Methylene Insertion into Nitrogen-Heteroatom Single Bonds of 1,2-Azoles via a Zinc Carbenoid: An Alternative Tool for Skeletal Editing.

The nitrogen-heteroatom single bonds of 1,2-azoles and isoxazolines underwent methylene insertion in the presence of CH2 I2 (6 equiv.) and diethylzinc (3 equiv.) to produce a wide variety of the ring-expanded six-membered heterocycles. Density functional theory calculations suggest that the methylene insertion proceeds via cleavage of nitrogen-heteroatom single bonds followed by ring closure.

Chemoselective α-Trifluoroacetylation of Amides Using Highly Electrophilic Trifluoroacetic Anhydrides and 2,4,6-Collidine.

Chemoselective α-acylation of tertiary amides proceeded with highly electrophilic acid anhydrides and weak bases under mild conditions. ß-Ketoamides containing trifluoroacetyl or perfluoroacyl groups were selectively obtained even in the presence of other functional groups such as ketone, ester, etc. Density functional theory calculations suggest that 1-acyloxyenamine is the key intermediate for the chemoselective α-acylation.

Gut microbiota-dependent adaptor molecule recruits DNA methyltransferase to the TLR4 gene in colonic epithelial cells to suppress inflammatory reactions.

The intestine is inhabited by a large number of commensal bacteria that are immunologically non-self, potentially causing inflammation. However, in a healthy intestine, inflammation is strictly controlled at low levels to maintain homeostasis. We previously reported that the gut microbiota induce DNA methylation of the gene encoding Toll-like receptor (TLR) 4, a pattern recognition receptor that recognizes lipopolysaccharides of gram-negative bacteria, in colonic epithelial cells, suggesting its role in controlling intestinal inflammation. However, there remains a question of how gut microbiota cause methylation of only specific genes including TLR4, despite the fact that DNA methyltransferase (DNMT) is common to all genes targeted for methylation. Here, we identified RBM14 as an adaptor molecule that recruits DNMT to the TLR4 gene. RBM14 was shown to bind DNMT3 and be expressed at significantly higher levels in an intestinal epithelial cell (IEC) line with hypermethylated TLR4 gene than in an IEC line with hypomethylated TLR4 gene. In addition, RBM14 interacted with DNA regions of the TLR4 gene, and knockdown of RBM14 suppressed DNA methylation of the TLR4 gene in IECs. Furthermore, RBM14 expression was higher in colonic epithelial cells of conventional mice than in those of germ-free mice. Collectively, these results indicate that the gut microbiota induce methylation of the TLR4 gene in colonic epithelial cells by upregulating RBM14, which can recruit DNMT3 to the gene. The regulation of adaptor molecules such as RBM14, which bind to specific target genes and recruit DNMT, can explain, at least in part, how gut microbiota contribute to the maintenance of intestinal homeostasis through epigenetic control of specific gene expression in IECs.

Cecal Patches Generate Abundant IgG2b-Bearing B Cells That Are Reactive to Commensal Microbiota.

Gut-associated lymphoid tissue (GALT), such as Peyer's patches (PPs), are key inductive sites that generate IgA+ B cells, mainly through germinal center (GC) responses. The generation of IgA+ B cells is promoted by the presence of gut microbiota and dietary antigens. However, the function of GALT in the large intestine, such as cecal patches (CePs) and colonic patches (CoPs), and their regulatory mechanisms remain largely unknown. In this study, we demonstrate that the CePs possess more IgG2b+ B cells and have fewer IgA+ B cells than those in PPs from BALB/c mice with normal gut microbiota. Gene expression analysis of postswitched transcripts supported the differential expression of dominant antibody isotypes in B cells in GALT. Germ-free (GF) mice showed diminished GC B cells and had few IgA+ or IgG2b+ switched B cells in both the small and large intestinal GALT. In contrast, myeloid differentiation factor 88- (MyD88-) deficient mice exhibited decreased GC B cells and presented with reduced numbers of IgG2b+ B cells in CePs but not in PPs. Using ex vivo cell culture, we showed that CePs have a greater capacity to produce total and microbiota-reactive IgG2b, in addition to microbiota-reactive IgA, than the PPs. In line with the frequency of GC B cells and IgG2b+ B cells in CePs, there was a decrease in the levels of microbiota-reactive IgG2b and IgA in the serum of GF and MyD88-deficient mice. These data suggest that CePs have a different antibody production profile compared to PPs. Furthermore, the innate immune signals derived from gut microbiota are crucial for generating the IgG2b antibodies in CePs.

Synthesis of isoxazoloazaborines via gold(I)-catalyzed propargyl aza-Claisen rearrangement/borylative cyclization cascade.

Isoxazoloazaborines 3 and 5 have been synthesized from 4-N-propargylaminoisoxazole 1via gold(I)-catalyzed propargyl aza-Claisen rearrangement followed by electrophilic borylative cyclization in 27-86% yields. In situ generation of isoxazole 2 having an amino group and allenyl functionality is essential to give highly substituted isoxazoloazaborines 3 and 5, although the conventional propargyl aza-Claisen rearrangement readily affords the corresponding nitrogen-containing heterocycles, such as pyridines and pyrroles. The resulting isoxazoloazaborine 5a underwent the N-O bond insertion of zinc carbenoid to give oxazine-fused azaborine 6 in 48% yield.

Repeated exposure to kairomone-containing coffee odor improves abnormal olfactory behaviors in heterozygous oxytocin receptor knock-in mice.

The oxytocin receptor (OXTR) knockout mouse is a model of autism spectrum disorder, characterized by abnormalities in social and olfactory behaviors and learning. Previously, we demonstrated that OXTR plays a crucial role in regulating aversive olfactory behavior to butyric acid odor. In this study, we attempted to determine whether coffee aroma affects the abnormal olfactory behavior of OXTR-Venus knock-in heterozygous mice [heterozygous OXTR (±) mice] using a set of behavioral and molecular experiments. Four-week repeated exposures of heterozygous OXTR (±) mice to coffee odor, containing three kairomone alkylpyrazines, rescued the abnormal olfactory behaviors compared with non-exposed wild-type or heterozygous OXTR (±) mice. Increased Oxtr mRNA expression in the olfactory bulb and amygdala coincided with the rescue of abnormal olfactory behaviors. In addition, despite containing the kairomone compounds, both the wild-type and heterozygous OXTR (±) mice exhibited a preference for the coffee odor and exhibited no stress-like increase in the corticotropin-releasing hormone, instead of a kairomone-associated avoidance response. The repeated exposures to the coffee odor did not change oxytocin and estrogen synthetase/receptors as a regulator of the gonadotropic hormone. These data suggest that the rescue of abnormal olfactory behaviors in heterozygous OXTR (±) mice is due to the coffee odor exposure-induced OXTR expression.

SARS-CoV-2 infection of intestinal epithelia cells sensed by RIG-I and DHX-15 evokes innate immune response and immune cross-talk.

SARS-CoV-2 causes a spectrum of clinical symptoms from respiratory damage to gastrointestinal disorders. Intestinal infection of SARS-CoV-2 triggers immune response. However, the cellular mechanism that how SARS-CoV-2 initiates and induces intestinal immunity is not understood. Here, we exploited SARS-CoV-2-GFP/ΔN trVLP pseudo-virus system and demonstrated that RIG-I and DHX15 are required for sensing SARS-CoV-2 and inducing cellular immune response through MAVS signaling in intestinal epithelial cells (IECs) upon SARS-CoV-2 infection. NLRP6 also engages in the regulation of SARS-CoV-2 immunity by producing IL-18. Furthermore, primary cellular immune response provoked by SARS-CoV-2 in IECs further cascades activation of MAIT cells and produces cytotoxic cytokines including IFN-γ, granzyme B via an IL-18 dependent mechanism. These findings taken together unveil molecular basis of immune recognition in IECs in response to SARS-CoV-2, and provide insights that intestinal immune cross-talk with other immune cells triggers amplified immunity and probably contributes to immunopathogenesis of COVID-19.

Synthesis of 4-amino-5-allenylisoxazoles via gold(I)-catalysed propargyl aza-Claisen rearrangement.

Propargyl aza-Claisen rearrangement of 4-propargylaminoisoxazoles 1 proceeded in the presence of cationic gold(i) catalysts to give 4-amino-5-allenylisoxazoles 2 in good to high yields. The silyl group at the terminal alkyne and a cationic gold(i) catalyst bearing a sterically bulky ligand are essential for the generation of isolable allene intermediates. The N-protection of the generated 4-amino-5-allenylisoxazoles 2 allowed the isolation of 5-allenylisoxazoles 4 that have never been synthesized. N-Propargyl aniline 5 was successfully converted to the corresponding ortho-allenyl aniline 6 under the current reaction conditions.

Regulation of Gene Expression through Gut Microbiota-Dependent DNA Methylation in Colonic Epithelial Cells.

A huge number of commensal bacteria inhabit the intestine, which is equipped with the largest immune system in the body. Recently, the regulation of various physiological functions of the host by these bacteria has attracted attention. In this study, the effects of commensal bacteria on gene expression in colonic epithelial cells (CoECs) were investigated with focus on regulation of DNA methylation. RNA sequencing analyses of CoECs from conventional, germ-free, and MyD88-/- mice indicated that, out of the genes affected by commensal bacteria, those downregulated in a MyD88-independent manner were most frequently observed. Furthermore, when the 5' regions of genes downregulated by commensal bacteria in CoECs were captured using a customized array and immunoprecipitated with the anti-methyl cytosine Ab, a certain population of these genes was found to be highly methylated. Comprehensive analysis of DNA methylation in the 5' regions of genes in CoECs from conventional and germ-free mice upon pull-down assay with methyl-CpG-binding domain protein 2 directly demonstrated that DNA methylation in these regions was influenced by commensal bacteria. Actually, commensal bacteria were shown to control expression of Aldh1a1, which encodes a retinoic acid-producing enzyme and plays an important role in the maintenance of intestinal homeostasis via DNA methylation in the overlapping 5' region of Tmem267 and 3110070M22Rik genes in CoECs. Collectively, it can be concluded that regulation of DNA methylation in the 5' regions of a specific population of genes in CoECs acts as a mechanism by which commensal bacteria have physiological effects on the host.

Suppressive effect of dietary resistant maltodextrin on systemic immunity in a mouse model of food allergy.

Resistant maltodextrin (RMD) is a soluble dietary fibre that exerts several physiological functions as a result of its microbial degradation and changes in the intestinal environment. It has been reported that RMD enhanced immunoglobulin A (IgA) secretion, which protects the mucosa from foreign substances. However, the effect of RMD on excessive immunity has yet to be investigated. In this study, we aimed to investigate the effect of RMD on excessive immune responses such as food allergy. OVA23-3 mice were fed an AIN-76-based diet containing 20% egg-white protein with or without RMD. While RMD was shown to contribute to an increase in goblet cells, RMD did not change the overall inflammatory status when ingested with the egg-white diet. RMD suppressed IL-4 and IL-10 production from splenocytes but not cells from mesenteric lymph nodes. RMD also downregulated the serum levels of OVA-specific Th1- and Th2-related antibodies, which were elevated in the food-allergic condition. RMD significantly increased the total amount of short-chain fatty acids, especially acetate and propionate, in the caecum of OVA23-3 mice fed the egg-white diet. Our study demonstrated that dietary RMD modulates systemic rather than intestinal antigen-specific immune responses in the food-allergic condition of OVA23-3 mice. Although the relevant mechanism has yet to be investigated, RMD shows potential for alleviating food allergy through adjustment of systemic immunity.

A role for BATF3 in TH9 differentiation and T-cell-driven mucosal pathologies.

T helper 9 (TH9) cells are important for the development of inflammatory and allergic diseases. The TH9 transcriptional network converges signals from cytokines and antigen presentation but is incompletely understood. Here, we identified TL1A, a member of the TNF superfamily, as a strong inducer of mouse and human TH9 differentiation. Mechanistically, TL1A induced the expression of the transcription factors BATF and BATF3 and facilitated their binding to the Il9 promoter leading to enhanced secretion of IL-9. BATF- and BATF3-deficiencies impaired IL-9 secretion under TH9 and TH9-TL1A-polarizing conditions. In vivo, using a T-cell transfer model, we demonstrated that TL1A promoted IL-9-dependent, TH9 cell-induced intestinal and lung inflammation. Neutralizing IL-9 antibodies attenuated TL1A-driven mucosal inflammation. Batf3-/- TH9-TL1A cells induced reduced inflammation and cytokine expression in vivo compared to WT cells. Our results demonstrate that TL1A promotes TH9 cell differentiation and function and define a role for BATF3 in T-cell-driven mucosal inflammation.

Dietary Fructo-Oligosaccharides Attenuate Early Activation of CD4+ T Cells Which Produce both Th1 and Th2 Cytokines in the Intestinal Lymphoid Tissues of a Murine Food Allergy Model.

BACKGROUND: Fructo-oligosaccharides (FOS) are prebiotic agents with immunomodulatory effects involving improvement of the intestinal microbiota and metabolome. In this study, we investigated the cellular mechanisms through which FOS modulate intestinal antigen-specific CD4+ T cell responses in food allergy, using OVA23-3 mice. METHODS: OVA23-3 mice were fed an experimental diet containing either ovalbumin (OVA) or OVA and FOS for 1 week. Body weight and mucosal mast cell protease 1 in the serum were measured as the indicator of intestinal inflammation. Single-cell suspensions were prepared from intestinal and systemic lymphoid tissues for cellular analysis. Cytokine production was measured by ELISA. Activation markers and intracellular cytokines in CD4+ T cells were analyzed by flow cytometry. Activated CD4+ T cells were purified to examine cytokine production. RESULTS: Dietary intake of FOS provided moderate protection from the intestinal inflammation induced by the OVA-containing diet. FOS significantly reduced food allergy-induced Th2 cytokine responses in intestinal tissues but not in systemic tissues. FOS decreased OVA diet-induced IFN-γ+IL-4+ double-positive CD4+ T cells and early-activated CD45RBhighCD69+CD4+ T cells in the mesenteric lymph nodes. Furthermore, we confirmed that these CD45RBhighCD69+CD4+ T cells are able to produce high levels of IFN-γ and moderate level of IL-4, IL-10, and IL-13. CONCLUSIONS: Dietary intake of FOS during the development of food allergy attenuates the induction of intestinal Th2 cytokine responses by regulating early activation of naïve CD4+ T cells, which produce both Th1 and Th2 cytokines. Our results suggest FOS might be a potential food agent for the prevention of food allergy by modulating oral sensitization to food antigens.

Commensal microbiota-induced microRNA modulates intestinal epithelial permeability through the small GTPase ARF4.

The intestinal tract contains many commensal bacteria that modulate various physiological host functions. Dysbiosis of commensal bacteria triggers dysfunction of the intestinal epithelial barrier, leading to the induction or aggravation of intestinal inflammation. To elucidate whether microRNA plays a role in commensal microbiome-dependent intestinal epithelial barrier regulation, we compared transcripts in intestinal epithelial cells (IECs) from conventional and germ-free mice and found that commensal bacteria induced the expression of miR-21-5p in IECs. miR-21-5p increased intestinal epithelial permeability and up-regulated ADP ribosylation factor 4 (ARF4), a small GTPase, in the IEC line Caco-2. We also found that ARF4 expression was up-regulated upon suppression of phosphatase and tensin homolog (PTEN) and programmed cell death 4 (PDCD4), which are known miR-21-5p targets, by RNAi. Furthermore, ARF4 expression in epithelial cells of the large intestine was higher in conventional mice than in germ-free mice. ARF4 suppression in the IEC line increased the expression of tight junction proteins and decreased intestinal epithelial permeability. These results indicate that commensal microbiome-dependent miR-21-5p expression in IECs regulates intestinal epithelial permeability via ARF4, which may therefore represent a target for preventing or managing dysfunction of the intestinal epithelial barrier.

A Bacterial Artificial Chromosome Reporter System for Expression of the Human FOXP3 Gene in Mouse Regulatory T-Cells.

The transcription factor FOXP3 plays key roles in the development and function of regulatory T cells (Treg) capable of preventing and correcting immunopathology. There has been much interest in exploiting Treg as adoptive cell therapy in man, but issues of lack of nominal antigen-specificity and stability of FoxP3 expression in the face of pro-inflammatory cytokines have been a concern. In order to enable fundamental studies of human FOXP3 (hFOXP3) gene regulation and to provide preclinical tools to guide the selection of drugs that might modulate hFOXP3 expression for therapeutic purposes, we generated hFOXP3/AmCyan bacterial artificial chromosome (BAC) transgenic mice and transfectants, wherein hFOXP3 expression was read out as AmCyan expression. Using the transgenic mice, one can now investigate hFOXP3 gene expression under defined experimental conditions used for mouse Foxp3 (mFoxp3) studies. Here, we demonstrate that hFOXP3 gene expression in BAC transgenic mice is solely restricted to CD4+ T-cells, as for mFoxp3 gene expression, showing that hFOXP3 expression in Treg cells depends on fundamentally similar processes to mFoxp3 expression in these cells. Similarly, hFOXP3 expression could be observed in mouse T-cells through TCR stimulation in the presence of TGF-ß. These data suggest that, at least in part, cell type-specific human and mouse foxp3 gene expression is regulated by common regulatory regions which for the human, are located within the 110-kb human FOXP3 BAC DNA. To investigate hFOXP3 gene expression further and to screen potential therapeutics in modulating hFOXP3 gene expression in vitro, we also generated hFOXP3/AmCyan expression reporter cell lines. Using the reporter cells and transcription factor inhibitors, we showed that, just as for mFoxp3 expression, inhibitors of NF-κB, AP1, STAT5, Smad3, and NFAT also block hFOXP3 expression. hFOXP3 induction in the reporter cells was also TGF-ß dependent, and substantially enhanced by an mTOR inhibitor, Torin1. In both the reporter transgenic mice and cell lines, histone H4 molecules in the hFOXP3 promoter and enhancers located in human CNS1 and CNS2 regions were highly acetylated in natural Treg and TCR/TGF-ß-induced Treg, indicating hFOXP3 gene expression is regulated by mechanisms similar to those previously identified for the mFoxp3 gene.

The TNF family member TL1A induces IL-22 secretion in committed human Th17 cells via IL-9 induction.

TL1A contributes to the pathogenesis of several chronic inflammatory diseases, including those of the bowel by enhancing TH1, TH17, and TH2 responses. TL1A mediates a strong costimulation of these TH subsets, particularly of mucosal CCR9+ T cells. However, the signaling pathways that TL1A induces in different TH subsets are incompletely understood. We investigated the function of TL1A on human TH17 cells. TL1A, together with TGF-ß, IL-6, and IL-23, enhanced the secretion of IL-17 and IFN-γ from human CD4+ memory T cells. TL1A induced expression of the transcription factors BATF and T-bet that correlated with the secretion of IL-17 and IFN-γ. In contrast, TL1A alone induced high levels of IL-22 in memory CD4+ T cells and committed TH17 cells. However, TL1A did not enhance expression of IL-17A in TH17 cells. Expression of the transcription factor aryl hydrocarbon receptor, which regulates the expression of IL-22 was not affected by TL1A. Transcriptome analysis of TH17 cells revealed increased expression of IL-9 in response to TL1A. Blocking IL-9 receptor antibodies abrogated TL1A-induced IL-22 secretion. Furthermore, TL1A increased IL-9 production by peripheral TH17 cells isolated from patients with Crohn's disease. These data suggest that TL1A differentially induces expression of TH17 effector cytokines IL-17, -9, and -22 and provides a potential target for therapeutic intervention in TH17-driven chronic inflammatory diseases.

α-Defensin 5 gene expression is regulated by gut microbial metabolites.

α-Defensin 5 is important to both maintenance of a gut microbiota and host immunity. While previous reports have shown that gut bacteria are able to upregulate α-defensin 5 through Toll-like receptor signaling, we demonstrate here that α-defensin 5 expression can also be regulated by microbial metabolites. Among these, lactate appeared to significantly suppress α-defensin 5 gene transcription. Actually, fractions of <3 kD compounds obtained from the ceca of SPF mice were suppressed α-defensin 5 gene transcription at specific concentrations. Our results also suggest that cecal content may include as yet unidentified factors that can enhance α-defensin 5 expression. Our data point to a novel function for the gut microbial metabolites in controlling the expression of antimicrobial peptides in the intestine.

Post-Transcriptional Regulation of Toll-Interacting Protein in the Intestinal Epithelium.

Immune responses against gut microbiota should be minimized to avoid unnecessary inflammation at mucosal surface. In this study, we analyzed the expression patterns of Toll-interacting protein (Tollip), an inhibitor of TLRs and IL-1 family cytokine-related intracellular signaling, in intestinal epithelial cells (IECs). Comparable mRNA expression was observed in murine small and large IECs (S-IECs and L-IECs). However, Tollip protein was only detected in L-IECs, but not in S-IECs. Similar results were obtained in germ-free mice, indicating that L-IEC-specific TOLLIP expression does not depend on bacterial colonization. Next, to understand the mechanisms underlying the post-transcriptional repression of Tollip, 3´-UTR-mediated translational regulation was evaluated. The region +1876/+2398 was responsible for the repression of Tollip expression. This region included the target sequence of miR-31. The inhibition of miR-31 restored the 3´-UTR-meditaed translational repression. In addition, miR-31 expression was significantly higher in S-IECs than in L-IECs, suggesting that miR-31 represses the translation of Tollip mRNA in S-IECs. Collectively, we conclude that the translation of Tollip is inhibited in S-IECs, at least in part, by miR-31 to yield L-IEC-specific high-level expression of the Tollip protein, which may contribute to the maintenance of intestinal homeostasis.

Gene expression in the Gitr locus is regulated by NF-κB and Foxp3 through an enhancer.

Glucocorticoid-induced TNFR (Gitr) and Ox40, two members of the TNFR superfamily, play important roles in regulating activities of effector and regulatory T cells (Treg). Their gene expression is induced by T cell activation and further upregulated in Foxp3+ Treg. Although the role of Foxp3 as a transcriptional repressor in Treg is well established, the mechanisms underlying Foxp3-mediated transcriptional upregulation remain poorly understood. This transcription factor seems to upregulate expression not only of Gitr and Ox40, but also other genes, including Ctla4, Il35, Cd25, all critical to Treg function. To investigate how Foxp3 achieves such upregulation, we analyzed its activity on Gitr and Ox40 genes located within a 15.1-kb region. We identified an enhancer located downstream of the Gitr gene, and both Gitr and Ox40 promoter activities were shown to be upregulated by the NF-κB-mediated enhancer activity. We also show, using the Gitr promoter, that the enhancer activity was further upregulated in conjunction with Foxp3. Foxp3 appears to stabilize NF-κB p50 binding by anchoring it to the enhancer, thereby enabling local accumulation of transcriptional complexes containing other members of the NF-κB and IκB families. These findings may explain how Foxp3 can activate expression of certain genes while suppressing others.

TGF-ß-mediated Foxp3 gene expression is cooperatively regulated by Stat5, Creb, and AP-1 through CNS2.

Foxp3 plays an important role in the development and the function of regulatory T cells (Treg). Both the induction and maintenance of Foxp3 gene expression are controlled by several regulatory regions including two enhancers in the conserved noncoding sequences (CNS). The functions of Enhancer 1 in CNS1 are well established, whereas those of Enhancer 2 in CNS2 remain unclear. Although CNS2 contains enhancer activity, methylated CpG sequences in this region prevent Foxp3 gene expression in Foxp3(-) T cells. These sequences are, however, demethylated in Foxp3(+) Treg by mechanisms as yet unknown. To investigate the role of CNS2, we have determined the Enhancer 2 core sequence by luciferase reporter assays in the absence of methylation to exclude the inhibitory effect and shown that transcription factors AP-1, Stat5, and Creb cooperate in regulating Enhancer 2 activity. We have then determined the methylation sensitivity of each of the transcription factors. AP-1 was found to be methylation sensitive as has previously been described for Creb. However, Stat5 was active even when its binding site in CNS2 was methylated. Stat5 binding to Enhancer 2 occurred early and preceded that of AP-1 and Creb during Treg induction. In addition, Stat5 activation is itself dependent on TGF-ß signaling through Smad3-mediated blockade of Socs3 expression. These findings suggest that Stat5 is a key regulator for opening up the CNS2 region during induced Treg induction, whereas AP-1 and Creb maintain Enhancer 2 activity.

IgA production in the large intestine is modulated by a different mechanism than in the small intestine: Bacteroides acidifaciens promotes IgA production in the large intestine by inducing germinal center formation and increasing the number of IgA+ B cells.

It has been demonstrated that intestinal commensal bacteria induce immunoglobulin (Ig) A production by promoting the development of gut-associated lymphoid tissues in the small intestine. However, the precise mechanism whereby these bacteria modulate IgA production in the large intestine, which harbors the majority of intestinal commensals, is poorly understood. In addition, it is not known which commensal bacteria induce IgA production in the small intestine and which induce production in the large intestine. To address these issues, we generated gnotobiotic mice mono-associated with different murine commensal bacteria by inoculating germ-free (GF) mice with Lactobacillus johnsonii or Bacteroides acidifaciens. In GF mice, IgA production was barely detectable in the small intestine and was not detected in the large intestine. Interestingly, total IgA secretion in the large intestinal mucosa of B. acidifaciens mono-associated (BA) mice was significantly greater than that of GF and L. johnsonii mono-associated (LJ) mice. However, there was no difference in total IgA production in the small intestine of GF, LJ and BA mice. In addition, in the large intestine of BA mice, the expression of IgA(+) cells and germinal center formation were more remarkable than in GF and LJ mice. Furthermore, B. acidifaciens-specific IgA was detected in the large intestine of BA mice. These results suggest that the production of IgA in the large intestine may be modulated by a different mechanism than that in the small intestine, and that B. acidifaciens is one of the predominant bacteria responsible for promoting IgA production in the large intestine.