Semen Strychni Pulveratum and vomicine alleviate neuroinflammation in amyotrophic lateral sclerosis through cGAS-STING-TBK1 pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Amyotrophic lateral sclerosis (ALS) is a fetal neuromuscular disorder characterized by the gradual deterioration of motor neurons. Semen Strychni pulveratum (SSP), a processed version of Semen Strychni (SS) powder, is widely used to treat ALS in China. Vomicine is one of the most primary components of SS. However, their pharmacological effects and mechanisms for ALS remain elusive. AIM OF THE STUDY: This study aimed to evaluate the neuroprotective and anti-neuroinflammatory effects of SSP and vomicine, as well as to explore their protective roles in ALS and the underlying mechanisms. MATERIALS AND METHODS: In vivo, 8-week-old hSOD1-WT mice and hSOD1-G93A mice were orally administered different concentrations of SSP (SSP-L = 5.46 mg/ml, SSP-M = 10.92 mg/ml or SSP-H = 16.38 mg/ml) once every other day for 8 weeks. A series of experiments, including body weight measurement, footprint tests, Hematoxylin & Eosin staining, and Nissl staining, were performed to evaluate the preventive effect of SSP. Immunofluorescence staining, western blotting, and RT-qPCR were subsequently performed to evaluate activation of the cGAS-STING-TBK1 pathway in the spinal cord. In vitro, hSOD1G93A NSC-34 cells were treated with vomicine to further explore the pharmacological mechanism of vomicine in the treatment of ALS via the cGAS-STING-TBK1 pathway. RESULTS: SSP improved motor function, body weight loss, gastrocnemius muscle atrophy, and motor neuron loss in the spine and cortex of hSOD1-G93A mice. Furthermore, the cGAS-STING-TBK1 pathway was activated in the spinal cord of hSOD1-G93A mice, with activation predominantly observed in neurons and microglia. However, the levels of cGAS, STING, and pTBK1 proteins and cGAS, IRF3, IL-6, and IL-1ß mRNA were reversed following intervention with SSP. Vomicine not only downregulated the levels of cGAS, TBK1, IL-6 and IFN-ß mRNA, but also the levels of cGAS and STING protein in hSOD1G93A NSC-34 cells. CONCLUSION: This study demonstrated that SSP and vomicine exert neuroprotective and anti-neuroinflammatory effects in the treatment of ALS. SSP and vomicine may reduce neuroinflammation by regulating the cGAS-STING-TBK1 pathway, and could thereby play a role in ALS treatment.

A TBK1-independent primordial function of STING in lysosomal biogenesis.

Stimulator of interferon genes (STING) is activated in many pathophysiological conditions, leading to TBK1-dependent interferon production in higher organisms. However, primordial functions of STING independent of TBK1 are poorly understood. Here, through proteomics and bioinformatics approaches, we identify lysosomal biogenesis as an unexpected function of STING. Transcription factor EB (TFEB), an evolutionarily conserved regulator of lysosomal biogenesis and host defense, is activated by STING from multiple species, including humans, mice, and frogs. STING-mediated TFEB activation is independent of TBK1, but it requires STING trafficking and its conserved proton channel. GABARAP lipidation, stimulated by the channel of STING, is key for STING-dependent TFEB activation. STING stimulates global upregulation of TFEB-target genes, mediating lysosomal biogenesis and autophagy. TFEB supports cell survival during chronic sterile STING activation, a common condition in aging and age-related diseases. These results reveal a primordial function of STING in the biogenesis of lysosomes, essential organelles in immunity and cellular stress resistance.

TBK1 is involved in M-CSF-induced macrophage polarization through mediating the IRF5/IRF4 axis.

TANK binding kinase 1 (TBK1) is an important kinase that is involved in innate immunity and tumor development. Macrophage colony-stimulating factor (M-CSF) regulates the differentiation and function of macrophages towards the immunosuppressive M2 phenotype in the glioblastoma multiforme microenvironment. The role of TBK1 in macrophages, especially in regulating macrophage polarization in response to M-CSF stimulation, remains unclear. Here, we found high TBK1 expression in human glioma-infiltrating myeloid cells and that phosphorylated TBK1 was highly expressed in M-CSF-stimulated macrophages but not in granulocyte-macrophage CSF-induced macrophages (granulocyte-macrophage-CSF is involved in the polarization of M1 macrophages). Conditional deletion of TBK1 in myeloid cells induced M-CSF-stimulated bone marrow-derived macrophages to exhibit a proinflammatory M1-like phenotype with increased protein expression of CD86, interleukin-1ß and tumor necrosis factor-α, as well as decreased expression of arginase 1. Mechanistically, TBK1 deletion or inhibition by amlexanox or GSK8612 reduced the expression of the transcription factor interferon-regulatory factor (IRF)4 and increased the level of IRF5 activation in macrophages stimulated with M-CSF, leading to an M1-like profile with highly proinflammatory factors. IRF5 deletion reversed the effect of TBK1 inhibition on M-CSF-mediated macrophage polarization. Our findings suggest that TBK1 contributes to the regulation of macrophage polarization in response to M-CSF stimulation partly through the IRF5/IRF4 axis.

bcIRF5 activates bcTBK1 phosphorylation to enhance PANoptosis during GCRV infection.

TBK1 is an important IFN antiviral signalling factor, and in previous work black carp TBK1 (bcTBK1) and black carp IRF5 (bcIRF5) together promoted cell death in GCRV-infected cells. In this research, bcTBK1 and bcIRF5 were investigated both in vivo and in vitro to delineate their individual and combined functions. This study demonstrated that both bcTBK1 and bcIRF5 expressions were modulated in response to GCRV infection across the intestine, gill, kidney and spleen. In bcgill cells, overexpression of bcTBK1 and bcIRF5 initially suppressed the expression of cell death-related genes, including RIPK1, caspase1, caspase3 and bax, but this suppression was negated upon GCRV infection. In vivo, mRNA expression levels of RIPK1 and related genes varied by tissue following bcTBK1 or bcIRF5 overexpression and GCRV infection. Notably, intracellular co-overexpression of bcTBK1 and bcIRF5 led to significant upregulation of caspase3, caspase1, bax, and IL1ß, along with enhanced caspase3 activity post-GCRV infection. This co-expression correlated with higher survival rates in black carp during GCRV infection and increased caspase3 mRNA in the spleen and gills. Hematoxylin-eosin (HE) staining indicated disorganized spleen tissue and edematous, hyperplastic gill changes in co-transfected groups after infection. TUNEL staining of tissue sections showed that DNA breakage was significantly stronger in the co-transfected group than in the other groups during GCRV infection. Further phosphorylation experiments showed that bcIRF5 promoted phosphorylation modification of bcTBK1. Thus, these data suggest that bcIRF5 activates bcTBK1 by enhancing its phosphorylation and promotes PANoptosis in GCRV-infected cells.

Mycobacterial peptidyl prolyl isomerase A activates STING-TBK1-IRF3 signaling to promote IFNß release in macrophages.

Peptidyl prolyl isomerases (PPIases) are well-conserved protein-folding enzymes that moonlight as regulators of bacterial virulence. Peptidyl prolyl isomerase A, PPiA (Rv0009) is a secretory protein of Mycobacterium tuberculosis that possesses sequence and structural similarity to eukaryotic cyclophilins. In this study, we validated the interaction of PPiA with stimulator of interferon genes (STING) using both, Escherichia coli-based and mammalian in vitro expression systems. In vitro pull-down assays confirmed that the cytosolic domain of STING interacts with PPiA, and moreover, we found that PPiA could induce dimerization of STING in macrophages. In silico docking analyses suggested that the PXXP (PDP) motif of PPiA is crucial for interaction with STING, and concordantly, mutations in the PDP domain (PPiA MUT-II) abrogated this interaction, as well as the ability of PPiA to facilitate STING dimerization. In agreement with these observations, fluorescence microscopy demonstrated that STING and wild-type PPiA, but not PPiA MUT-II, could colocalize when expressed in HEK293 cells. Highlighting the importance of the PDP domain further, PPiA, but not PPiA MUT-II could activate Tank binding kinase 1 (TBK1)-interferon regulatory factor 3 (IRF3) signaling to promote the release of interferon-beta (IFNß). PPiA, but not PPiA MUT-II expressed in Mycobacterium smegmatis induced IFNß release and facilitated bacterial survival in macrophages in a STING-dependent manner. The PPiA-induced release of IFNß was c-GAS independent. We conclude that PPiA is a previously undescribed mycobacterial regulator of STING-dependent type I interferon production from macrophages.

Suppression of Interferon Response and Antiviral Strategies of Bunyaviruses.

The order Bunyavirales belongs to the class of Ellioviricetes and is classified into fourteen families. Some species of the order Bunyavirales pose potential threats to human health. The continuously increasing research reveals that various viruses within this order achieve immune evasion in the host through suppressing interferon (IFN) response. As the types and nodes of the interferon response pathway are continually updated or enriched, the IFN suppression mechanisms and target points of different virus species within this order are also constantly enriched and exhibit variations. For instance, Puumala virus (PUUV) and Tula virus (TULV) can inhibit IFN response through their functional NSs inhibiting downstream factor IRF3 activity. Nevertheless, the IFN suppression mechanisms of Dabie bandavirus (DBV) and Guertu virus (GTV) are mostly mediated by viral inclusion bodies (IBs) or filamentous structures (FSs). Currently, there are no effective drugs against several viruses belonging to this order that pose significant threats to society and human health. While the discovery, development, and application of antiviral drugs constitute a lengthy process, our focus on key targets in the IFN response suppression process of the virus leads to potential antiviral strategies, which provide references for both basic research and practical applications.

A novel role of ETV6 as a pro-viral factor in host response by inhibiting TBK1 phosphorylation.

E26-transforming specific (ETS) variant 6 (ETV6) is a transcription factor regulating the expression of interferon stimulating genes (ISGs) and involved in the embryonic development and hematopoietic regulation, but the role of ETV6 in host response to virus infection is not clear. In this study, we show that ETV6 was upregulated in DF-1 cells with poly(I:C) stimulation or IBDV, AIV and ARV infection via engagement of dsRNA by MDA5. Overexpression of ETV6 in DF-1 cells markedly inhibited IBDV-induced type I interferon (IFN-I) and ISGs expressions. In contrast, knockdown, or knockout of ETV6 remarkably inhibited IBDV replication via promoting IFN-I response. Furthermore, our data show that ETV6 negatively regulated host antiviral response to IBDV infection by interaction with TANK binding kinase 1 (TBK1) and subsequently inhibited its phosphorylation. These results uncovered a novel role of ETV6 as a pro-viral factor in host response by inhibiting TBK1 phosphorylation, furthering our understandings of RNA virus immunosuppression and providing a valuable clue to the development of antiviral reagents for the control of avian RNA virus infection.

Amlexanox targeted inhibition of TBK1 regulates immune cell function to exacerbate DSS-induced inflammatory bowel disease.

Amlexanox (ALX) is a small molecule drug for the treatment of inflammatory, autoimmune, metabolic and tumor diseases. At present, there are no studies on whether ALX has a therapeutic effect on inflammatory bowel disease (IBD). In this study, we used a mouse model of dextran sulfate sodium (DSS)-induced colitis to investigate the effect of ALX targeted inhibition of TBK1 on colitis. We found that the severity of colitis in mice was correlated with TBK1 expression. Notably, although ALX inhibited the activation of the TBK1-NF-κB/TBK1-IRF3 pro-inflammatory signaling pathway, it exacerbated colitis and reduced survival in mice. The results of drug safety experiments ruled out a relationship between this exacerbating effect and drug toxicity. In addition, ELISA results showed that ALX promoted the secretion of IL-1ß and IFN-α, and inhibited the production of cytokines IL-6, TNF-α, IL-10, TGF-ß and secretory IgA. Flow cytometry results further showed that ALX promoted T cell proliferation, activation and differentiation, and thus played a pro-inflammatory role; Also, ALX inhibited the generation of dendritic cells and the polarization of macrophages to M1 type, thus exerting anti-inflammatory effect. These data suggest that the regulation of ALX on the function of different immune cells is different, so the effect on the inflammatory response is bidirectional. In conclusion, our study demonstrates that simply inhibiting TBK1 in all immune cells is not effective for the treatment of colitis. Further investigation the anti-inflammatory mechanism of ALX on dendritic cells and macrophages may provide a new strategy for the treatment of IBD.

Azamollugin, a mollugin derivative, has inhibitory activity on MyD88- and TRIF-dependent pathways.

Previously, we reported that azamollugin, an aza-derivative of mollugin, exhibited potent inhibitory activity on NO production in LPS-stimulated RAW 264.7 cells. Further investigations in this study revealed that azamollugin not only suppressed iNOS gene expression regulated by NF-κB, but also inhibited LPS-induced IFN-ß expression, which is known to be regulated by IRF3. Azamollugin exhibited an inhibitory activity on LPS-induced IRAK1 activation, suggesting inhibitory effect on the MyD88-dependent pathway. Furthermore, azamollugin inhibited LPS-induced phosphorylation of IRF3 and its upstream factor, TBK1/IKKε, suggesting an inhibitory effect on the TRIF-dependent pathway via TLR4. In addition, azamollugin also suppressed poly(I:C)-induced phosphorylation of TBK1 and IRF3, suggesting an inhibitory effect on the TRIF-dependent pathway via TLR3. These results suggest that azamollugin has inhibitory activity against both the MyD88-dependent and TRIF-dependent pathways, respectively.

TBK1-Zyxin signaling controls tumor-associated macrophage recruitment to mitigate antitumor immunity.

Mechanical control is fundamental for cellular localization within a tissue, including for tumor-associated macrophages (TAMs). While the innate immune sensing pathways cGAS-STING and RLR-MAVS impact the pathogenesis and therapeutics of malignant diseases, their effects on cell residency and motility remain incompletely understood. Here, we uncovered that TBK1 kinase, activated by cGAS-STING or RLR-MAVS signaling in macrophages, directly phosphorylates and mobilizes Zyxin, a key regulator of actin dynamics. Under pathological conditions and in STING or MAVS signalosomes, TBK1-mediated Zyxin phosphorylation at S143 facilitates rapid recruitment of phospho-Zyxin to focal adhesions, leading to subsequent F-actin reorganization and reduced macrophage migration. Intratumoral STING-TBK1-Zyxin signaling was evident in TAMs and critical in antitumor immunity. Furthermore, myeloid-specific or global disruption of this signaling decreased the population of CD11b+ F4/80+ TAMs and promoted PD-1-mediated antitumor immunotherapy. Thus, our findings identify a new biological function of innate immune sensing pathways by regulating macrophage tissue localization, thus providing insights into context-dependent mitigation of antitumor immunity.

IFIT3 mediates TBK1 phosphorylation to promote activation of pDCs and exacerbate systemic sclerosis in mice.

OBJECTIVE: To assess the impact of the IFIT3/TBK1 signalling pathway in activating plasmacytoid dendritic cells (pDCs) and its role in the development of SSc. METHODS: Utilized single-cell RNA sequencing (scRNA-seq) and high-throughput transcriptome RNA sequencing to reveal the differential abundance of pDCs and the role of the key gene IFIT3 in SSc. Conducted in vitro cell experiments to evaluate the effect of IFIT3/TBK1 signalling pathway intervention on pDC activation cytokine release and fibroblast function. Constructed an IFIT3-/- mouse model using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) gene editing to assess the potential benefits of intervening in the IFIT3/TBK1 signalling pathway on skin and lung fibrosis in the SSc mouse model. RESULTS: The IFIT3/TBK1 signalling pathway plays a crucial role in activating pDCs, with IFIT3 acting as an upstream regulator of TBK1. Intervention in the IFIT3/TBK1 signalling pathway can inhibit pDC activation cytokine release and impact fibroblast function. The IFIT3-/- mouse model shows potential benefits of targeting the IFIT3/TBK1 signalling pathway in reducing skin and lung fibrosis in the SSc mouse model. CONCLUSION: This study provides new insights into potential therapeutic targets for SSc, highlighting the critical role of the IFIT3/TBK1 signalling pathway in SSc development. HIGHLIGHTS: This study elucidates the pivotal role of plasmacytoid dendritic cells (pDCs) in systemic sclerosis (SSc). This study identified the key regulatory gene involved in systemic sclerosis (SSc) as IFIT3. This study has found that IFIT3 functions as an upstream regulatory factor, activating TBK1. This study provides Evidence of the regulatory effects of the IFIT3/TBK1 pathway on plasmacytoid dendritic cells (pDCs). This study validated the therapeutic potential using the IFIT3-/- mouse model.

Mechanism of TBK1 activation in cancer cells.

TANK-binding kinase 1 (TBK1) is a serine/threonine kinase with well-established roles as a central player in innate immune signaling. Dysregulation of TBK1 activity has been implicated in a variety of pathophysiologic conditions, including cancer. Generally, TBK1 acts as an oncogene and increased TBK1 activity, indicated by increased phosphorylation at the Ser172 residue, can be observed in multiple human cancers. TBK1 can be activated either by autophosphorylation of Ser172 or transphosphorylation at this site by upstream kinases. Serving as a hub for integrating numerous extracellular and intracellular signals, TBK1 can be activated through multiple signaling pathways. However, the direct upstream kinase responsible for TBK1 activation remains elusive, which limits our comprehensive understanding of its activation mechanism and potential therapeutic application targeting TBK1-related signaling especially in cancer. In this review, we summarize the findings on mechanisms of TBK1 activation in cancer cells and recent discoveries that shed light on the direct upstream kinases promoting TBK1 activation.

Selective autophagy receptor p62/SQSTM1 inhibits TBK1-IRF7 innate immune pathway in triploid hybrid fish.

The production of type I interferon is tightly regulated to prevent excessive immune activation. However, the role of selective autophagy receptor SQSTM1 in this regulation in teleost remains unknown. In this study, we cloned the triploid fish SQSTM1 (3nSQSTM1), which comprises 1371 nucleotides, encoding 457 amino acids. qRT-PCR data revealed that the transcript levels of SQSTM1 in triploid fish were increased both in vivo and in vitro following spring viraemia of carp virus (SVCV) infection. Immunofluorescence analysis confirmed that 3nSQSTM1 was mainly distributed in the cytoplasm. Luciferase reporter assay results showed that 3nSQSTM1 significantly blocked the activation of interferon promoters induced by 3nMDA5, 3nMAVS, 3nTBK1, and 3nIRF7. Co-immunoprecipitation assays further confirmed that 3nSQSTM1 could interact with both 3nTBK1 and 3nIRF7. Moreover, upon co-transfection, 3nSQSTM1 significantly inhibited the antiviral activity mediated by TBK1 and IRF7. Mechanistically, 3nSQSTM1 decreased the TBK1 phosphorylation and its interaction with 3nIRF7, thereby suppressing the subsequent antiviral response. Notably, we discovered that 3nSQSTM1 also interacted with SVCV N and P proteins, and these viral proteins may exploit 3nSQSTM1 to further limit the host's antiviral innate immune responses. In conclusion, our study demonstrates that 3nSQSTM1 plays a pivotal role in negatively regulating the interferon signaling pathway by targeting 3nTBK1 and 3nIRF7.

Control of innate immunity and lipid biosynthesis in neurodegeneration.

The cGAS-STING innate immunity pathway and the SREBP-activated cholesterol and fatty acid synthesis pathway are abnormally co-regulated in neurodegenerative disease. Activation of STING signaling occurs at the endoplasmic reticulum (ER) membrane with STING anchored by INSIG1 along with SREBP and the sterol-bound SREBP cleavage activating protein (SCAP) when sterols are in abundance. When sterols are low, the INSIG-dependent STING pathway is inactivated and the SREBP-SCAP complex is translocated to the Golgi where SREBP is cleaved and translocated to the nucleus to transactivate genes for cholesterol and fatty acid synthesis. Thus, there is inverse activation of STING vs. SREBP: when innate immunity is active, pathways for cholesterol and fatty acid synthesis are suppressed, and vice versa. The STING pathway is stimulated by foreign viral cytoplasmic nucleic acids interacting with the cyclic GMP-AMP synthase (cGAS) DNA sensor or RIG-I and MDA5 dsRNA sensors, but with neurodegeneration innate immunity is also activated by self-DNAs and double-stranded RNAs that accumulate with neuronal death. Downstream, activated STING recruits TBK1 and stimulates the transactivation of interferon stimulated genes and the autophagy pathway, which are both protective. However, chronic activation of innate immunity contributes to microglia activation, neuroinflammation and autophagy failure leading to neurodegeneration. STING is also a proton channel that when activated stimulates proton exit from STING vesicles leading to cell death. Here we review the salient features of the innate immunity and cholesterol and fatty acid synthesis pathways, observations of abnormal STING and SREBP signaling in neurodegenerative disease, and relevant therapeutic approaches.

TBK1 is paradoxical in tumor development: a focus on the pathway mediating IFN-I expression.

TANK-binding kinase 1 (TBK1) is a member of the IKK family and plays a crucial role in the activation of non-canonical NF-κB signaling and type I interferon responses. The aberrant activation of TBK1 contributes to the proliferation and survival of various types of tumor cells, particularly in specific mutational or tumorous contexts. Inhibitors targeting TBK1 are under development and application in both in vivo and in vitro settings, yet their clinical efficacy remains limited. Numerous literatures have shown that TBK1 can exhibit both tumor promoting and tumor inhibiting effects. TBK1 acts as a pivotal node within the innate immune pathway, mediating anti-tumor immunity through the activation of innate immune responses. Facilitating interferon-I (IFN-I) production represents a critical mechanism through which TBK1 bridges these processes. IFN has been shown to exert both beneficial and detrimental effects on tumor progression. Hence, the paradoxical role of TBK1 in tumor development may necessitate acknowledgment in light of its downstream IFN-I signaling cascade. In this paper, we review the signaling pathways mediated by TBK1 in various tumor contexts and summarize the dual roles of TBK1 and the TBK1-IFN pathways in both promoting and inhibiting tumor progression. Additionally, we highlight the significance of the TBK1-IFN pathway in clinical therapy, particularly in the context of immune response. We anticipate further advancements in the development of TBK1 inhibitors as part of novel cancer treatment strategies.

Lysosomal TBK1 responds to amino acid availability to relieve Rab7-dependent mTORC1 inhibition.

Lysosomes play a pivotal role in coordinating macromolecule degradation and regulating cell growth and metabolism. Despite substantial progress in identifying lysosomal signaling proteins, understanding the pathways that synchronize lysosome functions with changing cellular demands remains incomplete. This study uncovers a role for TANK-binding kinase 1 (TBK1), well known for its role in innate immunity and organelle quality control, in modulating lysosomal responsiveness to nutrients. Specifically, we identify a pool of TBK1 that is recruited to lysosomes in response to elevated amino acid levels. This lysosomal TBK1 phosphorylates Rab7 on serine 72. This is critical for alleviating Rab7-mediated inhibition of amino acid-dependent mTORC1 activation. Furthermore, a TBK1 mutant (E696K) associated with amyotrophic lateral sclerosis and frontotemporal dementia constitutively accumulates at lysosomes, resulting in elevated Rab7 phosphorylation and increased mTORC1 activation. This data establishes the lysosome as a site of amino acid regulated TBK1 signaling that is crucial for efficient mTORC1 activation. This lysosomal pool of TBK1 has broader implications for lysosome homeostasis, and its dysregulation could contribute to the pathogenesis of ALS-FTD.

Phosphorylation of the selective autophagy receptor TAX1BP1 by TBK1 and IKBKE/IKKi promotes ATG8-family protein-dependent clearance of MAVS aggregates.

TAX1BP1 is a selective macroautophagy/autophagy receptor that inhibits NFKB and RIGI-like receptor (RLR) signaling to prevent excessive inflammation and maintain homeostasis. Selective autophagy receptors such as SQSTM1/p62 and OPTN are phosphorylated by the kinase TBK1 to stimulate their selective autophagy function. However, it is unknown if TAX1BP1 is regulated by TBK1 or other kinases under basal conditions or during RNA virus infection. Here, we found that TBK1 and IKBKE/IKKi function redundantly to phosphorylate TAX1BP1 and regulate its autophagic turnover through canonical macroautophagy. TAX1BP1 phosphorylation promotes its localization to lysosomes, resulting in its degradation. Additionally, we found that during vesicular stomatitis virus infection, TAX1BP1 is targeted to lysosomes in an ATG8-family protein-independent manner. Furthermore, TAX1BP1 plays a critical role in the clearance of MAVS aggregates, and phosphorylation of TAX1BP1 controls its MAVS aggrephagy function. Together, our data support a model whereby TBK1 and IKBKE license TAX1BP1-selective autophagy function to inhibit MAVS and RLR signaling.Abbreviations: ATG: autophagy related; BafA1: bafilomycin A1; CALCOCO2: calcium binding and coiled-coil domain 2; GFP: green fluorescent protein; IFA: indirect immunofluorescence assay; IFN: interferon; IκB: inhibitor of nuclear factor kappa B; IKK: IκB kinase; IRF: interferon regulatory factor; KO: knockout; LAMP1: lysosomal associated membrane protein 1; LIR: LC3-interacting region; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAVS: mitochondrial antiviral signaling protein; MEF: mouse embryonic fibroblast; MOI: multiplicity of infection; IKBKG/NEMO: inhibitor of nuclear factor kappa B kinase regulatory subunit gamma; NFKB: nuclear factor kappa B; OPTN: optineurin; Poly(I:C): polyinosinic-polycytidylic acid; RB1CC1/FIP200: RB1 inducible coiled-coil 1; RIGI: RNA sensor RIG-I; RLR: RIGI-like receptor; SDD-AGE: semi-denaturing detergent-agarose gel electrophoresis; SeV: Sendai virus; SLR: SQSTM1-like receptor; SQSTM1: sequestosome 1; TAX1BP1: Tax1 binding protein 1; TBK1: TANK binding kinase 1; TNF: tumor necrosis factor; TRAF: TNF receptor associated factor; VSV: vesicular stomatitis virus; ZnF: zinc finger.

Post-Translational Variants of Major Proteins in Amyotrophic Lateral Sclerosis Provide New Insights into the Pathophysiology of the Disease.

Post-translational modifications (PTMs) affecting proteins during or after their synthesis play a crucial role in their localization and function. The modification of these PTMs under pathophysiological conditions, i.e., their appearance, disappearance, or variation in quantity caused by a pathological environment or a mutation, corresponds to post-translational variants (PTVs). These PTVs can be directly or indirectly involved in the pathophysiology of diseases. Here, we present the PTMs and PTVs of four major amyotrophic lateral sclerosis (ALS) proteins, SOD1, TDP-43, FUS, and TBK1. These modifications involve acetylation, phosphorylation, methylation, ubiquitination, SUMOylation, and enzymatic cleavage. We list the PTM positions known to be mutated in ALS patients and discuss the roles of PTVs in the pathophysiological processes of ALS. In-depth knowledge of the PTMs and PTVs of ALS proteins is needed to better understand their role in the disease. We believe it is also crucial for developing new therapies that may be more effective in ALS.

STRAP upregulates antiviral innate immunity against PRV by targeting TBK1.

Serine/threonine kinase receptor-associated protein (STRAP) serves as a scaffold protein and is engaged in a variety of cellular activities, although its importance in antiviral innate immunity is unknown. We discovered that STRAP works as an interferon (IFN)-inducible positive regulator, facilitating type I IFN signaling during pseudorabies virus infection. Mechanistically, STRAP interacts with TBK1 to activate type I IFN signaling. Both the CT and WD40 7 - 6 domains contribute to the function of STRAP. Furthermore, TBK1 competes with PRV-UL50 for binding to STRAP, and STRAP impedes the degradation of TBK1 mediated by PRV-UL50, thereby increasing the interaction between STRAP and TBK1. Overall, these findings reveal a previously unrecognized role for STRAP in innate antiviral immune responses during PRV infection. STRAP could be a potential therapeutic target for viral infectious diseases.

Crosstalk and Prospects of TBK1 in Inflammation.

BACKGROUND: TANK-binding kinase 1 (TBK1) is a pivotal mediator of innate immunity, activated by receptors such as mitochondrial antiviral signaling protein (MAVS), stimulator of interferon genes (STING), and TIR-domain-containing adaptor inducing interferon-ß (TRIF). It modulates immune responses by exerting influence on the type I interferons (IFN-Is) signaling and the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathways, Over the past few years, TBK1 multifaceted role in both immune and inflammatory responses is increasingly recognized. METHODS AND RESULTS: This review aims to scrutinize how TBK1 operates within the NF-κB pathway and the interferon regulatory transcription factor 3 (IRF3)-dependent IFN-I pathways, highlighting the kinases and other molecules involved in these processes. This analysis reveals the distinctive characteristics of TBK1's involvement in these pathways. Furthermore, it has been observed that the role of TBK1 in exerting anti-inflammatory or pro-inflammatory effects is contingent upon varying pathological conditions, indicating a multifaceted role in immune regulation. DISCUSSION: TBK1's evolving role in various diseases and the potential of TBK1 inhibitors as therapeutic agents are explored. Targeting TBK1 may provide new strategies for treating inflammatory disorders and autoimmune diseases associated with IFN-Is, warranting further investigation.