Immune Proteins Research Articles

Effects of immunoproteasome inhibition on acute respiratory infection with murine hepatitis virus strain 1.

Inflammatory responses triggered by acute infection by respiratory viruses such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) drive morbidity and mortality. Infection of mice with murine hepatitis virus strain 1 (MHV-1), a mouse coronavirus, is a useful model to study the pathogenesis of coronavirus respiratory infections. The immunoproteasome is an inducible component of the ubiquitin proteasome system that is poised to contribute to multiple aspects of immune function, inflammation, and protein homeostasis during an infection. We used the MHV-1 model to define the role of the immunoproteasome in coronavirus pathogenesis. We found that immunoproteasome subunit expression increases in the lungs and the liver during acute MHV-1 respiratory infection. Inhibition of immunoproteasome activity did not affect MHV-1 replication but increased MHV-1-induced weight loss, mortality, and inflammation in lungs and livers. Thus, our findings indicate that the immunoproteasome is a critical protective host factor during coronavirus respiratory infection.

Preclinical assessment of the efficacy of B7-H3 CAR-T in renal cell carcinoma

B7-H3 is a type I transmembrane protein that belongs to the B7 immune checkpoint protein family, is aberrantly expressed in cancer cells, but rarely expressed in normal tissues, making it an attractive target for cancer therapy. Here, we found B7-H3 is highly expressed in the renal cell carcinoma (RCC) tumor tissues and RCC cell lines, but is undetectable in normal renal tissues. Therefore, we engineered second-generation CAR-T cells targeting B7-H3, incorporating either CD28 or 4–1BB co-stimulatory domains. Both CAR-T cell variants demonstrated potent antitumor activity against RCC tumors in vitro and in metastatic and orthotopic RCC mouse models. Furthermore, the B7-H3 CAR-T cells exhibited remarkable proliferation and robust cytokine release when co-cultured with RCC cancer cells. These findings demonstrated that targeting B7-H3 by CAR-T cells potentially offering a new treatment option for RCC patients.

A widespread phage-encoded kinase enables evasion of multiple host antiphage defence systems.

DNA degradation (Dnd) is a widespread bacterial antiphage defence system that relies on DNA phosphorothioate (PT) modification for self/non-self discrimination and subsequent degradation of unmodified DNA. Phages employ counterstrategies to evade host immunity, but anti-Dnd immunity has not been characterized. Here we report an immune evasion protein encoded by the Salmonella phage JSS1 that contributes to subverting Dnd and other defence systems. Using quantitative proteomic and phosphoproteomic analyses, we show that the protein JSS1_004 employs N-terminal Ser/Thr/Tyr protein kinase activity to catalyse the multisite phosphorylation of host DndFGH. Notably, JSS1_004 also phosphorylates other bacterial immune systems to varying degrees, including CRISPR‒Cas, QatABCD, SIR2+HerA and DUF4297+HerA. Given that JSS1_004 and its homologues are widespread in phylogenetically diverse phages, we suggest that this strategy constitutes a family of immune evasion proteins that increases the chances of phage proliferation even when a host deploys multiple defence systems.

MMP2 enzyme-responsive extracellular vesicles as dual-targeted carriers to promote the phagocytosis of macrophages

Combination therapy using inhibition of tumor cell escape and alteration of the tumor microenvironment offers a new strategy for cancer treatment. This study aimed to develop an extracellular vesicle (EV) carrier that regulates tumor cells and the tumor microenvironment to achieve efficient tumor immunotherapy. The ligand modified on carriers targets the immune checkpoint CD47 protein, blocking tumor cell escape. This ligand is cleaved by the MMP2 enzyme and assembles into nanofibers, extending the retention time in the tumor. The carriers target the CD206 protein, enabling efficient uptake by M2 macrophages. Carriers with a high density of ligands (anti-CD206) exhibit strong receptorligand interactions with tumor cells. Due to their high rigidity, these EVs have difficulty deforming during the transmembrane process, reducing resistance and resulting in low uptake efficiency by M2 cells. The optimal uptake efficiency by M2 macrophages is achieved when the mass ratio of ligand to EVs is 1:25. Crocin loaded in EVs facilitates the polarization of M2 macrophages into M1 cells, which can phagocytize tumor cells. This study reveals a potential strategy for using extracellular vesicles in tumor treatment.

Structure-aware annotation of leucine-rich repeat domains.

Protein domain annotation is typically done by predictive models such as HMMs trained on sequence motifs. However, sequence-based annotation methods are prone to error, particularly in calling domain boundaries and motifs within them. These methods are limited by a lack of structural information accessible to the model. With the advent of deep learning-based protein structure prediction, existing sequenced-based domain annotation methods can be improved by taking into account the geometry of protein structures. We develop dimensionality reduction methods to annotate repeat units of the Leucine Rich Repeat solenoid domain. The methods are able to correct mistakes made by existing machine learning-based annotation tools and enable the automated detection of hairpin loops and structural anomalies in the solenoid. The methods are applied to 127 predicted structures of LRR-containing intracellular innate immune proteins in the model plant Arabidopsis thaliana and validated against a benchmark dataset of 172 manually-annotated LRR domains.

2023 Nobel Prize in Physics and Medicine

The 2023 Nobel Prize in Physiology and Medicine was awarded jointly to Katalin Karikó and Drew Weissman for their contributions to the development of mRNA vaccines against COVID-19. Dr. Karikó obtained her doctoral degree from the University of Szeged before moving to the United States and obtaining the position of an adjunct professor at the University of Pennsylvania.1 Dr. Weissman obtained an M.D. and a Ph.D. degree at Boston University and subsequently obtained a faculty position at the Perelman School of Medicine at the University of Pennsylvania. The two researchers met during their shared time at the University of Pennsylvania and collaborated in exploring mechanisms in which mRNA can be used to stimulate immunity development in the body.2 By modifying nucleotides in mRNAs, the pair discovered that the subsequent introduction of mRNAs into cells led to reduced inflammatory responses and increased immune protein production. Before the COVID-19 pandemic, most vaccines stimulated immune responses via attenuated viruses, proteins, or viral genetic code-carrying vectors. These vaccines, while effective, required substantial resources to produce the number of cell cultures needed to synthesize adequate supplies of vaccine to combat pandemics, such as COVID-19.3 Contrastingly, mRNA vaccines can be prepared without cell cultures through in vitro transcription and nucleotides can be modified to adapt to mutating pathogens - flexibilities that allowed more than 12 billion COVID-19 vaccines to be produced and administered in less than three years of the start of the pandemic.4,5

Immunometabolic Contributions of Atopobiaceae Family Members in Human Papillomavirus Infection, Cervical Dysplasia and Cancer.

In the cervicovaginal environment, HPV acquisition and cervical cancer progression are linked to non-Lactobacillus dominance, of which Atopobiaceae are key taxa. We hypothesize that Atopobiaceae modulates the cervicovaginal microenvironment to promote HPV persistence and progression to cancer. However, the extent to which Atopobiaceae impact the immunometabolic microenvironment is poorly understood. We investigated Atopobiaceae in a cohort of primarily Hispanic and non-Hispanic White women who were HPV-negative (n=20), HPV-positive (n=31) without dysplasia, diagnosed with cervical dysplasia (n=38), or newly diagnosed with invasive cervical carcinoma (n=9). Microbiome data was integrated with clinical and demographic surveys, immunoproteomics, and metabolomics data. Atopobiaceae identified were Fannyhessea vaginae, Fannyhessea massiliense, Fannyhessea species type 2, Lancefieldella deltae, and an unclassified species. A higher prevalence of Atopobiaceae was observed in women who were Hispanic and had higher gravidity and parity. F. species type 2 and F. vaginae were observed with infections of high-risk HPV genotypes 31 and 52. Atopobiacaeae were negatively correlated with Lactobacillus and positively correlated to Sneathia, Dialister, Anaerococcus, Prevotella, and Bifidobacterium/Gardnerella. Proinflammatory cytokines (IL-1α, IL-1β, IL-12, TNFα), immune checkpoint proteins (PD-L1, LAG3), and cancer biomarkers (CEA, MIF, TRAIL) were positively associated with Atopobiaceae-rich profiles. Pro-oncogenic metabolites, including 4-hydroxybutyrate and sphingosine, were also elevated in women colonized by Atopobiaceae. Our data implicate Atopobiaceae in lipid modulation, oxidative stress, inflammatory responses, and immune evasion, which may contribute to cancer. This study highlights a key family of pathogenic cervicovaginal bacteria that could be exploited to monitor HPV persistence and/or targeted to prevent HPV-mediated cancer.

Optimising Growth, Immunity, and Gene Expression in Broiler Chickens Through Dietary Threonine Levels and Oil Inclusion.

The inclusion of synthetic amino acids in poultry nutrition plays a crucial role in both enhancing the synthesis of immunoglobulins and elevating the overall comprehensiveness of the amino acid profile. This research examined the effects of consuming threonine (Thr) in various forms levels with low or high oil on broiler chickens' growth and immunity. We investigate the growth performance, feed efficiency, immune response, intestinal morphology, absorptive capacity, and expression of some genes related to the feed intake (Pro-opiomelanocortin [POMC]), fatty acid synthesis (Acetyl-CoA Carboxylase [ACC]), immunity (lipopolysaccharide-induced tumour necrosis like alpha factor [LITAF]), and heat shock protein 70 (HSP70). Eight groups of chicks were used, including four dietary Thr levels (100%, 115%, 130%, or 145%) with two oil levels (mixture of sunflower 50% and soybean oils 50%): (control) and high. The higher dietaryThrlevel (145%) with high oil inclusion significantly increased ACC and POMC gene expression, resulting in the lowest feed intake, body weight gain (BWG), and liver fat content. Combining high oil with 115% Thr was the optimum for the broilers. The birds have significant (p ≤.05) growth performance, immune parameters, and intestinal health, as well as the lowest expression of ACC, POMC, HSP70, and LITAF, which was reflected in better feed conversion ratio and lower incidence of fatty liver, thermo-resistance, and immune status of the birds. The combination of high oil and 115% Thr levels optimises broiler health and productivity, enhancing growth, immune function, and gut health. This diet lowers the expression of genes associated with fatty liver and stress, leading to better feed efficiency, thermo-resistance, and overall well-being. Adopting these dietary adjustments can improve broiler performance and economic viability in poultry farming by enhancing essential productivity metrics.

A small microcin plays a big role in V. cholerae interbacterial competition

Microcins are antibacterial small proteins secreted by gram-negative bacteria. In this issue of Cell Host & Microbe, Kim etal. report the discovery of a V.cholerae microcin, MvcC. MvcC shows antibacterial activity against non-self V.cholerae strains, which do not encode the cognate immunity protein.

The antibacterial activity of a prophage-encoded fitness factor is neutralized by two cognate immunity proteins

The antibacterial activity of a prophage-encoded fitness factor is neutralized by two cognate immunity proteins

Compression force promotes glioblastoma progression through the Piezo1‑GDF15‑CTLA4 axis.

Glioma, a type of brain tumor, is influenced by mechanical forces in its microenvironment that affect cancer progression. However, our understanding of the contribution of compression and its associated mechanisms remains limited. The objective of the present study was to create an environment in which human brain glioma H4 cells experience pressure and thereby investigate the compressive mechanosensors and signaling pathways. Subsequent time‑lapse imaging and wound healing assays confirmed that 12 h of compression significantly increased cell migration, thereby linking compression with enhanced cell motility. Compression upregulated the expression of Piezo1, a mechanosensitive ion channel, and growth differentiation factor 15 (GDF15), a TGF‑β superfamily member. Knockdown experiments targeting PIEZO1 or GDF15 using small interfering RNA resulted in reduced cell motility, with Piezo1 regulating GDF15 expression. Compression also upregulated CTLA4, a critical immune checkpoint protein. The findings of the present study therefore suggest that compression enhances glioma progression by stimulating Piezo1, promoting GDF15 expression and increasing CTLA4 expression. Thus, these findings provide important insights into the influence of mechanical compression on glioma progression and highlight the involvement of the Piezo1‑GDF15 signaling pathway. Understanding tumor responses to mechanical forces in the brain microenvironment may guide the development of targeted therapeutic strategies to mitigate tumor progression and improve patient outcomes.

Soluble immune checkpoints associated with disease activity and treatment response in GD and TED.

Soluble immune checkpoints play an important role in peripheral tolerance that has seldom been investigated in Graves' disease (GD) and thyroid eye disease (TED). The objective of this work is to examine the alteration of soluble immune checkpoints in GD and TED. We performed a quantitative multiplex analysis of 17 immune checkpoint proteins in serum from 50 GD patients without TED, 28 GD patients with TED and 40 healthy controls. The association with demographical, serological, clinical features and 27 cytokines was analyzed. A follow-up was conducted in GD patients without TED. Functional outcomes of sLAG-3 and sGITR were assessed in cell cultures using rh-LAG3, rh-GITR, an antagonistic LAG-3 antibody and an antagonistic GITR antibody. GD patients with TED had distinct sICP and cytokine profiles compared with GD patients without TED. Active TED patients exhibited elevation in the levels of sBTLA, sLAG-3, sGITR, sCD80, sCD86 and sPD-L1. Further, GD patients without TED with high sBTLA, sCD27 and sCD40 levels at baseline showed a better improvement in thyrotropin receptor antibody (TRAb) titers after antithyroid drug (ATD) treatment. Adding recombinant human GITR and LAG-3 to PBMC cultures resulted in increased inflammatory cytokine secretion and decreased anti-inflammatory cytokine secretion. The present study uncovers disturbed soluble immune checkpoints and cytokines in GD patients with and without TED, and may pave the way for novel immunological screening, allowing for identification of TED patients at higher risk of developing active disease and GD patients with better treatment response after ATD treatment.

Development of a PD-L1 Overexpression System in HEK293T Cells: A Platform for Studying Immune Evasion Mechanisms

Programmed death-ligand 1 (PD-L1) is a key immune checkpoint protein that facilitates tumor immune evasion by interacting with PD-1 receptors on T cells, making the PD-1/PD-L1 axis a critical target in cancer immunotherapy. However, resistance to these therapies remains a challenge, necessitating further exploration of PD-L1’s functions and regulatory mechanisms. In this study, we amplified the PD-L1 coding sequence (CDS) from human cDNA, cloned it into a pcDNA3.1 vector, and transfected it into HEK293T cells to establish a robust in vitro PD-L1 overexpression system. Quantitative RT-qPCR demonstrated a 300-400 fold increase in PD-L1 mRNA expression compared to controls. These results confirm the successful construction of the PD-L1 overexpression plasmid and validate its application as a tool for studying PD-L1’s role in immune modulation and its potential implications in overcoming resistance to immunotherapy. This system provides a valuable platform for future investigations into PD-L1-mediated mechanisms of immune evasion and cancer progression.

Low plasma levels of BTLA and LAG-3 before HCV therapy are associated with metabolic disorders after HCV eradication in persons with HIV/HCV coinfection: a retrospective study.

Understanding the predictors of metabolic disorders in persons with HIV/HCV coinfection post-HCV therapy is crucial for improving patient outcomes. Since immune checkpoint proteins are usually upregulated in these persons with HIV/HCV coinfection, we aimed to evaluate the association between plasma immune checkpoint proteins at baseline (before HCV therapy) and metabolic disturbances during the follow-up (about 5years after successful HCV treatment) in persons with HIV/HCV coinfection. We performed a retrospective study on 80 persons with HIV/HCV coinfection with advanced fibrosis or cirrhosis who cleared HCV infection after successful HCV therapy and were followed for about 5 years after completion of HCV treatment. Plasma samples were collected at baseline. Immune checkpoint proteins were analyzed using a Luminex 200™ analyzer. Outcomes were the development of a metabolic event (type 2 diabetes mellitus and/or dyslipidemia) and the change in Triglycerides and glucose (TyG) index. During follow-up, 21 (26%) patients developed metabolic events (type 2 diabetes mellitus/dyslipidemia), and 29 (46.0%) patients had an increase in TyG during the follow-up. Low baseline values of BTLA and LAG-3, two immune checkpoint proteins, were associated with the development of metabolic events (aAMR = 0.69 and aAMR = 0.71, respectively) and with increases in TyG values (aAMR = 0.72 and aAMR = 0.70, respectively). In addition, other immune checkpoint proteins were also inversely associated with increases in TyG. We discovered that low plasma levels of BTLA and LAG-3 before HCV therapy significantly correlate with an increased risk of developing metabolic disorders after treatment.

Immunoproteomics: Approach to Diagnostic and Vaccine Development.

The study of large protein sets (proteomics) involved in the immunological reaction is known as immunoproteomics. The methodology of immunoproteomics plays a major role in identifying possible vaccine candidates that could protect against pathogenic infection. The study of immunogenic proteins that are expressed during the outset of infection is the focus of the crosstalk between proteomics and immune protection antigens utilizing serum. Peptide presentation by MHC provides the new 'window' into changes that occur in the cell. Thus, there is strong, intense pressure on the pathogen that has been mutated in such an unusual manner that it can bypass the MHC peptide presentation by the MHC molecule. The pathogen's ability to evade the immune system is strongly restricted by the two unique distinct properties of MHC molecules, i.e., polygenic and polymorphic properties. MHC-I restriction epitope identification has traditionally been accomplished using genetic motif prediction. The study of immune system proteins and their interactions is the main emphasis of the specialist field of immunoproteomics within proteomics. Methodologies include mass spectrometry (MS), SRM assay, MALDI-TOF, Chromatography, ELISA, 2DG PAGE, and bioinformatics tools. Challenges are the complexity of the immune system, protein abundance and dynamics, sample variability, post-translational modifications (PTMs), and data integration. Current advancements are enhanced mass spectrometry techniques, single-cell proteomics, artificial intelligence and machine learning, advanced protein labeling techniques, integration with other omics technologies, and functional proteomics. However, the recently emerging field of immunoproteomics has more promising possibilities in the field of peptide-based vaccines and virus-like particle vaccines. The importance of immunoproteomics technologies and methodologies, as well as their use in the field of vaccinomics, are the main topics of this review. Here, we have discussed immunoproteomics in relation to a step towards the future of vaccination.

Immunohistochemical Expression of Immune Regulatory Proteins in Interface Dermatoses.

Cutaneous immune-related adverse events (irAEs) of immunotherapies, such as anti-programmed cell death protein-1 (PD-1), suggest that immune checkpoint factors may contribute to the pathobiology of lichenoid interface dermatitis in immunotherapy-naïve patients. Our study aimed to describe innate and adaptive immune markers via immunohistochemical (IHC) staining of lichenoid interface dermatoses. We studied the staining patterns of PD-L1, STING, IL-36 gamma, CD8, PD-1, and LAG-3 in five interface dermatoses: oral lichen planus (LP) (n = 10), cutaneous LP (n = 10), chronic cutaneous lupus erythematosus (CLE) (n = 11), erythema multiforme (EM) (n = 11), and toxic epidermal necrolysis (TEN) (n = 13), by immunohistochemistry (IHC) analysis. Expression was evaluated semi-quantitively according to the percentage of keratinocytes and dermal lymphocytes stained compared to keratinocytes and resident pericapillary lymphocytes in normal human skin. All interface dermatoses evaluated showed increased expression of PD-L1 on keratinocytes and LAG-3 in lymphocytes. STING was increased on the keratinocytes of most specimens. Expression of IL-36 gamma, in basal layer keratinocytes was more extensive in oral LP and cutaneous LP and varied in CLE, EM, and TEN. Lymphocytic infiltration expressing PD-1 was elevated in oral LP, cutaneous LP, and CLE. Current thinking is that interface dermatitis is the result of a cell-mediated immune reaction involving cytotoxic CD8+ T-cell-mediated apoptosis of keratinocytes. The findings of this study suggest that in addition to cell-mediated immunity, innate immune factors may contribute to pathobiology.

Antibody blockade of the PSGL-1 immune checkpoint enhances T-cell responses to B-cell lymphoma.

Despite advancements in cancer immunotherapy, most lymphomas remain unresponsive to checkpoint inhibitors. P-selectin glycoprotein ligand-1 (PSGL-1), recently identified as a promoter of T-cell exhaustion in murine melanoma models, has emerged as a novel immune checkpoint protein and promising immunotherapeutic target. In this study, we investigated the potential of PSGL-1 antibody targeting in B-cell lymphoma. Using allogeneic co-culture systems, we demonstrated that targeted antibody interventions against human PSGL-1 enhanced T-cell activation and effector cytokine production in response to lymphoma cells. Moreover, in vitro treatment of primary lymphoma cell suspensions with PSGL-1 antibody resulted in increased activation of autologous lymphoma-infiltrating T cells. Using the A20 syngeneic B-cell lymphoma mouse model, we found that PSGL-1 antibody treatment significantly slowed tumor development and reduced the endpoint tumor burden. This antitumoral effect was accompanied by augmented tumor infiltration of CD4+ and CD8+ T cells and reduced infiltration of regulatory T cells. Finally, anti-PSGL-1 administration enhanced the expansion of CAR T cells previously transferred to mice bearing the aggressive Eμ-Myc lymphoma cells and improved disease control. These results demonstrate that PSGL-1 antibody blockade bolsters T-cell activity against B-cell lymphoma, suggesting a potential novel immunotherapeutic approach for treating these malignancies.

Impairment of spinal CSF flow precedes immune cell infiltration in an active EAE model

Accumulation of immune cells and proteins in the subarachnoid space (SAS) is found during multiple sclerosis and in the animal model experimental autoimmune encephalomyelitis (EAE). Whether the flow of cerebrospinal fluid (CSF) along the SAS of the spinal cord is impacted is yet unknown. Combining intravital near-infrared (NIR) imaging with histopathological analyses, we observed a significantly impaired bulk flow of CSF tracers within the SAS of the spinal cord prior to EAE onset, which persisted until peak stage and was only partially recovered during chronic disease. The impairment of spinal CSF flow coincided with the appearance of fibrin aggregates in the SAS, however, it preceded immune cell infiltration and breakdown of the glia limitans superficialis. Conversely, cranial CSF efflux to cervical lymph nodes was not altered during the disease course. Our study highlights an early and persistent impairment of spinal CSF flow and suggests it as a sensitive imaging biomarker for pathological changes within the leptomeninges.

Toxoplasma gondii ROP5 Enhances Type I IFN Responses by Promoting Ubiquitination of STING.

Toxoplasma gondii is a widely spread opportunistic pathogen that can infect nearly all warm-blooded vertebrates and cause serious toxoplasmosis in immunosuppressed animals and patients. However, the relationship between the host's innate immune system and effector proteins is poorly understood, particularly with regard to how effectors antagonize cGAS-STING signaling during T. gondii infection. In this study, the ROP5 from the PRU strain of T. gondii was found to promote cGAS-STING-mediated immune responses. Mechanistically, ROP5 interacted with STING through predicted domain 2 and modulated cGAS-STING signaling in a predicted domain 3-dependent manner. Additionally, ROP5 strengthened cGAS-STING signaling by enhancing the K63-linked ubiquitination of STING. Consistently, ROP5 deficient PRU (PRUΔROP5) induced fewer type I IFN-related immune responses and replicated faster than the parental strain in RAW264.7 cells. Taken together, this study provides new insights into the mechanism by which ROP5 regulates T. gondii infection and provides new clues for strategies to prevent and control toxoplasmosis.

A percolation-type criticality threshold controls immune protein coating of surfaces.

When a material enters the body, it is immediately attacked by hundreds of proteins, organized into complex networks of binding interactions and reactions. How do such complex systems interact with a material, "deciding" whether to attack? We focus on the "complement" system of ∼40 blood proteins that bind microbes, nanoparticles, and medical devices, initiating inflammation. We show a sharp threshold for complement activation upon varying a fundamental material parameter, the surface density of potential complement attachment points. This sharp threshold manifests at scales spanning single nanoparticles to macroscale pathologies, shown here for diverse engineered and living materials. Computational models show these behaviors arise from a minimal subnetwork of complement, manifesting percolation-type critical transitions in the complement response. This criticality switch explains the "decision" of a complex signaling network to interact with a material, and elucidates the evolution and engineering of materials interacting with the body.