Dissertations / Theses on the topic 'Ash2L'

In eukaryotes, the SET1 family of methyltransferases carry out the methylation of Lysine 4 on Histone H3. Alone, these enzymes exhibit low enzymatic activity and require the presence of additional regulatory proteins, which include RbBP5, Ash2L, WDR5 and DPY-30, to stimulate their catalytic activity. While previous structural studies established the structural basis underlying the interaction between RbBP5, Ash2L and WDR5, the formation of the Ash2L/DPY-30 complex remains elusive. Here we report the crystal structure of the Ash2L/DPY-30 complex solved at 2.2Å. Our results show that a Cterminal amphipathic α-helix on Ash2L makes several hydrophobic interactions with the DPY-30 homodimer. Moreover, the structure reveals that a tryptophan residue on Ash2L, which directly precedes its C-terminal amphipathic α-helix, makes key interactions with one of DPY-30 α-helix. Finally, biochemical studies of Ash2L revealed a hitherto unknown ability of this protein to bind anionic lipids.

Background. Different histone post-translational modifications (PTMs) are crucial in the regulation of chromatin, including methylations of H3 at Lysine 4 by the MLL complex. A relevant issue is how this is causally correlated to the binding of specific transcription factors (TFs) in regulatory regions. NF-Y is a TF that regulates 30% of mammalian promoters containing the widespread CCAAT element. We and others established that the presence of H3K4me3 is dependent upon the binding of NF-Y. Here, we investigate the mechanisms of H3K4me3 deposition by NF-Y. Methods. We employed Chromatin Immunoprecipitation in cells in which Ash2L and NF-Y subunits were knocked down by RNAi, to monitor the presence of histones PTMs and components of the MLL complex. We performed gene expression profiling of Ash2L-knocked down cells and analyzed the regulated genes. We performed ChIPs in leukemic cells in which MLL1 is devoid of the methyltransferase domain and fused to the AF4 gene. Results. Knock down of the Ash2L subunit of MLL leads to a decrease in global H3K4me3 with a concomitant increase in H3K79me2. Knock down of NF-Y subunits prevents promoter association of Ash2L, but not MLL1, nor WDR5, and H3K4me3 drops dramatically. Endogenous NF-Y and Ash2L specifically interact in vivo. Analysis of the promoters of Ash2L regulated genes, identified by transcriptional profiling, suggests that a handful TF binding sites are moderately enriched, among which the CCAAT box. Finally, leukemic cells carrying the MLL-AF4 translocation show a decrease of H3K4me3, absence of Ash2L and increase in H3K79me2, while NF-Y binding was not significantly affected. Conclusions. Three types of conclusions are reached: (i) H3K4 methylation is not absolutely required for NF-Y promoter association. (ii) NF-Y acts upstream of H3K4me3 deposition by recruiting Ash2L. (iii) There is a general cross-talk between H3K4me3 and H3K79me2 which is independent from the presence of MLL oncogenic fusions.

The mixed lineage leukemia (MLL) proteins regulate an array of developmental and differentiation processes. Similar to other members of the SET1 family, association of MLL1-4 with Ash2L, RbBP5 and WDR5, collectively termed the MLL core complex, is required for MLL mediated histone H3 Lys-4 di/tri-methylation. Each member of the core complex has a unique role in modulating the activity of MLL1. WDR5 is key in nucleating the formation of the core complex by acting as a structural scaffold, whereas Ash2L and RbBP5 are responsible for stimulating MLL methyltransferase activity. Currently, the structural and biochemical mechanisms utilized by the core complex to regulate MLL1 activity are unknown. Through structural and biochemical dissection of the core complex we have assigned specific functions to core complex subunits and have identified the minimal structural requirements for methyltransferase activity. Furthermore, through structure based drug design, we have identified a peptidomimetic inhibitor of MLL1 methyltransferase activity.

A l'inici del desenvolupament, les cèl·lules d'un organisme comencen a diferenciar-se en resposta a estímuls intrínsecs i del seu entorn cel·lular. Aquests estímuls provoquen l'expressió de factors reguladors de la transcripció que estableixen uns patrons d'expressió gènica específics per a cada tipus cel·lular. Malgrat l'expressió d'aquests factors és transitòria, els patrons d'expressió gènica establerts inicialment es mantenen al llarg del desenvolupament, transmetent-se de generació en generació a les cèl·lules filles. El fet que una cèl·lula expressi uns gens i no uns altres durant molts cicles de divisió cel·lular, li confereix una identitat pròpia i fa sobreentendre que hi ha d'haver un mecanisme, basat en senyals epigenètics, que reconegui i preservi el patró de gens que s'ha d'activar o silenciar.
Els gens del grup Polycomb (PcG) i trithorax (trxG) codifiquen per proteïnes reguladores de la cromatina implicades en el manteniment de les identitats cel·lulars. La seva funció com a reguladors dels gens homeòtics ha estat molt ben documentada però es coneix molt poc sobre els efectes que poden tenir en altres gens diana o processos del desenvolupament. En aquesta Tesi s'utilitza el patró de venes i intervenes de l'ala com a model per entendre la funció del gen trxG absent, small or homeotic discs 2 (ash2). Es mostra que ash2 es requereix per mantenir l'activació dels gens promotors d'intervena blistered i net, i per reprimir l'expressió de rhomboid, un component de la via de DER, necessària per la formació de venes. Els nostres resultats també mostren que ash2 actua com a repressor del gen knirps (kni), organitzador de la vena longitudinal L2. La inhibició que ash2 exerceix sobre kni és independent de spalt-major i spalt-related, els quals s'ha postulat que són els reguladors de kni. Tots aquests experiments indiquen que ash2 és essencial per dos processos durant el desenvolupament de l'ala: (1) mantenir el destí cel·lular d'intervena, ja sigui activant els gens promotors d'intervena o inhibint els de vena; i (2) mantenir kni en un estat reprimit en teixits fora de la vena L2.
A més, donat que els gens del grup PcG i trxG codifiquen per proteïnes que formen part de complexes multiproteics reguladors de la cromatina, en aquesta Tesi també s'ha intentat fer una aproximació al complexe d' ASH2. Mitjançant estudis de co-immunoprecipitació i marcatges en cromosomes politènics es va demostrar que les proteïnes ASH2 i Sin3A interaccionen amb la proteïna Host Cell Factor (HCF), suggerint que a Drosophila podria existir un complexe conservat, semblant al purificat en humans, amb capacitat histona metiltransferasa.
Differential gene expression results in cell diversity, although how different cell identities are established early in development and maintained throughout life is still poorly understood. Most of the transcription factors required for early developmental decisions are expressed transiently, but the gene expression patterns they trigger are maintained during cell division and inherited by daughter cells. Actively dividing cells must preserve individual genes in an on or off expression state after an initial commitment is made, especially given that some regulators disassemble from promoters during DNA replication or mitosis. Thus, developmental decisions may be maintained by the ability to deposit epigenetic marks involving chromatin modifying complexes to control the cellular memory of gene activity states. Genes of the Polycomb (PcG) and trithorax group (trxG) encode proteins that are engaged in the regulation of cellular memory.
absent, small or homeotic discs 2 (ash2) is a trxG protein that belongs to a 0.5 MDa complex thought to be involved in chromatin remodelling. In this thesis, to gain more insight into the function of ash2, we have used the Drosophila wing as a model system to examine whether vein- and intervein-specific genes and vein positioning genes act as putative targets of ash2 function. We found that ash2 is involved in activating intervein-promoting genes and downregulating the Egfr pathway. Moreover, ash2 also acts as a kni repressor independently of sal-C. These results strongly support a role for ash2 in maintaining vein/intervein developmental decisions and vein patterning in the developing wing.
We have also tried to do an approximation to the Drosophila ASH2 complex. We have demonstrated that ASH2 and Sin3A were found to colocalize on polytene chromosomes and coimmunoprecipitate with the Host Cell Factor (HCF). Together with the localization of ASH2 at sites of H3K4 trimethylation we propose that Drosophila might contain an ASH2 complex with histone methyltransferase activity similar to the one found in humans.

El gen absent, small, or homeotic discs 2 (ash2) és un membre del grup dels trithorax (trxG), reguladors positius de la transcripció dels gens homeòtics. Els al·lels mutants per ash2 són letals larvals o pupals i presenten anomalies en els discs imaginals i en el cervell. Hem identificat un nou transcrit, més petit que el descrit fins al moment, pel gen ash2. L'al·lel ash2I1 no presenta el transcrit més gran i és un mutant nul per la funció trxG d'ASH2. Per tal d'identificar gens diana d'ASH2, hem estudiat el patró d'expressió de larves mutants d'ash2 mitjançant microarrays de cADN. Entre els gens que varien els seus nivells d'expressió se n'hi troben d'involucrats en múltiples processos, com la proliferació, l'adhesió i el cicle cel·lulars. Alguns d'aquests gens han estat validats mitjançant tècniques d'expressió alternatives o estudis funcionals. D'altra banda, el gen ash2 ha estat relacionat funcionalment amb els gens absent, small or homeotic discs 1 (ash1) i trithorax (trx) mitjançant interaccions genètiques. ASH2 i ASH1 pertanyen a complexes multimèrics diferents de composició desconeguda i no es coneix amb exactitud de quina manera regulen la transcripció. L'examen minuciós dels patrons d'expressió de discs imaginals d'ala de mutants d'ash2 i d'ash1, mostra que hi ha un alt grau de solapament (overlap) entre els dos, tant en les comparacions gen a gen com en els processos que es troben alterats. Sorprenentment, la comparació dels transcriptomes de larva de mutants d'ash2 i de trx no mostren cap tipus de correlació. A més, ensenyem que la sub-expressió d'alguns d'aquests gens està directament correlacionada amb alguns dels fenotips observats en els mutants d'ash2. Per tal d'entendre millor el gen ash2, hem comparat els gens des-regulats en els seus mutants amb dades genòmiques provinents d'altres anàlisis in vivo i in silico. Una d'aquestes comparacions apunta cap a una putativa relació entre ASH2 i Sin3A, una proteïna associada a complexes desacetiladors d'histones. L'anàlisi en profunditat d'aquesta possibilitat ens ha portat a demostrar que tant ASH2 com Sin3A co-immunoprecipiten amb la proteïna Host Cell Factor (HCF), i que les tres co-localitzen en molts loci de cromosomes politènics. Aquestes evidències, juntament amb l'observació de que ASH2 co-localitza amb la tri-metilació de la H3K4, donen suport a un model en el qual ASH1 i ASH2 actuarien de forma seqüencial en la modificació de les histones per tal de mantenir els estats activats de transcripció.
The transcription factor absent, small, or homeotic discs 2 (ash2) gene is a member of the trithorax group of positive regulators of homeotic genes. Mutant alleles for ash2 are larval_pupal lethals and display imaginal disc and brain abnormalities. The allele used in this study is a true mutant for the trithorax function and lacks the longest transcript present in wild-type flies. In an attempt to identify gene targets of ash2, we have performed an expression analysis by using cDNA microarrays. Genes involved in cell cycle, cell proliferation, and cell adhesion are among these targets, and some of them are validated by functional and expression studies. Even though trithorax proteins act by modulating chromatin structure at particular chromosomal locations, evidence of physical aggregation of ash2-regulated genes has not been found.
Although ASH2 and ASH1 belong to distinct multimeric complexes and it is unclear how they act to regulate transcription, they are functionally related. In this study, examination of gene expression profiles in wing imaginal discs from ash2 and ash1 mutants revealed their transcriptomes are very similar and correlate with wing phenotypes. Comparison of the differentially expressed genes with results from other in vivo and in silico genome-wide analyses indicated, among others, a putative relationship between ASH2 and the histone deacetylase-associated protein Sin3A. ASH2 and Sin3A were found to colocalize on polytene chromosomes and coimmunoprecipitate with the Host Cell Factor. Together with the localization of ASH2 at sites of H3K4 trimethylation, our results support a model in which ASH2 and ASH1 are sequentially involved in histone modifications.

碩士
國立陽明大學
藥理學研究所
101
Induced pluripotent stem cells (iPSCs) were reprogrammed by somatic cells back to an embryonic stem cell (ESC) -like state by transfecting four factors Oct4, Nanog, Klf4 and c-myc. They shared many characteristics with embryonic stem cells. The structures of iPSCs were similar; they also expressed the same markers and genes. Importantly, iPSCs have the ability to grow indefinitely while maintaining pluripotency and the ability to differentiate into cells of all three germ layers. iPSCs have a general characteristic of open chromatin, a state that may be necessary for maintaining pluripotency. Recent studies established the importance of open chromatin, characterized by a predominance of euchromatin over heterochromatin, in maintenance of stem cell pluripotency. Ash2l is a key regulator of open chromatin in ES cells. In addition knockdown Ash2l would induce differentiation. Together all of above, Ash2l may play a major role in pluripotency maintenance. Ash2l has two splicing form Ash2l-a (long form) and Ash2l-b (short form) reported by previous studies. However the function of two isoform was still unclear. In this study compared with somatic cells, we observed the increased expression of ash2l-b in stem cells with western blotting. We also demonstrated that the expression of Ash2l associated with the expression of stemness genes including Oct4, Nanog and Nr5a2. In addition, we investigated whether the different isoform of Ash2l could contribute to regulate the pluripotency maintenance in iPSCs. We constructed the short hairpin RNA specific to knockdown Ash2l-a and Ash2l-b in induced pluripotent stem cells respectively. Our data showed that the suppression of the Ash2l-a not only diminished the expression of stemness genes such as Oct4, Nanog and Sox2 but also enhanced the expression of the ectodermal, mesodermal and endodermal-lineage markers. Therefore, these results indicate the isoforms of Ash2l may participate the important role involved in the maintenance of pluripotent status.

碩士
國立陽明大學
藥理學研究所
101
Pluripotent embryonic stem cells (ESCs) maintain self-renewal and the potential for rapid response to differentiation cues. Pluripotent stem cells (PSCs) have unique transcriptional regulatory networks and epigenetic states that are involved in maintaining pluripotency. There is well known about the establishment and maintenance of ES cell-specific transcription were regulated by a variety of specific transcriptional activators such as Oct4, Sox2 and Nanog. Despite the well-documented functions of these core activators, very little is known about an ES cell specific co-activator(s) involved in ES cell transcriptional core circuits. Ash2l, a core subunit of the histone 3 lysine 4 (H3K4) methyltransferase (HMT) complex, positively correlates with the undifferentiated state and is a regulator of ES cell self-renewal. However the function of Ash2l may be as a transcriptional regulator. The HMT complex components Wdr5 and Dpy30 had been find out contributed ESCs features. Therefore we sought to test the role of Ash2l during stem cell and cellular reprogramming by siRNA knockdown. We demonstrated that Ash2l, an ‘‘effector’’ of H3K4 methylation, interacts with the pluripotency transcription factor Oct4 by co-immunoprecipitation. The POU homeodomain transcription factor Oct4 (Pou5f1) is an essential mediator of the embryonic stem cell state. Next we confirmed the interaction between Ash2l and Oct4 enhances Oct4 bind its’ promoter by luciferase assay. In further to our data we found that Ash2l depletion induced the collapse of the extended ES cell transcriptional network by gene expression of stemness markers. Notably, we observed depletion of Ash2l led to drastic reduction of iPS cell formation. Overall we established that Ash2l is required for the initial reconfiguration phase of somatic cell reprogramming. We propose that the Ash2l-Oct4 partnership enhances Oct4 expression through transcriptional activation of its promoter and activated downstream genes expression. Summarized our finding, Ash2l is essential for maintenance for of ES cell pluripotency as well as somatic cell reprogramming by acting as a stem cell cofactor.

碩士
國立陽明大學
藥理學研究所
104
Abstract Recent study showed that Ash2l is necessary for ES cell pluripotency to open chromatin and carry out its function through multiple target proteins. However, a few transcription factors have shown importance in stem cells by interacting with master transcription factors, such as Oct4, Sox2 and Nanog. Our previous data suggested that Ash2l is essential for efficient somatic cell reprogramming. However, we also found that Ash2l directly interacts with Oct4, one of master transcription factor. According to studies and data, we would like to investigate the gene regulation between Ash2l and stemness genes in maintaing pluripotency and self-renewal in iPSCs. Our results showed that Ash2l may recruits master transcription factor Oct4 to facilitate stemness gene expression. We observed that Ash2l and Oct4 occupy at loci of several stemness-associated genes and transcription-regulated genes. In order to dissect the effect of Ash2l in gene regulation, we selected some genes from ChIP-seq database, which have significant Oct4, Ash2l, H3K27ac binding. Therefore, we believe that Ash2l may be an enhancer binding protein and promote gene expression. To prove our hypothesis, we chose Nanog, one of master transcription factors, as a model, and observed its regulation by Ash2l. We found Ash2l is required for Oct4 binding to enhancer regions and it positively regulates Nanog enhancer activity through recruitment of co-factor p300 and RNA polymerase II transcription subunit 1 protein (Med1). Therefore, it is able to promote initiation of transcription, and activate pluripotency genes expression.

碩士
國立陽明大學
解剖學及細胞生物學研究所
102
Abstract Histone H3K4 methylation is a crucial histone mark in regulating the stemness and developmental genes in embryonic stem cells. Ash2l belongs to the Trithorax complexes that add methyl groups to histone H3K4 and contribute to H3K4me1, me2, and me3 marks across the genome. Human ASH2L is downregulated rapidly and dramatically in K562, Hel and Dami cells differentiation with megakaryocytic features, suggesting that Ash2l play a role in hematopoiesis. Human ASH2L and its alternative splicing isoform ASH2L1 and ASH2L2 are homologous to mouse Ash2l isoforms were called Ash2l-long form and Ash2l-short from in this study. However, their functional role of in stem cell maintenance and iPSCs reprogramming are not investigated. In our study, we significantly identify two Ash2l isoform in mESC, iPSC and PSA1. In the comparison, the expression of Ash2l-short form was significantly high in the stem cell, whereas, Ash2-long form was remaining a similar level compared to the somatic cells. By using isoform-specific siRNA to investigate the functional role in iPSCs, we found that both isoforms maintain stemness in iPSCs. In the other hand, by using Retrovirus mediated overexpression of isoforms, we concluded that Ash2l-short form could enhance the reprogramming efficiency, such as alkaline phosphatase activity, colony formation and the expression of stemness genes. Meanwhile, Ash2l-long form provides a physical interaction with Oct4. In LC-MS proteomic analysis, we identify the Ash2l associated partner in the iPSCs, such as Parp1, Akt, HMGB2, Brd2, Smarcc1 and Wdr5. Those partners are known protein that contributes to the establishment of stemness. My thesis revealed the importance of isoforms in stem cell maintenance and reprogramming, and provides insight into how the reprogramming efficiency is enhanced by Ash2l-short in iPSC.

The mammalian immune system consists of the innate and adaptive arm/s that protect the host against pathogenic infections in a highly coordinated process involving multiple steps. Innate immune cells such as macrophages and dendritic cells (DCs) are responsible for determining the initiation of specific events and thus they tailor specific immune responses to eliminate the invading pathogen. Host innate immune responses are triggered by sensing of PAMPs (pathogen associated molecular patterns) or DAMPs (damage associated molecular patterns) via pattern recognition receptors (PRRs) and these inturn facilitate adaptive immune responses. The four major families of PRRs including Toll- like receptors (TLRs), NOD-like receptors (NLRs), RIG-I like receptors (RLRs) and C-type lectin receptors (CLRs) recognize a wide range of PAMPs and DAMPs. Engagement of PRRs with these stimuli promotes the differential induction of PRRs-driven signaling cascades such as inflammation, apoptosis, and autophagy among others that result in organized actions of multiple immune cells to eradicate microbial infection. However, in spite of having such effective and efficient immune system, some of the pathogens are able to breach immune layers and establish a successful infection while escaping host key immune surveillance mechanisms. One such pathogen of rising concern, Mycobacterium tuberculosis (Mtb) causing tuberculosis (TB), has evolved with mankind and causes an alarming 1.4 million deaths annually, in accordance with recent WHO reports. Mtb is an intracellular pathogen, whose primary target cells are macrophages. These are crucial effector immune cells that provide defense against a vast array of pathogens through the presentation of abundant cell surface receptors including TLRs that sensitize the host and execute the tailoring of immune responses during mycobacterial infection. In particular, TLR2 has been shown to elicit inflammatory responses including the increased expression of effector molecules such as tumor necrosis factor (TNF) α, interferon gamma (IFN-γ), xiinterleukins (ILs), chemokines and inflammatory cytokines in this process. Accumulating evidences indicate the role of phagosomal nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX)2 in producing oxidative stress in order to clear pathogen. Recent reports have suggested essential role of NOX2 in efficient killing of mycobacteria. For example, p47-phox (NOX2 subunit) null mouse showed increased susceptibility to mycobacteria infection. Interestingly, another recent study suggested the crucial role of host NAD(P)H quinone oxidoreductase 1 (NQO1) in promoting mycobacterial survival. Yet the contribution of NQO1 in regulating immune responses during pathogenic mycobacteria infection needs further investigation. Additionally, few reports indicate the role for hostderived reductases in regulating apoptosis. More importantly, several studies propose that mycobacteria can inhibit apoptosis and promote its survival. Therefore, dissecting the molecular mechanism in regulating host-derived reductases during mycobacterial infection will provide a further insight into comprehending the success of Mtb infection which depends on its ability to evade host immune responses by modulating host signaling and the related expression of immuno-regulatory molecules. In line with the above observations, defined signaling downstream of TLRs plays a central role in generating effective immune responses at the site of infection. Several immunological modules act as important accessory events to TLR-triggered signaling. Among the many signaling pathways, the canonical WNT-β-catenin pathway has been recently shown to play a crucial role in controlling the expression of inflammatory molecules during infection. However, information on the roles of WNT signaling in mediating inflammatory responses remains scanty. In particular, a growing number of studies have indicated the potential involvement of epigenetic factors to determine the host cell inflammatory responses. Modifications by such factors including DNA methylation, histone modification and noncoding RNAs are shown to be potential regulators of TLR mediated inflammatory responses. Histone methylation plays a crucial role in mediating TLR-triggered immune responses. For example, H3K27 demethylase Jumonji domain-containing 3 (JMJD3) was shown to determine the M2 macrophage development upon TLR stimulation during helminth infection. Furthermore, methyltransferases including SETDB2 and ASH1L, repress the expression of CXC-chemokine ligand 1 (CXCL1) and LPS-induced production of inflammatory cytokines respectively. However, it has not yet been determined, how pathogen modulates the recruitment of these epigenetic modifiers at specific promoters, thus orchestrating changes in complex phenomena such as inflammatory responses and apoptosis. Based on these evidences, the former part of the present study is focused on the expression of WNT-responsive epigenetic effectors which mediate the inflammatory responses and apoptosis during mycobacterial infection in mice. The investigation demonstrates that Mtb H37Rv triggers a robust activation of WNT-β-catenin signaling via its adaptor molecule protein kinase C (PKC) ζ. Indeed, emerging studies suggest a potential contribution of miRNAs in modulating immune responses during pathogenic infection. In line with these observations, we show that the WNT-responsive Mir-30e-3p stabilizes H4K20me1 methyltransferase SET8 by modulating the expression of its negative regulator, CDT2 (E3 ubiquitin ligase) during mycobacterial infection. Notably, stabilised SET8 leads to increased monomethylation of H4K20 on the promotors of Nqo1 and Trxrn1, which reflects in their significantly high RNA and protein levels. Despite being bonafide antioxidants, NQO1 is known to play a crucial role in regulating inflammation and TRXR1 is considered as an apoptosis controller via the regulation of ASK1 activity. Thus the depletion of NQO1 and TRXR1 by using specific siRNA reveals the important roles for NQO1 and TRXR1 in regulating inflammatory responses and apoptosis respectively during mycobacterial infection. Furthermore, experiments based on the use of dicoumarol (NQO1 inhibitor) and auranofin (TRXR1 inhibitor) in macrophages followed by pathogenic mycobacterial infection suggested that inflammatory responses and apoptosis are dependent on reductase activity of NQO1 and TRXR1. Most importantly, these functions of TRXR1 and NQO1 were explored in an experimental mouse TB model. Interestingly, mice administered with dicoumarol and auranofin showed a significantly reduced lung and spleen Mtb CFU. This was also corroborated by lung histopathology which alluded the reduced severity of TB through the analysis of granuloma, underscoring the importance of NQO1 and TRXR1 in TB pathogenesis. Collectively, we suggest the crucial role for WNT-responsive Mir-30e-3p - histone methyltransferase SET8 axis in regulating host-derived antioxidants NQO1 and TRXR1 in suppressing host immune responses. Thus, epigenetic reprogramming of the host cell by SET8 promotes Mtb survival in macrophages by regulating inflammation and apoptosis. Additionally, another histone methyltransferase, ASH2L, has been established as an immune regulator via facilitating activatory H3K4me3 (trimethylation) on the promoters of target genes at CG-rich DNA motifs. The participation of H3K4me3 performed by ASH2L methyltransferase, is highlighted in regulation of immune-related genes. However contribution of AHS2L in regulating host immune responses during mycobacterial infection has been not addressed. Based on these evidences, we established that Mtb-induced ASH2L, coupled with β-catenin directly regulates the expression of 5-LO and 15-LO. As a functional consequence, WNT/β-catenin and its responsive 5-LO and 15-LO were found to regulate expression of Gpr18 transcript which in turn regulates host inflammatory responses. In addition to membrane bound TLRs, emerging studies suggest the effective role for cytosolic NLRs in sensing PAMPs or DAMPs with great efficiency. Among the NLRs, nucleotide-binding oligomerization domain-containing protein (NOD) 1 and NOD2 are well characterized cytosolic receptors in determining inflammatory responses. Although, inflammation is a highly regulated fundamental defensive host mechanism, its hyper and chronic activation in response to different stimuli is associated with serious inflammatory disorders. Importantly, recent reports implicated the close association of NOD2 and WNT signaling pathways during the development of Crohn’s disease. In continuation, evidence suggests that NOD2-driven inflammatory disorders are associated with impairment of inflammasome function. Activation of WNT signaling is also implicated in the development of arthritis. However, precise mechanism of NOD2-WNT-inflammasome crosstalk or the pivotal role of WNT signaling during inflammatory arthritis requires extensive investigation. In this context, the present investigation demonstrates that upon activation with muramyl dipeptide (MDP, NOD2 specific agonist), NOD2 interacts positive regulator of WNT signaling Ly6/PLAUR domain-containing protein 6 (LYPD6) to stimulate and mediate WNT signaling activation. Strikingly, canonical adaptor molecules of NOD2 signaling, RIP2 and TAK1 were proven to be dispensable in NOD2-triggered WNT signaling activation. Furthermore, we found that WNT-responsive X-linked inhibitor of apoptosis (XIAP) leads to activation of NOD like receptors family pyrin domain-containing 3 (NLRP3) inflammasome complex. Further, NOD2-stimulated formation of active caspase-1 and secretion of IL-1β were found to be dependent on WNT and its responsive XIAP. Consistent with this in vitro data, mice administered with WNT-signaling inhibitor, XIAP inhibitor or Caspase-1 inhibitor displayed compromised ability to develop MDP-triggered acute arthritis. Taken together, our study contributes new biological insights towards understanding of NOD2-associated inflammatory responses. Altogether, our findings lay the groundworks for comprehending conceptual framework in orchestrating TLR and NLR responses by WNT/β-catenin signaling. More importantly, our study pays a tribute to novel mechanistic and functional insights into Mtb pathogenesis and inflammatory diseases, which promises to provide important leads in diagnostic and therapeutic approaches for immune-associated disorder