Nucleotide Oligomerization Domain Research Articles

Ischemia/reperfusion injured intestinal epithelial cells cause cortical neuron death by releasing exosomal microRNAs associated with apoptosis, necroptosis, and pyroptosis

To date, there is no good evidence that intestine epithelial cells (IEC) affected by ischemia/reperfusion (I/R) injury are able to cause cortical neuron injury directly. Additionally, it remains unclear whether the neuronal damage caused by I/R injured IEC can be affected by therapeutic hypothermia (TH, 32 °C). To address these questions, we performed an oxygen–glucose deprivation (OGD) affected IEC-6-primary cortical neuron coculture system under normothermia (37 °C) or TH (32 °C) conditions. It was found that OGD caused hyperpermeability in IEC-6 cell monolayers. OGD-preconditioned IEC-6 cells caused cortical neuronal death (e.g., decreased cell viability), synaptotoxicity, and neuronal apoptosis (evidenced by increased caspase-3 expression and the number of TUNEL-positive cells), necroptosis (evidenced by increased receptor-interacting serine/threonine-protein kinase-1 [RIPK1], RIPK3 and mixed lineage kinase domain-like pseudokinase [MLKL] expression), and pyroptosis (evidenced by an increase in caspase-1, gasdermin D [GSDMD], IL-1β, IL-18, the apoptosis-associated speck-like protein containing a caspase recruitment domain [ASC], and nucleotide oligomerization domain [NOD]-like receptor [NLRP]-1 expression). TH did not affect the intestinal epithelial hyperpermeability but did attenuate OGD-induced neuronal death and synaptotoxicity. We also performed quantitative real-time PCR to quantify the genes encoding 84 exosomal microRNAs in the medium of the control-IEC-6, the control-neuron, the OGD-IEC-6 at 37 °C, the OGD-IEC-6 at 32 °C, the neuron cocultured with OGD-IEC-6 at 37 °C, and the neurons cocultured with OGD-IEC-6 at 32 °C. We found that the control IEC-6 cell s or cortical neurons are able to secrete a basal level of exosomal miRNAs in their medium. OGD significantly up-regulated the basal level of each parameter for IEC-6 cells. As compared to those of the OGD-IEC-6 cells or the control neurons, the OGD-IEC-6 cocultured neurons had significantly higher levels of 19 exosomal miRNAs related to apoptosis, necroptosis, and/or pyroptosis events. Our results identify that I/R injured intestinal epithelium cells can induce cortical neuron death via releasing paracrine mediators such as exosomal miRNAs associated with apoptosis, necroptosis, and/or pyroptosis, which can be counteracted by TH.

The Role of the Bacterial Muramyl Dipeptide in the Regulation of GLP-1 and Glycemia.

The host’s intestinal microbiota contributes to endocrine and metabolic responses, but a dysbiosis in this environment can lead to obesity and insulin resistance. Recent work has demonstrated a role for microbial metabolites in the regulation of gut hormones, including the metabolic hormone, glucagon-like peptide-1 (GLP-1). Muramyl dipeptide (MDP) is a bacterial cell wall component which has been shown to improve insulin sensitivity and glucose tolerance in diet-induced obese mice by acting through the nucleotide oligomerization domain 2 (NOD2) receptor. The purpose of this study was to understand the effects of MDP on GLP-1 secretion and glucose regulation. We hypothesized that MDP enhances glucose tolerance by inducing intestinal GLP-1 secretion through NOD2 activation. First, we observed a significant increase in GLP-1 secretion when murine and human L-cells were treated with a fatty acid MDP derivative (L18-MDP). Importantly, we demonstrated the expression of the NOD2 receptor in mouse intestine and in L-cells. In mice, two intraperitoneal injections of MDP (5 mg/kg body weight) caused a significant increase in fasting total GLP-1 in chow-fed mice, however this did not lead to an improvement in oral glucose tolerance. When mice were exposed to a high-fat diet, they eventually lost this MDP-induced GLP-1 release. Finally, we demonstrated in L-cells that hyperglycemic conditions reduce the mRNA expression of NOD2 and GLP-1. Together these findings suggest MDP may play a role in enhancing GLP-1 during normal glycemic conditions but loses its ability to do so in hyperglycemia.

Amyloid Signaling in Filamentous Fungi and Bacteria.

Amyloids are implicated in many protein misfolding diseases. Amyloid folds, however, also display a range of functional roles particularly in the microbial world. The templating ability of these folds endows them with specific properties allowing their self-propagation and protein-to-protein transmission in vivo. This property, the prion principle, is exploited by specific signaling pathways that use transmission of the amyloid fold as a way to convey information from a receptor to an effector protein. I describe here amyloid signaling pathways involving fungal nucleotide binding and oligomerization domain (NOD)-like receptors that were found to control nonself recognition and programmed cell death processes. Studies on these fungal amyloid signaling motifs stem from the characterization of the fungal [Het-s] prion protein and have led to the identification in fungi but also in multicellular bacteria of several distinct families of signaling motifs, one of which is related to RHIM [receptor-interacting protein (RIP) homotypic interaction motif], an amyloid motif regulating mammalian necroptosis.

NLRP12 controls arthritis severity by acting as a checkpoint inhibitor of Th17 cell differentiation.

Nucleotide oligomerization domain (NOD)-like receptor-12 (NLRP12) has emerged as a negative regulator of inflammation. It is well described that the Th17 cell population increases in patients with early Rheumatoid Arthritis (RA), which correlates with the disease activity. Here, we investigated the role of NLRP12 in the differentiation of Th17 cells and the development of experimental arthritis, using the antigen-induced arthritis (AIA) murine model. We found that Nlrp12-/- mice develop severe arthritis characterized by an exacerbated Th17-mediated inflammatory response with increases in the articular hyperalgesia, knee joint swelling, and neutrophil infiltration. Adoptive transfer of Nlrp12-/- cells into WT mice recapitulated the hyperinflammatory response seen in Nlrp12-/- mice and the treatment with anti-IL-17A neutralizing antibody abrogated arthritis development in Nlrp12-/- mice, suggesting that NLRP12 works as an inhibitor of Th17 cell differentiation. Indeed, Th17 cell differentiation markedly increases in Nlrp12-/- T cells cultured under the Th17-skewing condition. Mechanistically, we found that NLRP12 negatively regulates IL-6-induced phosphorylation of STAT3 in T cells. Finally, pharmacological inhibition of STAT3 reduced Th17 cell differentiation and abrogated hyperinflammatory arthritis observed in Nlrp12-/- mice. Thus, we described a novel role for NLRP12 as a checkpoint inhibitor of Th17 cell differentiation, which controls the severity of experimental arthritis.

MCC950 Inhibits NLRP3 Inflammasome and Alleviates Axonal Injures in Early Stages of Diffuse Axonal Injury in Rats.

Increasing evidence has revealed that neuroinflammation plays a pivotal role in axonal injures. Nucleotide oligomerization domain (NOD)-like receptor protein (NLRP3) inflammasome is reported to be widely involved with the pathology of central nervous system disorders. But the role of NLRP3 in diffuse axonal injury (DAI) are rarely reported. The purpose of this study was to investigate the expression of NLRP3 after diffuse axonal injury and the role of NLRP3 in axonal injures. The lateral head rotation device was used to establish DAI model of rats. Immunohistochemical staining for β-amyloid precursor protein and Bielschowsky silver staining were used to assess axonal injures and axonal loss. Terminal Deoxynucleotidyl Transferase-Mediated Digoxigenin-dUTP-Biotin Nick-End Labelling Assay was used to detect cell apoptosis. Brain water content was used to assess cerebral edema and the modified Neurologic Severity Score was used to assess the neurological deficits. Components of NLRP3 inflammasome, such as NLRP3, apoptosis-associated speck-like (ASC) adapter protein and caspase-1, and pro-inflammatory cytokines, for example IL-18 and IL-1β, were over-expressed in early stages of DAI. MCC950, a selective small-molecule inhibitor of NLRP3 inflammasome, inhibited the over-expression of NLRP3 inflammasome and pro-inflammatory cytokines after DAI. MCC950 alleviated axonal injures and cell apoptosis. MCC950 also decreased brain water content and alleviated neurologic deficits 1day and 3days after DAI but not 7days after DAI. These results suggest that MCC950 treatment in the early stages of DAI has a time limiting effect in preventing from axonal injuries and neurological deficits, and that NLRP3 inflammasome plays an important role in axonal injures and may be a potential candidate for axonal injures following DAI.

A Novel X-Linked Inhibitor of Apoptosis Deficient Variant Showing Attenuated Epstein-Barr Virus Response.

We report on 2 Asian siblings with X-linked inhibitor of apoptosis deficiency that arose from a novel deletion that presented with Epstein-Barr virus disease and hemophagocytic lymphohistiocytosis. This disease is ascribed to dysfunction in the nucleotide binding and oligomerization domain receptor pathway, tested using a modified muramyl dipeptide-mediated assay.

Spinal cord NLRP1 inflammasome contributes to dry skin induced chronic itch in mice

BackgroundDry skin itch is one of the most common skin diseases and elderly people are believed to be particularly prone to it. The inflammasome has been suggested to play an important role in chronic inflammatory disorders including inflammatory skin diseases such as psoriasis. However, little is known about the role of NLRP1 inflammasome in dry skin-induced chronic itch.MethodsDry skin-induced chronic itch model was established by acetone-ether-water (AEW) treatment. Spontaneous scratching behavior was recorded by video monitoring. The expression of nucleotide oligomerization domain (NOD)-like receptor protein 1 (NLRP1) inflammasome complexes, transient receptor potential vanilloid type 1 (TRPV1), and the level of inflammatory cytokines were determined by western blot, quantitative real-time PCR, and enzyme-linked immunosorbent assay (ELISA) kits. Nlrp1a knockdown was performed by an adeno-associated virus (AAV) vector containing Nlrp1a-shRNA-eGFP infusion. H.E. staining was used to evaluate skin lesion.ResultsAEW treatment triggers spontaneous scratching and significantly increases the expression of NLRP1, ASC, and caspase-1 and the levels of IL-1β, IL-18, IL-6, and TNF-α in the spinal cord and the skin of mice. Spinal cord Nlrp1a knockdown prevents AEW-induced NLRP1 inflammasome assembly, TRPV1 channel activation, and spontaneous scratching behavior. Capsazepine, a specific antagonist of TRPV1, can also inhibit AEW-induced inflammatory response and scratching behavior. Furthermore, elderly mice and female mice exhibited more significant AEW-induced scratching behavior than young mice and male mice, respectively. Interestingly, AEW-induced increases in the expression of NLRP1 inflammasome complex and the levels of inflammatory cytokines were more remarkable in elderly mice and female mice than in young mice and male mice, respectively.ConclusionsSpinal cord NLRP1 inflammasome-mediated inflammatory response contributes to dry skin-induced chronic itch by TRPV1 channel, and it is also involved in age and sex differences of chronic itch. Inhibition of NLRP1 inflammasome may offer a new therapy for dry skin itch.

Explorations of the Apoptotic Protease Activating Factor 1 (Apaf‐1) in its Formation of the Seven‐Spoked Apoptosome Responsible for Programmed Cell Death

Multicellular life forms first appeared in the fossil record in a burst of evolution termed during the Cambrian period. With multicellularity came the development and specialization of cells, alongside their coordination in the formation of tissues and organs. An adult human, composed of 30–40 trillion cells, progresses through repeated choreographed rounds of growth and cell division from its microscopic begins as a single celled zygote. Some have argued that this is the most critical stage of one’s life. Equally important for development, relatively overlooked, is the intrinsic process of cell death. For example, the webbing between our fingers and toes disappears around week six of gestation. This macroscopic observation during fetal development is the result of a complex series of molecular events initiated by the apoptotic protease activating factor 1 (Apaf‐1) as it forms the holo‐enzyme the apoptosome. The Blue Valley CAPS 2020 SMART Team (Students Modeling a Research Topic), with support from the CBM at MSOE, modeled the Apaf‐1 using JMol and 3D printing technologies in order to better understand how this molecular machine carries out programmed cell death. Structurally, Apaf‐1 is composed of 1249 amino acid residues that form a C‐terminal regulatory domain, a nucleotide binding and oligomerization domain (NOD), and an N‐terminal caspase recruitment domain (CARD). The regulatory domain is composed of two WD40 b‐propellers that are responsible for binding cytochrome c, which itself is released from the inner membrane of the mitochrondria under conditions of stress. Cytochrome c bonding causes a disruption in a salt bridge between Lys192 and Asp616 leading to changes in the orientation of the propellers. The nucleotide binding and oligomerization domain is composed of a number of helical subdomains. Conformational changes in the regulatory domain stimulates nucleotide exchange, where an ADP molecule bond in Apaf‐1’s inactive and closed monomeric state is exchanged for an ATP molecule leading to a more open monomeric state and the subsequent formation of the heptameric apoptosome. The Arg265 trigger which stabilizes an aspartate triad in the inactive monomeric state becomes compromised after ATP binding leading to conformational changes that open up Apaf‐1. Once the polymeric apoptosome forms, the CARD binds and activates procaspase‐9, which itself cleaves loops in procaspases 3 and 7 which become active and responsible of the apoptosomes proteolytic function. Proteolysis ultimately leads to cell death.

Polymorphism nucleotide oligomerization domain-2 (NOD2) in neonatal with early breastfeeding initiation

This study aimed to determine if NOD2 gene polymorphisms are found in neonatal with early breastfeeding initiation and neonatal without early breastfeeding initiation. This study used a Quasy Experiment type, with Non-equivalent Control Group Design. The sample in this study were 60 pregnant women; gestational age±34 and 36 weeks, normal delivery, and carried out early breastfeeding at the Siti Fatima maternal and child hospital and Public Health Center Bara-baraya. The sample was divided into two groups, i.e., the intervention group as many as 30 neonatal and the control group as many as 30 neonatal. Samples were taken from blood from the umbilical cord as much as three cc for the examination of NOD2 polymorphisms. The group that was given early breastfeeding initiation intervention had a polymorphism frequency of 1 person (3.33%), while the control group had a polymorphism frequency of 4 people (6.67%). The control group, i.e., infants who did not get early breastfeeding initiation had a higher frequency of NOD2 gene mutations compared to the group that received early breastfeeding initiation.

Teduglutide Promotes Epithelial Tight Junction Pore Function in Murine Short Bowel Syndrome to Alleviate Intestinal Insufficiency

BackgroundIn short bowel syndrome, epithelial surface loss results in impaired nutrient absorption and may lead to intestinal insufficiency or intestinal failure. Nucleotide oligomerization domain 2 (Nod2) dysfunction predisposes to the development of intestinal failure after intestinal resection and is associated with intestinal barrier defects. Epithelial barrier function is crucial for intestinal absorption and for intestinal adaptation in the short bowel situation.AimsThe aim of the study was to characterize the effects of the GLP-2 analogue Teduglutide in the small intestine in the presence and absence of Nod2 in a mouse model of short bowel syndrome.MethodsMice underwent 40% ICR and were thereafter treated with Teduglutide versus vehicle injections. Survival, body weight, stool water, and sodium content and plasma aldosterone concentrations were determined. Intestinal and kidney tissue was examined with light and fluorescence microscopy, Ussing chamber studies and quantitative PCR in wild type and transgenic mice.ResultsTeduglutide reduced intestinal failure incidence in Nod2 k.o. mice. In wt mice, Teduglutide attenuated intestinal insufficiency as indicated by reduced body weight loss and lower plasma aldosterone concentrations, lower stool water content, and lower stool sodium losses. Teduglutide treatment was associated with enhanced epithelial paracellular pore function and enhanced claudin-10 expression in tight junctions in the villus tips, where it colocalized with sodium–glucose cotransporter 1 (SGLT-1), which mediates Na-coupled glucose transport.ConclusionsIn the SBS situation, Teduglutide not only maximizes small intestinal mucosal hypertrophy but also partially restores small intestinal epithelial function through an altered distribution of claudin-10, facilitating sodium recirculation for Na-coupled glucose transport and water absorption.

Activation of the pattern recognition receptor NOD1 augments colon cancer metastasis

While emerging data suggest nucleotide oligomerization domain receptor 1 (NOD1), a cytoplasmic pattern recognition receptor, may play an important and complementary role in the immune response to bacterial infection, its role in cancer metastasis is entirely unknown. Hence, we sought to determine the effects of NOD1 on metastasis. NOD1 expression in paired human primary colon cancer, human and murine colon cancer cells were determined using immunohistochemistry and immunoblotting (WB). Clinical significance of NOD1 was assessed using TCGA survival data. A series of in vitro and in vivo functional assays, including adhesion, migration, and metastasis, was conducted to assess the effect of NOD1. C12-iE-DAP, a highly selective NOD1 ligand derived from gram-negative bacteria, was used to activate NOD1. ML130, a specific NOD1 inhibitor, was used to block C12-iE-DAP stimulation. Stable knockdown (KD) of NOD1 in human colon cancer cells (HT29) was constructed with shRNA lentiviral transduction and the functional assays were thus repeated. Lastly, the predominant signaling pathway of NOD1-activation was identified using WB and functional assays in the presence of specific kinase inhibitors. Our data demonstrate that NOD1 is highly expressed in human colorectal cancer (CRC) and human and murine CRC cell lines. Clinically, we demonstrate that this increased NOD1 expression negatively impacts survival in patients with CRC. Subsequently, we identify NOD1 activation by C12-iE-DAP augments CRC cell adhesion, migration and metastasis. These effects are predominantly mediated via the p38 mitogen activated protein kinase (MAPK) pathway. This is the first study implicating NOD1 in cancer metastasis, and thus identifying this receptor as a putative therapeutic target.

Overexpression of MicroRNA-9a-5p Ameliorates NLRP1 Inflammasome-mediated Ischemic Injury in Rats Following Ischemic Stroke

The nucleotide oligomerization domain (NOD)-like receptor (NLR) pyrin domain-containing protein 1 (NLRP1) inflammasome has been shown to contribute to brain injury after ischemic stroke. Our previous study showed that microRNA-9a-5p (miR-9a-5p) ameliorates ischemic injury by regulating neuronal autophagy in rats subjected to middle cerebral artery occlusion (MCAO) surgery. The aims of this study were to investigate whether miR-9a-5p can influence the NLRP1 inflammasome following ischemic stroke and to clarify the mechanism involved. We found that MCAO in rats increased the level of NLRP1 inflammasome proteins, including NLRP1 receptor, ASC and precursor caspase-1, which induced higher levels of cleaved caspase-1, mature interleukin-1β (IL-1β) and interleukin-18 (IL-18). Similarly, the levels of the NLRP1 inflammasome proteins, cleaved caspase-1, mature IL-1β and IL-18 were elevated in SY-5Y cells exposed to oxygen-glucose deprivation (OGD). Further investigation showed that NLRP1 was a target of miR-9a-5p and was downregulated by miR-9a-5p overexpression and upregulated by miR-9a-5p inhibition. Moreover, overexpression of miR-9a-5p not only decreased the levels of NLRP1, ASC and precursor caspase-1 but also reduced the levels of IL-1β and IL-18 in MCAO rats and OGD cells. Therefore, we conclude that miR-9a-5p is involved in NLRP1 inflammasome-mediated ischemic injury, which further suggests that the overexpression of miR-9 may be an effective way to ameliorate brain injury following ischemic stroke.

Elevated expression of IL-18 but not IL-1β gene is associated with NALP3 and AIM2 inflammasome in Polycystic Ovary Syndrome

Inflammasome complex mediated interleukin 1β (IL-1β) and interleukin 18 (IL-18) production may be involved in immunopathogenesis of polycystic ovary syndrome (PCOS). Therefore, this study was conducted to investigate involved inflammasome pathways in PCOS. Therefore, inflammasome genes expression and serum level of IL-1β were evaluated in 30 patients with confirmed PCOS and 30 women without PCOS. A remarkable increase in expression of the nucleotide binding and oligomerization domain (NOD)-like receptor (NLR) family pyrin domain-containing 3 (NALP3), absent in melanoma 2 (AIM2), IL-18 and associated speck-like protein containing a caspase recruitment domain (CARD); (ASC) genes in PCOS were observed (p < 0.05). In contrast, expression level of NALP1, NALP12, NLR family apoptosis inhibitory proteins (NAIP), NLR family caspase recruitment domain (CARD) domain containing 4 (NLRC4) and IL-1β genes was not significant. Although the IL-1β protein level in serum of COS patients with BMI ≥ 25 was significantly higher than PCOS patient with BMI < 25, but there was no significant difference in non-PCOS individuals with BMI < 25 or ≥25. Furthermore, significant correlation between expression of AIM2 (r = 0.83, p = 0.032) and NALP3 (r = 0.59, p = 0.0001) was observed with IL-18, while a positive correlation (r = 0.84, p = 0.0001) was revealed between NAIP and IL-1β. Based on the obtained results on inflammasome components along with increased expression of IL-1β especially in overweight patients, it can be concluded that IL-18 expression as well as IL-1β is probably due to activation of AIM2, NALP3 or NAIP inflammasome, which may play a critical role in immunopathology of PCOS.

Functional characterization of NLRX1 in rabbit during enterohemorrhagic Escherichia coli infection

Nucleotide oligomerization domain (NOD) like receptor X1 (NLRX1) is a member of pattern recognition receptor, which has been linked to viral response, cancer, and inflammatory diseases. In this study, rabbit NLRX1 (rNLRX1) was firstly cloned from RK-13 cells, which protein contained a NACHT domain and seven LRRs. rNLRX1 was widely expressed in tissues of rabbits, and highly increased in liver, spleen, kidney, and colon after infected with enterohemorrhagic Escherichia coli (EHEC). Overexpression of rNLRX1 negatively regulated NF-κB signaling, and impaired the expression of pro-inflammatory cytokines and defensins. Moreover, deficient of rNLRX1 in RK-13 cells was performed to investigate the possible roles of rNLRX1. Upon EHEC stimulation, knockdown of rNLRX1 markedly enhanced NF-κB activation and downstream responsive cytokines (IL1β and TNFα) and β-defensins (DEFB114, DEFB124, and DEFB125). Furthermore, overexpression of rNLRX1 promoted the proliferation of EHEC, whereas knockdown of rNLRX1 inhibited its growth. Our study identified that rNLRX1 acts as a negative regulatory in anti-microbial responses after EHEC infection.

Innate signaling within the central nervous system recruits protective neutrophils

There is great interest in understanding how the central nervous system (CNS) communicates with the immune system for recruitment of protective responses. Infiltrating phagocytic monocytes and granulocytes are implicated in neuroinflammation in multiple sclerosis and its animal model experimental autoimmune encephalomyelitis (EAE). To investigate how CNS endogenous signals can be harnessed to promote anti-inflammatory programs, we have used a particulate Toll-like receptor 9 and nucleotide-oligomerization domain 2 bispecific innate ligand (MIS416), to address whether its phagocytosis within the CNS recruits protective myeloid cells. We find that MIS416 injected intrathecally into the cerebrospinal fluid via the cisterna magna induced a local chemokine response that recruited blood-derived monocytes and neutrophils to the CNS. These cells phagocytosed MIS416. The increase in EAE severity normally seen from time of onset did not occur in mice receiving MIS416. This suppression of disease symptoms was dependent on expression of the type I interferon receptor (IFNAR). Transfer of intrathecal MIS416-induced neutrophils suppressed EAE in recipient mice, while monocytes did not transfer protection. MIS416-induced neutrophils showed increased IL-10 expression that was IFNAR1-driven. In contrast to intrathecal administration, intravenous administration of MIS416 led to monocyte but not neutrophil infiltration to the CNS. We thus identify a CNS-intrinsic and -specific phagocytosis-induced recruitment of anti-inflammatory neutrophils that contribute to CNS homeostasis and may have therapeutic potential.

Inferring RPW8-NLRs's evolution patterns in seed plants: case study in Vitis vinifera.

Genomic and transcriptomic studies in plants and, more in deep, in grapevine reveal that the disease-resistance RNL gene family is highly variable. RNLs (RPW8-NLRs) are a phylogenetically distinct class of nucleotide oligomerization domain (NOD)-like receptors (NLRs) identified in plants. Two RNLs, namely, the NRG1 (N Requirement Gene 1) and the ADR1 (Activated Disease Resistance 1), have been characterized; however, little is known about the RNL evolutionary history in higher plants. To trace the diversification of RNL gene subfamily, we scanned the NLR proteins of 73 plant genomes belonging to 29 taxa, revealing a noticeable diversification across species and within the same genus or botanic family together with a conspicuous expansion in important crop species. To explore the RNL variability in Vitis vinifera and gain information with respect to their structure, evolutionary diversification of five grape genomes ('Aglianico', 'Falanghina', 'Sultanina', 'Tannat', and 'Nebbiolo') has been compared to the reference genome ('Pinot Noir'). The number of RNLs ranged from 6 ('Sultanina') to 14 ('Nebbiolo'), in contrast to the 10 'Pinot Noir' RNLs. The phylogenetic study on grapevine RNLs revealed that all collapsed into NRG1-clade, rather than four. To investigate more in depth the means of intraspecific variability of grape RNL copies, a transcriptomic profiling in response to powdery mildew (PM) infection was carried out through qRT-PCRs and public databases interrogation. The RNL expression variability identified in transcriptome data sets supports the hypothesis of a functional expansion/contraction in grapevine varieties. Although no direct correlations between grapevine PM-resistance and RNL expression was identified, our work can provide good candidates for functional studies able to elucidate the putative "helper" role of RNLs in grape immune signalling.

The role of NOD2/CARD15 gene of antigen-presenting system in functioning of various organs and systems

The article presents the literature review devoted to NOD2/CARD15 gene. Genetic variability affects the susceptibility and development of certain human diseases such as autoimmune diseases and infections, affecting numerous cellular processes, and thus modulating the response to environmental and internal factors. The NOD2/CARD15 gene plays a major role in the development and course of various diseases such as Grohn's disease, Blau syndrome, as well as the risk of developing severe complications of the graft versus host reaction after allogeneic stem cell transplantation. NOD (Nucleotide Oligomerization Domain) is the domain of nucleotide oligomerization. NOD-like receptors play an important regulatory role in the response on infectious agents and at activation of the adaptive immune response. It is known that the mechanism of action of NOD-like receptors is based on the response to the pathogen of associated molecular patterns mainly of bacterial origin, which leads to the formation and activation of inflammasome. Recently, another NOD-like receptor activation mechanism has been revealed that provides innate virus recognition. The review presents Toll-like receptors, which are part of the innate immune system. Innate immunity is an inherited system of protection of the body against pathogenic and non-pathogenic microorganisms. The mechanisms of innate immunity develop very quickly. In newborns, the immune system is mainly dependent on components of the innate or antigen-independent immune system including phagocytes, natural killer cells, antigen-presenting cells, humoral inflammatory mediators and complement system.

Inflammatory Responses of Porcine MoDC and Intestinal Epithelial Cells in a Direct-Contact Co-culture System Following a Bacterial Challenge.

Intestinal epithelial cells (IEC) and immune cells, such as dendritic cells (DC), jointly control the immune response towards luminal pathogens in the intestinal mucosa. Crosstalk between IEC and DC is crucial for coordinating immune responses and occurs via soluble factors and direct cell-cell contacts. The present study aimed at establishing a direct-contact co-culture model of porcine IEC and DC to mimic these interactions. The effects of (1) co-cultivation of the two cell types and (2) bacterial infection on the inflammatory response patterns of each of the cell types were determined with a special focus on the canonical and non-canonical inflammasome signaling pathways. In infection experiments, in vitro cultures were exposed to either the probiotic Enterococcus (E.) faecium NCIMB 10415 or enterotoxigenic Escherichia coli (ETEC). In porcine IEC (IPEC-J2), co-cultivation with porcine monocyte-derived DC (MoDC) resulted in reduced basal NLRP3 (nucleotide oligomerization domain [NOD]-like receptor [NLR] family, pyrin domain containing 3) inflammasome mRNA levels in unstimulated conditions. In porcine MoDC, the presence of IPEC-J2 cells evoked a noticeable decrease of interleukin (IL)-8 and transforming growth factor-β (TGF-β) mRNA and protein expression. ETEC, in contrast to E. faecium, modulated the inflammasome pathway in IPEC-J2 cells and porcine MoDC. Co-cultured IPEC-J2 cells showed an augmented inflammasome response to ETEC infection. By contrast, MoDC revealed a weakened ETEC response under such co-culture conditions as indicated by a reduction of inflammasome-related IL-1β protein release. Our data indicate that the close contact between IEC and resident immune cells has a major effect on their immunological behavior.

Palmitoylation of NOD1 and NOD2 is required for bacterial sensing.

The nucleotide oligomerization domain (NOD)-like receptors 1 and 2 (NOD1/2) are intracellular pattern-recognition proteins that activate immune signaling pathways in response to peptidoglycans associated with microorganisms. Recruitment to bacteria-containing endosomes and other intracellular membranes is required for NOD1/2 signaling, and NOD1/2 mutations that disrupt membrane localization are associated with inflammatory bowel disease and other inflammatory conditions. However, little is known about this recruitment process. We found that NOD1/2 S-palmitoylation is required for membrane recruitment and immune signaling. ZDHHC5 was identified as the palmitoyltransferase responsible for this critical posttranslational modification, and several disease-associated mutations in NOD2 were found to be associated with defective S-palmitoylation. Thus, ZDHHC5-mediated S-palmitoylation of NOD1/2 is critical for their ability to respond to peptidoglycans and to mount an effective immune response.

NODs require S-palmitoylation to signal

Innate Immunity The compartmentalization of proteins within the cell is essential for their function. The addition of lipid molecules redistributes proteins to the cell surface or to membrane-bound organelles. Working in transgenic mice and in tissue cultured cells, Lu et al. found that nucleotide oligomerization domain–like receptors 1 and 2 (NOD1 and NOD2), two proteins responsible for detecting bacterial products, required lipid modifications for their recruitment to the cell membrane and function. The specific modification, palmitoylation at a cysteine thiol, was mediated by the enzyme ZDHHC5. Loss of ZDHHC5 or removal of key modification residues in NOD1 and NOD2 abolished their function, compromising antibacterial responses. Human variants of NOD2 display altered palmitoylation, which could help to explain many inflammatory conditions, such as irritable bowel syndrome. Science , this issue p. [460][1] [1]: /lookup/doi/10.1126/science.aau6391