Helicobacter pylori exemplifies one of the most favourable bacterial pathogens worldwide. The bacterium colonizes the gastric mucosa in about half of the human population and constitutes a major risk factor for triggering gastric diseases such as stomach cancer. H. pylori infection represents a prime example of chronic inflammation and cancer-inducing bacterial pathogens. The microbe utilizes a remarkable set of virulence factors and strategies to control cellular checkpoints of inflammation and oncogenic signal transduction. This chapter emphasizes on the pathogenicity determinants of H. pylori such as the cytotoxin-associated genes pathogenicity island (cagPAI)-encoded type-IV secretion system (T4SS), effector protein CagA, lipopolysaccharide (LPS) metabolite ADP-glycero-β-D-manno-heptose (ADP-heptose), cytotoxin VacA, serine protease HtrA, and urease, and how they manipulate various key host cell signaling networks in the gastric epithelium. In particular, we highlight the H. pylori-induced disruption of cell-to-cell junctions, pro-inflammatory activities, as well as proliferative, pro-apoptotic and anti-apoptotic responses. Here we review these hijacked signal transduction events and their impact on gastric disease development.

The atypical protein kinase ALPK1 is activated by the bacterial nucleotide sugar ADP-heptose and phosphorylates TIFA to switch on a signaling pathway that combats microbial infection. In contrast, ALPK1 mutations cause two human diseases: the ALPK1[T237M] and ALPK1[Y254C] mutations underlie ROSAH syndrome (retinal dystrophy, optic nerve oedema, splenomegaly, anhidrosis, and migraine headache), while the ALPK1[V1092A] mutation accounts for 45% of spiradenoma and 30% of spiradenocarcinoma cases studied. In this study, we demonstrate that unlike wild-type (WT) ALPK1, the disease-causing ALPK1 mutants trigger the TIFA-dependent activation of an NF-κB/activator protein 1 reporter gene in the absence of ADP-heptose, which can be suppressed by either of two additional mutations in the ADP-heptose binding site that prevent the activation of WT ALPK1 by ADP-heptose. These observations are explained by our key finding that although ALPK1[T237M] and ALPK1[V1092A] are activated by bacterial ADP-heptose, they can also be activated by nucleotide sugars present in human cells (UDP-mannose, ADP-ribose, and cyclic ADP-ribose) which can be prevented by disruption of the ADP-heptose binding site. The ALPK1[V1092A] mutant was also activated by GDP-mannose, which did not activate ALPK1[T237M]. These are new examples of disease-causing mutations permitting the allosteric activation of an enzyme by endogenous molecules that the WT enzyme does not respond to. We propose that the loss of the specificity of ALPK1 for bacterial ADP-heptose underlies ROSAH syndrome and spiradenoma/spiradenocarcinoma caused by ALPK1 mutation.

BACKGROUND: Scalp psoriasis seriously affects the appearance and psychological status of patients. The aim of this study was to investigate the effect and potential mechanism of RPL9 and TIFA in scalp psoriasis, so as to provide a precise and effective way for the clinical treatment of scalp psoriasis.
METHODS: The Gene Expression Omnibus (GEO) database was employed to download the GSE75343 dataset to search for differentially expressed genes (DEGs) in scalp psoriasis through Sangerbox. Gene set enrichment analysis (GSEA) and gene set variation analysis (GSVA) enrichment analysis, functional enrichment analysis, immune cell infiltration analysis, immune responses and correlation analysis with 12 hub genes were performed. Then, STRING was used to develop a protein-protein interaction (PPI) network, used Cytoscape to locate hub genes, and SVM-RFE and random forest were utilized to identified RPL9 as the targeted gene. TIFA-RPL9 interaction predictions were made viathe Open Targets Platform and Uniprot. Further, the RPL9 and TIFA expression, molecular mechanism, and function were assessed in scalp psoriasis.
RESULTS: Immunohistochemistry, qPCR, and western blotting verified that RPL9 and TIFA were highly expressed in lesional tissues of scalp psoriasis and IL17A-stimulated HaCaT cells. RPL9 knockdown effectively suppressed the proliferative capacity of IL17A-stimulated HaCaT cells in the CCK8 assay. The co-immunoprecipitation results revealed that RPL9 could interact with TIFA in IL17A-stimulated HaCaT cells. In qPCR and western blotting, RPL9 knockdown significantly inhibited TIFA at the mRNA and protein levels in IL17A-stimulated HaCaT cells. In ELISA, the secretion of TNF-α was markedly inhibited after downregulating RPL9 in IL17A-stimulated HaCaT cells.
CONCLUSIONS: To our knowledge, we have elucidated the expression and role of RPL9 and TIFA in scalp psoriatic skin and keratinocytes, and our findings confirm that RPL9 might act as a candidate therapeutic target for scalp psoriasis.

Due to the global crisis caused by the dramatic rise of drug resistance among Gram-negative bacteria, there is an urgent need for a thorough understanding of the pathogenesis of infections of such an etiology. In light of the limited availability of new antibiotics, therapies aimed at host-pathogen interactions emerge as potential treatment modalities. Thus, understanding the mechanism of pathogen recognition by the host and immune evasion appear to be the key scientific issues. Until recently, lipopolysaccharide (LPS) was recognized as a major pathogen-associated molecular pattern (PAMP) of Gram-negative bacteria. However, recently, ADP-L-glycero-β-D-manno-heptose (ADP-heptose), an intermediate carbohydrate metabolite of the LPS biosynthesis pathway, was discovered to activate the hosts\' innate immunity. Therefore, ADP-heptose is regarded as a novel PAMP of Gram-negative bacteria that is recognized by the cytosolic alpha kinase-1 (ALPK1) protein. The conservative nature of this molecule makes it an intriguing player in host-pathogen interactions, especially in the context of changes in LPS structure or even in its loss by certain resistant pathogens. Here, we present the ADP-heptose metabolism, outline the mechanisms of its recognition and the activation of its immunity, and summarize the role of ADP-heptose in the pathogenesis of infection. Finally, we hypothesize about the routes of the entry of this sugar into cytosol and point to emerging questions that require further research.

Sepsis is the leading cause of acute kidney injury (AKI), and specific treatment options for septic AKI are very limited. Here, we used bulk RNA sequencing of a septic model of AKI to characterize the mRNA profile during AKI. The differentially expressed genes (DEGs) mainly participate in the inflammatory response and metabolic processes. Analysis of comprehensive mRNA-seq datasets revealed sepsis-induced AKI-specific cohorts of expressed genes, and six DEGs were tested in urine from septic patients with/without AKI. TRAF-interacting protein with forkhead-associated domain (TIFA) and fatty acid synthase (FASN) were differentially expressed in the urine from the sepsis-induced AKI group. Furthermore, we found that TIFA expression was significantly upregulated in mouse kidney tissue following cecal ligation and puncture (CLP). We sought to investigate its role in lipopolysaccharide (LPS) (TLR4 ligand)- and oligodeoxynucleotides (ODN) (TLR9 ligand)-treated human kidney cells and mouse. TIFA was located in Lotus tetragonolobus lectin (LTL) positive renal cells in kidney tissue, which was stained by immunofluorescence. Exposure of HK-2 cells to LPS and ODN caused disruption of the mitochondrial transmembrane potential. The results of transmission electron microscope (TEM) showed that mitochondrial damages were improved in TIFA-knockdown group. Moreover, knockdown of TIFA resulted in a decrease in the percentage of annexin V-positive and PI-negative cells after ODN treatment. The protein of NLRP3, Caspase-1 and GSDMD were also decreased when si-TIFA was transferred into HK-2 cells following LPS and ODN treatment. Activation of TIFA enhanced the expression of IL-1β and IL18. These results indicated that TIFA induced pyroptosis by activating the mitochondrial damage. Our study provides a detailed transcriptomic description of the renal cellular responses after sepsis. Our study suggest that TIFA is involved in pyroptosis by activating the mitochondrial damage and may be a therapeutic target to treat sepsis-induced kidney injury.

ADP-heptose activates the protein kinase ALPK1 triggering TIFA phosphorylation at Thr9, the recruitment of TRAF6 and the subsequent production of inflammatory mediators. Here, we demonstrate that ADP-heptose also stimulates the formation of Lys63- and Met1-linked ubiquitin chains to activate the TAK1 and canonical IKK complexes, respectively. We further show that the E3 ligases TRAF6 and c-IAP1 operate redundantly to generate the Lys63-linked ubiquitin chains required for pathway activation, which we demonstrate are attached to TRAF6, TRAF2 and c-IAP1, and that c-IAP1 is recruited to TIFA by TRAF2. ADP-heptose also induces activation of the kinase TBK1 by a TAK1-independent mechanism, which require TRAF2 and TRAF6. We establish that ALPK1 phosphorylates TIFA directly at Thr177 as well as Thr9 in vitro. Thr177 is located within the TRAF6-binding motif and its mutation to Asp prevents TRAF6 but not TRAF2 binding, indicating a role in restricting ADP-heptose signalling. We conclude that ADP-heptose signalling is controlled by the combined actions of TRAF2/c-IAP1 and TRAF6.

Previous studies have reported that TIFA plays different roles in various tumor types. However, the function of TIFA in colorectal cancer (CRC) remains unclear. Here, we showed that the expression of TIFA was markedly increased in CRC versus normal tissue, and positively correlated with CRC TNM stages. In agreement, we found that the CRC cell lines show increased TIFA expression levels versus normal control. The knockdown of TIFA inhibited cell proliferation but had no effect on cell apoptosis in vitro or in vivo. Moreover, the ectopic expression of TIFA enhanced cell proliferation ability in vitro and in vivo. In contrast, the expression of mutant TIFA (T9A, oligomerization site mutation; D6, TRAF6 binding site deletion) abolished TIFA-mediated cell proliferation enhancement. Exploration of the underlying mechanism revealed that the protein synthesis-associated kinase RSK and PRAS40 activation were responsible for TIFA-mediated CRC progression. In summary, these findings suggest that TIFA plays a role in mediating CRC progression. This could provide a promising target for CRC therapy.

A hallmark of infection by the pathogen Helicobacter pylori, which colonizes the human gastric epithelium, is the simultaneous activation of the classical and alternative nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathways, underlying inflammation and cell survival. Here, we report that the classical NF-κB target gene product A20 contributes to the negative regulation of alternative NF-κB signaling in gastric epithelial cells infected by H. pylori. Mechanistically, the de novo synthesized A20 protein interacts with tumor necrosis factor receptor-associated factor-interacting protein with forkhead-associated domain (TIFA) and thereby interferes with the association of TIFA with the NIK regulatory complex. We also show that alternative NF-κB activity contributes to the up-regulation of anti-apoptotic genes, such as baculoviral IAP repeat containing 2 (BIRC2), BIRC3 and B-cell lymphoma 2-related protein A1 (BCL2A1) in gastric epithelial cells. Furthermore, the observed over-expression of RelB in human gastric biopsies with type B gastritis and RelB-dependent suppression of apoptotic cell death emphasize an important role of the alternative NF-κB pathway in H. pylori infection.

[Figure: see text].

Microbial pathogens have evolved complex mechanisms to interface with host cells in order to evade host defenses and replicate. However, mammalian innate immune receptors detect the presence of molecules unique to the microbial world or sense the activity of virulence factors, activating antimicrobial and inflammatory pathways. We focus on how studies of the major virulence factor of one group of microbial pathogens, the type III secretion system (T3SS) of human pathogenic Yersinia, have shed light on these important innate immune responses. Yersinia are largely extracellular pathogens, yet they insert T3SS cargo into target host cells that modulate the activity of cytosolic innate immune receptors. This review covers both the host pathways that detect the Yersinia T3SS and the effector proteins used by Yersinia to manipulate innate immune signaling.