Epigenetic regulation of major histocompatibility complexes in gastrointestinal malignancies and the potential for clinical interception.

BACKGROUND: Gastrointestinal malignancies encompass a diverse group of cancers that pose significant challenges to global health. The major histocompatibility complex (MHC) plays a pivotal role in immune surveillance, orchestrating the recognition and elimination of tumor cells by the immune system. However, the intricate regulation of MHC gene expression is susceptible to dynamic epigenetic modification, which can influence functionality and pathological outcomes. MAIN BODY: By understanding the epigenetic alterations that drive MHC downregulation, insights are gained into the molecular mechanisms underlying immune escape, tumor progression, and immunotherapy resistance. This systematic review examines the current literature on epigenetic mechanisms that contribute to MHC deregulation in esophageal, gastric, pancreatic, hepatic and colorectal malignancies. Potential clinical implications are discussed of targeting aberrant epigenetic modifications to restore MHC expression and 0 the effectiveness of immunotherapeutic interventions. CONCLUSION: The integration of epigenetic-targeted therapies with immunotherapies holds great potential for improving clinical outcomes in patients with gastrointestinal malignancies and represents a compelling avenue for future research and therapeutic development.

NLRC5/MHC class I transactivator: A key target for immune escape by SARS-CoV-2.

Antigen presentation to CD8+ T cells by MHC class I molecules is essential for host defense against viral infections. Various mechanisms have evolved in multiple viruses to escape immune surveillance and defense to support viral proliferation in host cells. Through in vitro SARS-CoV-2 infection studies and analysis of COVID-19 patient samples, we found that SARS-CoV-2 suppresses the induction of the MHC class I pathway by inhibiting the expression and function of NLRC5, a major transcriptional regulator of MHC class I genes. In this review, we discuss the molecular mechanisms for suppression of the MHC class I pathway and clinical implications for COVID-19.

Brucella-driven host N-glycome remodeling controls infection.

Many powerful methods have been employed to elucidate the global transcriptomic, proteomic, or metabolic responses to pathogen-infected host cells. However, the host glycome responses to bacterial infection remain largely unexplored, and hence, our understanding of the molecular mechanisms by which bacterial pathogens manipulate the host glycome to favor infection remains incomplete. Here, we address this gap by performing a systematic analysis of the host glycome during infection by the bacterial pathogen Brucella spp. that cause brucellosis. We discover, surprisingly, that a Brucella effector protein (EP) Rhg1 induces global reprogramming of the host cell N-glycome by interacting with components of the oligosaccharide transferase complex that controls N-linked protein glycosylation, and Rhg1 regulates Brucella replication and tissue colonization in a mouse model of brucellosis, demonstrating that Brucella exploits the EP Rhg1 to reprogram the host N-glycome and promote bacterial intracellular parasitism, thereby providing a paradigm for bacterial control of host cell infection.

The balance between nuclear import and export of NLRC5 regulates MHC class I transactivation.

Major histocompatibility complex (MHC) class I molecules play an essential role in regulating the adaptive immune system by presenting antigens to CD8 T cells. CITA (MHC class I transactivator), also known as NLRC5 (NLR family, CARD domain-containing 5), regulates the expression of MHC class I and essential components involved in the MHC class I antigen presentation pathway. While the critical role of the nuclear distribution of NLRC5 in its transactivation activity has been known, the regulatory mechanism to determine the nuclear localization of NLRC5 remains poorly understood. In this study, a comprehensive analysis of all domains in NLRC5 revealed that the regulatory mechanisms for nuclear import and export of NLRC5 coexist and counterbalance each other. Moreover, GCN5 (general control non-repressed 5 protein), a member of HATs (histone acetyltransferases), was found to be a key player to retain NLRC5 in the nucleus, thereby contributing to the expression of MHC class I. Therefore, the balance between import and export of NLRC5 has emerged as an additional regulatory mechanism for MHC class I transactivation, which would be a potential therapeutic target for the treatment of cancer and virus-infected diseases.

Targeted demethylation and activation of NLRC5 augment cancer immunogenicity through MHC class I.

Impaired expression of MHC (major histocompatibility complex) class I in cancers constitutes a major mechanism of immune evasion. It has been well documented that the low level of MHC class I is associated with poor prognosis and resistance to checkpoint blockade therapies. However, there is lmited approaches to specifically induce MHC class I to date. Here, we show an approach for robust and specific induction of MHC class I by targeting an MHC class I transactivator (CITA)/NLRC5, using a CRISPR/Cas9-based gene-specific system, designated TRED-I (Targeted reactivation and demethylation for MHC-I). The TRED-I system specifically recruits a demethylating enzyme and transcriptional activators on the NLRC5 promoter, driving increased MHC class I antigen presentation and accelerated CD8+ T cell activation. Introduction of the TRED-I system in an animal cancer model exhibited tumor-suppressive effects accompanied with increased infiltration and activation of CD8+ T cells. Moreover, this approach boosted the efficacy of checkpoint blockade therapy using anti-PD1 (programmed cell death protein) antibody. Therefore, targeting NLRC5 by this strategy provides an attractive therapeutic approach for cancer.

FBXO11 constitutes a major negative regulator of MHC class II through ubiquitin-dependent proteasomal degradation of CIITA.

Major histocompatibility complex (MHC) class I and II molecules play critical roles in the activation and regulation of adaptive immunity through antigen presentation to CD8+ and CD4+ T cells, respectively. Strict regulation of MHC expression is critical for proper immune responses. CIITA (MHC class II transactivator), an NLR (nucleotide-binding domain, leucine-rich-repeat containing) protein, is a master regulator of MHC class II (MHC-II) gene transcription. Although it has been known that CIITA activity is regulated at the transcriptional and protein levels, the mechanism to determine CIITA protein level has not been elucidated. Here, we show that FBXO11 is a bona fide E3 ligase of CIITA and regulates CIITA protein level through ubiquitination-mediated degradation. A nonbiased proteomic approach for CIITA-binding protein identified FBXO11, a member of the Skp1-Cullin-1-F-box E3 ligase complex, as a binding partner of CIITA but not MHC class I transactivator, NLRC5. The cycloheximide chase assay showed that the half-life of CIITA is mainly regulated by FBXO11 via the ubiquitin-proteasome system. The expression of FBXO11 led to the reduced MHC-II at the promoter activity level, transcriptional level, and surface expression level through downregulation of CIITA. Moreover, human and mouse FBXO11-deficient cells display increased levels of MHC-II and related genes. In normal and cancer tissues, FBXO11 expression level is negatively correlated with MHC-II. Interestingly, the expression of FBXO11, along with CIITA, is associated with prognosis of cancer patients. Therefore, FBXO11 is a critical regulator to determine the level of MHC-II, and its expression may serve as a biomarker for cancer.

Engineering live attenuated vaccines: Old dogs learning new tricks.

Autoimmune diseases such as rheumatoid arthritis and type 1 diabetes are increasingly common global problems. Concerns about increases in the prevalence of such diseases and the limited efficacy of conventional treatment regimens necessitates new therapies to address these challenges. Autoimmune disease severity and dysbiosis are interconnected. Although probiotics have been established as a therapy to rebalance the microbiome and suppress autoimmune symptoms, these microbes tend to lack a number of advantageous qualities found in non-commensal bacteria. Through attenuation and genetic manipulation, these non-commensal bacteria have been engineered into recombinant forms that offer malleable platforms capable of addressing the immune imbalances found in RA and T1D. Such bacteria have been engineered to express valuable gene products known to suppress autoimmunity such as anti-inflammatory cytokines, autoantigens, and enzymes synthesizing microbial metabolites. This review will highlight current and emerging trends in the field and discuss how they may be used to prevent and control autoimmune diseases.

A metabolically engineered bacterium controls autoimmunity and inflammation by remodeling the pro-inflammatory microenvironment.

Immunotherapy has led to impressive advances in the treatment of autoimmune and pro-inflammatory disorders; yet, its clinical outcomes remain limited by a variety of factors including the pro-inflammatory microenvironment (IME). Discovering effective immunomodulatory agents, and the mechanisms by which they control disease, will lead to innovative strategies for enhancing the effectiveness of current immunotherapeutic approaches. We have metabolically engineered an attenuated bacterial strain (i.e., Brucella melitensis 16M ∆vjbR, Bm∆vjbR::tnaA) to produce indole, a tryptophan metabolite that controls the fate and function of regulatory T (Treg) cells. We demonstrated that treatment with Bm∆vjbR::tnaA polarized macrophages (Mφ) which produced anti-inflammatory cytokines (e.g., IL-10) and promoted Treg function; moreover, when combined with adoptive cell transfer (ACT) of Treg cells, a single treatment with our engineered bacterial strain dramatically reduced the incidence and score of autoimmune arthritis and decreased joint damage. These findings show how a metabolically engineered bacterium can constitute a powerful vehicle for improving the efficacy of immunotherapy, defeating autoimmunity, and reducing inflammation by remodeling the IME and augmenting Treg cell function.

Regulation of MHC Class I Expression in Lung Epithelial Cells during Inflammation.

Lung infections are a perennial leading cause of death worldwide. The lung epithelium comprises three main cell types: alveolar type I (AT1), alveolar type II (AT2), and bronchiolar cells. Constitutively, these three cell types express extremely low amounts of surface MHC class I (MHC I) molecules, that is, <1% of levels found on medullary thymic epithelial cells (ECs). We report that inhalation of the TLR4 ligand LPS upregulates cell surface MHC I by ∼25-fold on the three subtypes of mouse lung ECs. This upregulation is dependent on Nlrc5, Stat1, and Stat2 and caused by a concerted production of the three IFN families. It is nevertheless hampered, particularly in AT1 cells, by the limited expression of genes instrumental in the peptide loading of MHC I molecules. Genes involved in production and response to cytokines and chemokines were selectively induced in AT1 cells. However, discrete gene subsets were selectively downregulated in AT2 or bronchiolar cells following LPS inhalation. Genes downregulated in AT2 cells were linked to cell differentiation and cell proliferation, and those repressed in bronchiolar cells were primarily involved in cilium function. Our study shows a delicate balance between the expression of transcripts maintaining lung epithelium integrity and transcripts involved in Ag presentation in primary lung ECs.

Live attenuated bacterium limits cancer resistance to CAR-T therapy by remodeling the tumor microenvironment.

The tumor microenvironment (TME) is characterized by the activation of immune checkpoints, which limit the ability of immune cells to attack the growing cancer. To overcome immune suppression in the clinic, antigen-expressing viruses and bacteria have been developed to induce antitumor immunity. However, the safety and targeting specificity are the main concerns of using bacteria in clinical practice as antitumor agents. In our previous studies, we have developed an attenuated bacterial strain (Brucella melitensis 16M ∆vjbR, henceforth Bm∆vjbR) for clinical use, which is safe in all tested animal models and has been removed from the select agent list by the Centers for Disease Control and Prevention. In this study, we demonstrated that Bm∆vjbR homed to tumor tissue and improved the TME in a murine model of solid cancer. In addition, live Bm∆vjbR promoted proinflammatory M1 polarization of tumor macrophages and increased the number and activity of CD8+ T cells in the tumor. In a murine colon adenocarcinoma model, when combined with adoptive transfer of tumor-specific carcinoembryonic antigen chimeric antigen receptor CD8+ T cells, tumor cell growth and proliferation was almost completely abrogated, and host survival was 100%. Taken together, these findings demonstrate that the live attenuated bacterial treatment can defeat cancer resistance to chimeric antigen receptor T-cell therapy by remodeling the TME to promote macrophage and T cell-mediated antitumor immunity.

SARS-CoV-2 inhibits induction of the MHC class I pathway by targeting the STAT1-IRF1-NLRC5 axis.

The MHC class I-mediated antigen presentation pathway plays a critical role in antiviral immunity. Here we show that the MHC class I pathway is targeted by SARS-CoV-2. Analysis of the gene expression profile from COVID-19 patients as well as SARS-CoV-2 infected epithelial cell lines reveals that the induction of the MHC class I pathway is inhibited by SARS-CoV-2 infection. We show that NLRC5, an MHC class I transactivator, is suppressed both transcriptionally and functionally by the SARS-CoV-2 ORF6 protein, providing a mechanistic link. SARS-CoV-2 ORF6 hampers type II interferon-mediated STAT1 signaling, resulting in diminished upregulation of NLRC5 and IRF1 gene expression. Moreover, SARS-CoV-2 ORF6 inhibits NLRC5 function via blocking karyopherin complex-dependent nuclear import of NLRC5. Collectively, our study uncovers an immune evasion mechanism of SARS-CoV-2 that targets the function of key MHC class I transcriptional regulators, STAT1-IRF1-NLRC5.

NLRC5/CITA expression correlates with efficient response to checkpoint blockade immunotherapy.

Checkpoint blockade-mediated immunotherapy is emerging as an effective treatment modality for multiple cancer types. However, cancer cells frequently evade the immune system, compromising the effectiveness of immunotherapy. It is crucial to develop screening methods to identify the patients who would most benefit from these therapies because of the risk of the side effects and the high cost of treatment. Here we show that expression of the MHC class I transactivator (CITA), NLRC5, is important for efficient responses to anti-CTLA-4 and anti-PD1 checkpoint blockade therapies. Melanoma tumors derived from patients responding to immunotherapy exhibited significantly higher expression of NLRC5 and MHC class I-related genes compared to non-responding patients. In addition, multivariate analysis that included the number of tumor-associated non-synonymous mutations, predicted neo-antigen load and PD-L2 expression was capable of further stratifying responders and non-responders to anti-CTLA4 therapy. Moreover, expression or methylation of NLRC5 together with total somatic mutation number were significantly correlated with increased patient survival. These results suggest that NLRC5 tumor expression, alone or together with tumor mutation load constitutes a valuable predictive biomarker for both prognosis and response to anti-CTLA-4 and potentially anti-PD1 blockade immunotherapy in melanoma patients.

MHC class I transactivator NLRC5 in host immunity, cancer and beyond.

The presentation of antigenic peptides by major histocompatibility complex (MHC) class I and class II molecules is crucial for activation of the adaptive immune system. The nucleotide-binding domain and leucine-rich repeat receptor family members CIITA and NLRC5 function as the major transcriptional activators of MHC class II and class I gene expression, respectively. Since the identification of NLRC5 as the master regulator of MHC class I and class-I-related genes, there have been major advances in understanding the function of NLRC5 in infectious diseases and cancer. Here, we discuss the biological significance and mechanism of NLRC5-dependent MHC class I expression.

IFN-λ Enhances Constitutive Expression of MHC Class I Molecules on Thymic Epithelial Cells.

Regulation of MHC class I (MHC I) expression has been studied almost exclusively in hematolymphoid cells. We report that thymic epithelial cells (TECs), particularly the medullary TECs, constitutively express up to 100-fold more cell surface MHC I proteins than epithelial cells (ECs) from the skin, colon, and lung. Differential abundance of cell surface MHC I in primary ECs is regulated via transcription of MHC I and of genes implicated in the generation of MHC I-binding peptides. Superior MHC I expression in TECs is unaffected by deletion of Ifnar1 or Ifngr1, but is lessened by deletion of Aire, Ifnlr1, Stat1, or Nlrc5, and is driven mainly by type III IFN produced by medullary TECs. Ifnlr1 -/- mice show impaired negative selection of CD8 thymocytes and, at 9 mo of age, present autoimmune manifestations. Our study shows unanticipated variation in MHC I expression by ECs from various sites and provides compelling evidence that superior expression of MHC I in TECs is crucial for proper thymocyte education.

Class I transactivator, NLRC5: a central player in the MHC class I pathway and cancer immune surveillance.

Major histocompatibility complex (MHC) class I and class II molecules play critical roles in the activation of the adaptive immune system by presenting antigens to CD8+ and CD4+ T cells, respectively. Although it has been well known that CIITA (MHC class II transactivator), an NLR (nucleotide-binding domain, leucine-rich-repeat containing) protein, as a master regulator of MHC class II gene expression, the mechanism of MHC class I gene transactivation was unclear. Recently, another NLR protein, NLRC5 (NLR family, CARD domain-containing 5), was identified as an MHC class I transactivator (CITA). NLRC5 is a critical regulator for the transcriptional activation of MHC class I genes and other genes involved in the MHC class I antigen presentation pathway. CITA/NLRC5 plays a crucial role in human cancer immunity through the recruitment and activation of tumor killing CD8+ T cells. Here, we discuss the molecular function and mechanism of CITA/NLRC5 in the MHC class I pathway and its role in cancer.

The IL-33-PIN1-IRAK-M axis is critical for type 2 immunity in IL-33-induced allergic airway inflammation.

Interleukin 33 (IL-33) is among the earliest-released cytokines in response to allergens that orchestrate type 2 immunity. The prolyl cis-trans isomerase PIN1 is known to induce cytokines for eosinophil survival and activation by stabilizing cytokines mRNAs, but the function of PIN1 in upstream signaling pathways in asthma is unknown. Here we show that interleukin receptor associated kinase M (IRAK-M) is a PIN1 target critical for IL-33 signaling in allergic asthma. NMR analysis and docking simulations suggest that PIN1 might regulate IRAK-M conformation and function in IL-33 signaling. Upon IL-33-induced airway inflammation, PIN1 is activated for binding with and isomerization of IRAK-M, resulting in IRAK-M nuclear translocation and induction of selected proinflammatory genes in dendritic cells. Thus, the IL-33-PIN1-IRAK-M is an axis critical for dendritic cell activation, type 2 immunity and IL-33 induced airway inflammation.

Quiescent Tissue Stem Cells Evade Immune Surveillance.

Stem cells are critical for the maintenance of many tissues, but whether their integrity is maintained in the face of immunity is unclear. Here we found that cycling epithelial stem cells, including Lgr5+ intestinal stem cells, as well as ovary and mammary stem cells, were eliminated by activated T cells, but quiescent stem cells in the hair follicle and muscle were resistant to T cell killing. Immune evasion was an intrinsic property of the quiescent stem cells resulting from systemic downregulation of the antigen presentation machinery, including MHC class I and TAP proteins, and is mediated by the transactivator NLRC5. This process was reversed upon stem cell entry into the cell cycle. These studies identify a link between stem cell quiescence, antigen presentation, and immune evasion. As cancer-initiating cells can derive from stem cells, these findings may help explain how the earliest cancer cells evade immune surveillance.