigms.ConclusionsOur study highlights multiple gene drivers in neuroanatomical phenotypes, interacting with each other through complex relationships. It also provides the first evidence for the involvement of the major vault protein in the regulation of brain size and neuroanatomy, specifically in male mice.

Edwardsiella ictaluri T3SS effector EseN is a phosphothreonine lyase that inactivates ERK1/2, p38, JNK, and PDK1 and modulates cell death in infected macrophages.

EseN is an Edwardsiella ictaluri type III secretion system effector with phosphothreonine lyase activity. In this work, we demonstrate that EseN inactivates p38 and c-Jun-N-terminal kinase (JNK) in infected head-kidney-derived macrophages (HKDMs). We have previously reported inactivation of extracellular-regulated kinase 1/2 (ERK1/2). Also, for the first time, we demonstrated that EseN is involved in the inactivation of 3-phosphoinositide-dependent kinase 1 (PDK1), which has not been previously demonstrated for any of the EseN homologs in other species. We also found that EseN significantly affected mRNA expression of IL-10, pro-apoptotic baxa, and p53, but had no significant effect on anti-apoptotic bcl2 or pro-apoptotic apoptotic peptidase activating factor 1. EseN is also involved in the inhibition of caspase-8 and caspase-3/7 but does not affect caspase-9 activity. Repression of apoptosis was further confirmed with flow cytometry using Alexa Fluor 647-labeled annexin V and propidium iodide. In addition, we found that the E. ictaluri T3SS is essential for the inhibition of IL-1β maturation, but EseN is not involved in this process. EseN did not affect cell pyroptosis, as indicated by the lack of EseN impact on the release of lactate dehydrogenase from infected HKDM. The transmission electron microscopy data also indicate that HKDM infected with WT or an eseN mutant died by apoptosis, while HKDM infected with the T3SS mutant more likely died by pyroptosis. Collectively, our results indicate that E. ictaluri EseN is involved in inactivation of ERK1/2, p38, JNK, and PDK1 signaling pathways that lead to modulation of cell death among infected HKDMs. IMPORTANCE This work has global significance in the catfish industry, which provides food for increasing global populations. E. ictaluri is a leading cause of disease loss, and EseN is an important player in E. ictaluri virulence. The E. ictaluri T3SS effector EseN plays an essential role in establishing infection, but the specific role EseN plays is not well characterized. EseN belongs to a family of phosphothreonine lyase effectors that specifically target host mitogen activated protein kinase (MAPK) pathways important in regulating host responses to infection. No phosphothreonine lyase equivalents are known in eukaryotes, making this family of effectors an attractive target for indirect narrow-spectrum antibiotics. Targeting of major vault protein and PDK1 kinase by EseN has not been reported in EseN homologs in other pathogens and may indicate unique functions of E. ictaluri EseN. EseN targeting of PDK1 is particularly interesting in that it is linked to an extraordinarily diverse group of cellular functions.