[show abstract][hide abstract] ABSTRACT: Dectin-1, which specifically recognizes β-glucan of fungal cell walls, is a non-Toll-like receptor (TLR) pattern recognition receptor and a representative of C-type lectin receptors (CLRs). The importance of Dectin-1 in innate immune cells, such as dendritic cells and macrophages, has previously been well studied. However, the function of Dectin-1 in B cells is very poorly understood. To determine the role of Dectin-1 in B cell activation, we first investigated whether mouse B cells express Dectin-1 and then assessed the effect of Dectin-1 stimulation on B cell proliferation and antibody production. Mouse B cells express mRNAs encoding CLRs, including Dectin-1, and surface Dectin-1 was expressed in B cells of C57BL/6 rather than BALB/c strain. Dectin-1 agonists, heat-killed Candida albicans (HKCA) and heat-killed Saccharomyces cerevisiae (HKSC), alone induced B cell proliferation but not antibody production. Interestingly, HKSC, HKCA, and depleted zymosan (a selective Dectin-1 agonist) selectively enhanced LPS-driven IgG1 production. Taken together, these results suggest that, during fungal infection, β-glucan-stimulated Dectin-1 may cooperate with TLR4 to specifically enhance IgG1 production by mouse B cells.
[show abstract][hide abstract] ABSTRACT: The present study demonstrates that RA has activity of an IgA switch factor and is more specific than TGF-β1. RA independently caused only IgA switching, whereas TGF-β1 caused IgA and IgG2b switching. We found that RA increased IgA production and that this was a result of its ability to increase the frequency of IgA-secreting B cell clones. Increased IgA production was accompanied by an increase of GLTα. RA activity was abrogated by an antagonist of the RAR. Additionally, RA affected intestinal IgA production in mice. Surprisingly, RA, in combination with TGF-β1, notably enhanced not only IgA production and GLTα expression but also CCR9 and α4β7 expression on B cells. These results suggest that RA selectively induces IgA isotype switching through RAR and that RA and TGF-β have important effects on the overall gut IgA antibody response.
Journal of leukocyte biology 06/2013; · 4.99 Impact Factor
[show abstract][hide abstract] ABSTRACT: Activation-induced cytidine deaminase (AID) plays a key role in B cell immunoglobulin (Ig) class switch recombination (CSR) and somatic hypermutation (SHM). We have previously reported that the highly conserved homeodomain HoxC4 transcription factor binds to the Aicda (AID gene) promoter to induce AID expression. Here, we investigated the regulation of HoxC4 transcription by a proliferation-inducing ligand (APRIL) and B cell-activating factor belonging to the TNF family (BAFF) in mouse B cells. APRIL substantially increased both HoxC4 and AID expression, whereas BAFF induced the expression of AID but not HoxC4. To elucidate the underlying mechanisms, we constructed a HoxC4 gene promoter reporter vector and analyzed the promoter induction after APRIL stimulation. APRIL enhanced the HoxC4 promoter activity by 2.3-fold, and this increase disappeared when the second putative NF-κB-binding promoter element (NBE2) was mutated. Based on ChIP assays, we found that NF-κB bound to the HoxC4 promoter NBE2 region. Furthermore, the overexpression of NF-κB augmented the APRIL-induced HoxC4 promoter activity, while the expression of dominant negative-IκBα suppressed it. Taken together, our findings suggest that NF-κB mediates APRIL-induced HoxC4 transcription.
[show abstract][hide abstract] ABSTRACT: Immune cells express toll-like receptors (TLRs) and respond to molecular patterns of various pathogens. CpG motif in bacterial DNA activates innate and acquired immune systems through binding to TLR9 of immune cells. Several studies reported that CpG can directly regulate B cell activation, differentiation, and Ig production. However, the role of CpG in B cell growth and Ig production is not fully understood. In this study, we analyzed the effect of CpG on the kinetics of mouse B cell viability, proliferation, and Igs production. Overall, CpG enhanced mouse B cell growth and production of Igs in a dose-dependent manner. Unlike LPS, 100 nM CpG (high dose) did not support TGF-β1-induced IgA and IgG2b production. Moreover, 100 nM CpG treatment abrogated either LPS-induced IgM or LPS/TGF-β1-induced IgA and IgG2b production, although B cell growth was enhanced by CpG under the same culture conditions. We subsequently found that 10 nM CpG (low dose) is sufficient for B cell growth. Again, 10 nM CpG did not support TGF-β1-induced IgA production but, interestingly enough, supported RA-induced IgA production. Further, 10 nM CpG, unlike 100 nM, neither abrogated the LPS/TGF-β1-nor the LPS/RA-induced IgA production. Taken together, these results suggest that dose of CpG is critical in B cell growth and Igs production and the optimal dose of CpG cooperates with LPS in B cell activation and differentiation toward Igs production.
[show abstract][hide abstract] ABSTRACT: The inhibitory effect of polyphenol extracts (Seapolynol(™), SPN) of the marine brown algae Ecklonia cava and dieckol, a major component of SPN, on hyperlipidemia was investigated in ICR mice fed a high-fat diet (HFD) for five weeks. For analysis of the anti-hyperlipidemic effects of SPN and dieckol, these two agents were given orally on a daily basis to HFD-fed mice for four weeks, starting one week after the beginning of HFD feeding. Groups administered with SPN as well as dieckol showed lower body weight gains than the HFD only group. Administration of SPN and dieckol also resulted in a significant reduction of the level of total cholesterol (TCHO), triglyceride (TG), and low-density lipoprotein (LDL) cholesterol in the serum of HFD-fed mice. In Oil Red O staining using 3T3-L1 preadipocytes, it was shown that both SPN and dieckol markedly inhibited lipid accumulation of 3T3-L1 cells. Furthermore, SPN and dieckol (50 μg/mL) significantly inhibited 3-hydroxyl-methyl glutaryl coenzyme A (HMGCoA) reductase activity in vitro. Taken together, these results suggest that polyphenols of Ecklonia cava (SPN) and dieckol reduce body weight gain and fat accumulation in HFD-induced obese mice, and that their hypolipidemic effect is related to the inhibition of adipogenesis of adipocytes and HMGCoA reductase activity.
Preventive nutrition and food science. 03/2012; 17(1):1-7.
[show abstract][hide abstract] ABSTRACT: Angiogenesis is a multi-step process that involves the activation, proliferation, and migration of endothelial cells. We have recently shown that TGF-beta1 can induce mouse macrophages to produce VEGF, a potent angiogenic factor. In the present study, we explored whether TGF-beta1 has a similar effect on mouse dendritic cells. First, we show that under hypoxic conditions, TGF-beta1 induced the expression of VEGF transcripts in bone marrow-derived dendritic cells. Overexpression of Smad3/4 further augmented TGF-beta1-induced VEGF transcription, while overexpression of DN-Smad3 decreased VEGF transcription in DC2.4 cells, a mouse dendritic cell line. We also show that TGF-beta1 and Smads are involved in the induction of VEGF protein secretion. Interestingly, under the same conditions, the expression of VEGF receptor 1 (Flt-1) was also elevated at both the transcriptional and protein levels. Additionally, we found that the TGF-beta1-induced VEGF secretion in activated DC2.4 cells has wound-healing properties. Finally, Smad7 and Smurf1 negatively regulated the TGF-beta1-induced and Smad3/4-mediated VEGF expression. Taken together, these results indicate that TGF-beta1 can enhance the expression of VEGF and Flt-1 through the typical Smad pathway in mouse dendritic cells.
Experimental and Molecular Medicine 09/2010; 42(9):606-13. · 2.57 Impact Factor
[show abstract][hide abstract] ABSTRACT: Class switch DNA recombination (CSR) is the mechanism that diversifies the biological effector functions of antibodies. Activation-induced cytidine deaminase (AID), a key protein in CSR, targets immunoglobulin H (IgH) switch regions, which contain 5'-AGCT-3' repeats in their core. How AID is recruited to switch regions remains unclear. Here we show that 14-3-3 adaptor proteins have an important role in CSR. 14-3-3 proteins specifically bound 5'-AGCT-3' repeats, were upregulated in B cells undergoing CSR and were recruited with AID to the switch regions that are involved in CSR events (Smu-->Sgamma1, Smu-->Sgamma3 or Smu-->Salpha). Moreover, blocking 14-3-3 by difopein, 14-3-3gamma deficiency or expression of a dominant-negative 14-3-3sigma mutant impaired recruitment of AID to switch regions and decreased CSR. Finally, 14-3-3 proteins interacted directly with AID and enhanced AID-mediated in vitro DNA deamination, further emphasizing the important role of these adaptors in CSR.
[show abstract][hide abstract] ABSTRACT: p300 acts as a coactivator for TGF-β1-induced Ig germ-line (GL) α transcription. To understand the exact role of p300, we
analyzed known domains of p300 using several deletion mutants. p300 mutants lacking both the histone acetyltransferase (HAT)
and Smad-binding domains did not enhance GLα promoter activity. In addition, Smad-binding domain within p300 was not sufficient
for the promoter activity. Surprisingly, overexpression of a p300 mutant retaining the HAT domain but lacking the rest of
downstream segment, p300[N1736], greatly augmented the GLα promoter activity, regardless of TGF-β1 treatment, to levels much
higher than that of p300[WT]. Together, these results suggest that HAT activity of p300 is required for GLα promoter activity.
We found that p300[N1736] binds to the GLα promoter using ChIP assays. Further analysis of p300[N1736] indicated that both
Stat1 and HDAC1 can inhibit the enhancing effect of p300[N1736], suggesting the importance of the Stat1-binding domain and
the HAT domain. E1A, a repressor of p300[WT], had no effect on the activity of p300[N1736], which also lacks the E1A-binding
domain. Finally, when the amount of transfected p300[N1736] was reduced 40-fold, p300[N1736] functioned similarly to overexpressed
p300[WT]. Thus, p300 [N1736], though not having a Smad3-binding domain, augmented TGF-β1-induced GLα promoter activity, which
was inhibited by IFN-γ/Stat-1. Taken together, these results indicate that p300 acts as a basal element in GLα transcription
without binding to Smad3. Additionally, E1A, Stat1, and HDAC1 act as repressors of p300 in such transcription.
Keywordsp300-Germ-line α transcripts-IgA-TGF-β1-Smad3-FN-γ
[show abstract][hide abstract] ABSTRACT: IFN-gamma has been shown to either up- or down-regulate the expression of specific TGF-beta1-induced target genes. We investigated the effect of IFN-gamma on TGF-beta1-induced IgA isotype expression. We found that IFN-gamma inhibited not only TGF-beta1-induced germ-line (GL) alpha transcription, but also IgA secretion by TGF-beta1-stimulated murine B cells. Overexpression of Stat1 diminished TGF-beta1-induced, Smad3/4-and Runx3-mediated GL alpha promoter activity. Overexpression of p300 also increased the promoter activity, while its effect was abrogated by co-transfected Stat1. Stat1 interfered with the Smad3:p300 interaction, likely due to a stronger Stat1:p300 binding affinity. These results indicate that Stat1 can inhibit GL alpha transcription through binding to p300. Further, overexpression of SOCS1, a JAK inhibitor, diminished the antagonistic effect of IFN-gamma on TGF-beta1-induced GL alpha transcription and IgA secretion. These results indicate that JAK/Stat1-mediated IFN-gamma signaling antagonizes TGF-beta1-induced GL alpha transcription, mainly through deprivation of p300 from Smad3, resulting in decreased IgA synthesis.
Molecules and Cells 12/2009; 29(1):57-62. · 2.21 Impact Factor
[show abstract][hide abstract] ABSTRACT: APRIL (a proliferation-inducing ligand) is primarily expressed by macrophages and dendritic cells, and has profound effects on B cell physiology. In this study, we investigated the role of IL-4 in APRIL expression by mouse macrophages and the signaling mechanism involved. IL-4 markedly enhanced APRIL expression in mouse macrophages at the transcriptional and protein level. The p38MAPK inhibitor SB203580 completely abolished the IL-4 effect, whereas overexpression of CREB with IL-4 augmented APRIL expression. This increase was abolished by SB203580 treatment, indicating that p38MAPK may activate CREB. Overexpression of Stat6 also augmented IL-4-induced APRIL expression; this effect was partially abolished by SB203580 but not by the Jak inhibitor AG490, indicating that Stat6 mediates IL-4-induced APRIL expression in a Jak-independent manner and that p38MAPK acts as the intermediate. Our results demonstrate that IL-4 up-regulates APRIL expression through two divergent pathways in mouse macrophages, p38MAPK-CREB and p38MAPK-Stat6.
[show abstract][hide abstract] ABSTRACT: The cytidine deaminase AID (encoded by Aicda in mice and AICDA in humans) is critical for immunoglobulin class-switch recombination (CSR) and somatic hypermutation (SHM). Here we show that AID expression was induced by the HoxC4 homeodomain transcription factor, which bound to a highly conserved HoxC4-Oct site in the Aicda or AICDA promoter. This site functioned in synergy with a conserved binding site for the transcription factors Sp1, Sp3 and NF-kappaB. HoxC4 was 'preferentially' expressed in germinal center B cells and was upregulated by engagement of CD40 by CD154, as well as by lipopolysaccharide and interleukin 4. HoxC4 deficiency resulted in impaired CSR and SHM because of lower AID expression and not some other putative HoxC4-dependent activity. Enforced expression of AID in Hoxc4(-/-) B cells fully restored CSR. Thus, HoxC4 directly activates the Aicda promoter, thereby inducing AID expression, CSR and SHM.
[show abstract][hide abstract] ABSTRACT: TGF-beta1 directs class switch recombination to IgG2b as well as IgA. We have shown that Smad3/4, Runx3, and p300 mediate TGF-beta1-induced germ-line (GL) gamma2b transcription and that there is a potential Smad-binding element (SBE, CAGAC, -38/-34) and Runx-binding element (TGTGGGT, +41/+47) in the promoter region. Here, we have characterized more putative transcription factor-binding elements in the promoter. Site-directed mutagenesis revealed that two more putative SBE (GTCTG, -67/-63 and +38/+42) are relevant to TGF-beta1-induced GLgamma2b promoter activity, a finding that was confirmed by EMSA. However, neither overexpression of Ets (i.e. Elf-1, Fli-1, or Pu.1) nor a mutation deleting a putative Ets-binding element (CAGGAA, -4/+2) affected basal or TGF-beta1-induced promoter activity. On the other hand, NF-kappaB repressed promoter activity without direct binding to two putative NF-kappaB-binding elements (GGACTCCCC, -63/-55; GGGCCTTTCC,+237/+246). Instead, NF-kappaB overexpression increased the expression of Smad7 transcripts. Moreover, p300 overexpression failed to rescue the inhibitory effect of NF-kappaB on GLgamma2b promoter activity. These results indicate that there are multiple SBE relevant to GLgamma2b promoter activity and that NF-kappaB acts in cooperation with p300 to downregulate promoter activity through increasing the gene expression of inhibitory Smad7.
European Journal of Immunology 04/2009; 39(4):1157-66. · 4.97 Impact Factor
[show abstract][hide abstract] ABSTRACT: Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by the production of an array of pathogenic autoantibodies, including high-affinity anti-dsDNA IgG antibodies. These autoantibodies are mutated and class-switched, mainly to IgG, indicating that immunoglobulin (Ig) gene somatic hypermutation (SHM) and class switch DNA recombination (CSR) are important in their generation. Lupus-prone MRL/fas(lpr/lpr) mice develop a systemic autoimmune syndrome that shares many features with human SLE. We found that Ig genes were heavily mutated in MRL/fas(lpr/lpr) mice and contained long stretches of DNA deletions and insertions. The spectrum of mutations in MRL/fas(lpr/lpr) B cells was significantly altered, including increased dG/dC transitions, increased targeting of the RGYW/WRCY mutational hotspot and the WGCW AID-targeting hotspot. We also showed that MRL/fas(lpr/lpr) greatly upregulated CSR, particularly to IgG2a and IgA in B cells of the spleen, lymph nodes and Peyer's patches. In MRL/fas(lpr/lpr) mice, the significant upregulation of SHM and CSR was associated with increased expression of activation-induced cytidine deaminase (AID), which mediates DNA lesion, the first step in SHM and CSR, and translesion DNA synthesis (TLS) polymerase (pol) theta, pol eta and pol zeta, which are involved in DNA synthesis/repair process associated with SHM and, possibly, CSR. Thus, in lupus-prone MRL/fas(lpr/lpr) mice, SHM and CSR are upregulated, as a result of enhanced AID expression and, therefore, DNA lesions, and dysregulated DNA repair factors, including TLS polymerases, which are involved in the repair process of AID-mediated DNA lesions.
[show abstract][hide abstract] ABSTRACT: Activation-induced cytidine deaminase (AID) is an inducible gene that plays a critical role in Ig class switch recombination and somatic hypermutation in B cells. We explored the mechanisms by which IL-4 induces AID expression in mouse B cells. IL-4 increased AID expression and over-expression of Stat6 further augmented IL-4-induced promoter activity. The involvement of Stat6 in the promoter activity was confirmed using ChIP assays and site-directed mutagenesis. Treatment with H89, a PKA inhibitor, markedly decreased IL-4-induced AID expression, and over-expression of CREB enhanced it. These results indicate that Stat6 and PKA/CREB are involved in IL-4-induced AID expression. The relevance of these signal transducing molecules was verified using the TGFbeta1-induced IgA isotype switching model. Our results indicate that IL-4, through Stat6 and PKA/CREB, induces AID expression leading to Ig isotype switching event.
Biochemical and Biophysical Research Communications 10/2007; 361(2):398-403. · 2.41 Impact Factor
[show abstract][hide abstract] ABSTRACT: Expression and activity of activation-induced cytidine deaminase (AID) encoded by the aicda gene are essential for immunoglobulin (Ig) gene somatic hypermutation (SHM) and class switch DNA recombination (CSR). SHM and CSR unfold, in general, in germinal centers and/are central to the maturation of effective antibody responses. AID expression is induced by activated B-cell CD40 signaling, which is critical for the germinal center reaction, and is further enhanced by other stimuli, including interleukin-4 (IL-4) secreted from CD4+ T cells or Toll-like receptor (TLR)-activating bacterial and/or viral molecules. Integration of different intracellular signal transduction pathways, as activated by these stimuli, leads to a dynamic aicda-regulating program, which involves both positively acting trans-factors, such as Pax5, HoxC4, E47, and Irf8, and negative modulators, such as Blimp1 and Id2, to restrict aicda expression primarily to germinal center B cells. The phosphatidylinositol 3-kinase (PI 3-K), which functions downstream of activated B-cell receptor (BCR) signaling, likely plays an important role in triggering the downregulation of aicda expression in postgerminal center B cells and throughout plasmacytoid differentiation. In B cells undergoing SHM and CSR, AID activity, and, possibly, AID targeting to the Ig locus are regulated at a posttranslational level, including AID dimerization/oligomerization, nuclear/cytoplasmic AID translocation, and phosphorylation of the AID Ser38 residue by protein kinase A (PKA). Here, we discuss the role of B-cell activation signals, transcription regulation programs, and posttranslational modifications in controlling aicda expression and AID activity, thereby delineating an integrated model of modulation of SHM and CSR in the germinal center reaction.
Critical Reviews in Immunology 02/2007; 27(4):367-97. · 3.38 Impact Factor
[show abstract][hide abstract] ABSTRACT: Activation-induced cytidine deaminase (AID) is needed for Ig class switch recombination (CSR). We explored the effect of LPS on the expression of AID during B cell differentiation, and the role of AID in IgA isotype expression. In normal spleen B cells, LPS increased AID transcription up to 48 h post-stimulation, i.e. around the time of Ig CSR. TGF-b1 and AID were required for IgA expression, and LPS contributed to TGFb1-induced IgA production largely by inducing AID. Interestingly, LPS repressed AID transcription in sIgA+ B cells but still stimulated IgA production mainly by increasing the rate of IgA secretion. Our data indicate that LPS contributes to TGFb1-induced IgA isotype expression in at least two ways: by stimulating AID transcription before CSR and by enhancing the IgA secretion rate after CSR.
Molecules and Cells 07/2005; 19(3):445-51. · 2.21 Impact Factor
[show abstract][hide abstract] ABSTRACT: Transforming growth factor (TGF)-beta1 directs class switch recombination (CSR) to IgG2b as well as to IgA. Smad3/4, Runx3 and p300 mediate TGF-beta1-induced germ-line (GL) alpha transcription leading to IgA expression. However, the molecular mechanisms by which TGF-beta1 induces IgG2b CSR are unknown. We used luciferase reporter plasmids to investigate how TGF-beta1 regulates the activity of the promoter for GL transcripts of IgG2b constant gene (GLgamma2b promoter). Similarly to the GLalpha promoter, overexpression of Smad3/4 and Runx3 enhances TGF-beta1-induced GLgamma2b promoter activity. Mutation analysis of the promoter identified likely Smad- and Runx3-binding sites. Also similar to the GLalpha promoter, overexpression of p300 enhances Smad3/4-mediated promoter activity, whereas E1A represses promoter activity. Since these regulation mechanisms underlying both GLalpha and GLgamma2b transcription are similar, we explored the possibility that TGF-beta1 induces IgA CSR via transitional IgG2b CSR. TGF-beta1 enhances the expression of both Ialpha-Cmu and Ialpha-Cgamma2b circle transcripts, indicative of direct (Smu-->Salpha) and sequential CSR (Smu-->Sgamma2b-->Salpha).
European Journal of Immunology 04/2005; 35(3):946-56. · 4.97 Impact Factor
[show abstract][hide abstract] ABSTRACT: We have shown previously that Smad3 and Smad4 mediate TGF-beta1-induced IgA expression. In the present study, we examined the involvement of Runx3 in this process. Overexpression of Runx3 in mice increased germ-line alpha (GLalpha) transcription, and transcription was further augmented when B lymphoma and LPS-activated murine spleen cells were co-transfected with Smad3/4. Overexpression of Runx3 and Smad3/4 increased IgA secretion by both cell types in response to TGF-beta1. p300, which has histone acetyltransferase activity, further augmented TGF-beta1-induced GLalpha transcription promoted by Smad3/4 and Runx3. These observations were confirmed by examining the influence of Smad3/4, Runx3 and p300 on the expression of endogenous GLalpha and post-switch alpha transcripts.E1A, an inhibitor of p300, blocked both GLalpha promoter activity and the enhancement of endogenous GLalpha transcription by Smad3/4 and Runx3. We conclude that p300 cooperates with Smad3/4 and Runx3 in stimulating TGF-beta1-induced GLalpha transcription and subsequent IgA isotype expression, while E1A inhibits these cooperative effects.
European Journal of Immunology 01/2004; 33(12):3386-92. · 4.97 Impact Factor
[show abstract][hide abstract] ABSTRACT: Antibody class switch recombination (CSR) occurs after antigen activation of B cells. CSR is directed to specific heavy chain isotypes by cytokines and B cell activators that induce transcription from the unrearranged, or germline (GL), CH region genes. Transforming growth factor (TGF)-β1 is essential for switch recombination to IgA due to its ability to induce transcription from GL Ig α genes. It has been shown that the promoters which regulate transcription of mouse and human GL α RNAs contain a TGF-β1-responsive element that binds Smad and core binding factor (CBFα)/AML/PEBPα/Runx. They also contain other elements which bind the transcription factors CREB, BSAP and Ets family proteins. In this manuscript we demonstrate that two tandem Ets sites in the mouse GL α promoter bind the transcription factors Elf-1 and PU.1, and that the 3 site is essential for expression of a luciferase reporter gene driven by the GL α promoter. Binding of Elf-1 to the GL α promoter is inducible by lipopolysaccharide in nuclear extracts from splenic B cells. An NF-κB site is identified, although it does not contribute to expression of the promoter in reporter gene assays. Since CSR to IgA is greatly reduced in NF-κB/p50-deficient mice, these data support the hypothesis that NF-κB has roles in switching in addition to regulation of GL transcription. Finally, we demonstrate that nocodazole, which disrupts microtubules that sequester Smad proteins in the cytoplasm, stimulates transcription from the GL α promoter.
[show abstract][hide abstract] ABSTRACT: Retinoic acid (RA) is considered to possess an activity of IgA isotype switching. Thus far, TGF-β1 is known to be the most
powerful IgA isotype switch factor. To elucidate the molecular mechanisms underlying the Ig germ line (GL) α transcriptional
regulation by RA, we constructed three different sizes of mouse GLα promoter reporters; short-GLα(−130/+14), middle-GLα(−448/+72)
and long-GLα(−3028/+72). Based on luciferase assay, RA increased the activity of all three GLα promoter reporters by approximately
2-fold and the effect was further enhanced by TGF-β1. Overexpression of Smad3/4 increased TGF-β1-induced GLα promoter activities
but had no effect on RA-induced GLα promoter activities. In order to analyze the characteristics of the RA-inducible GLα promoter
region, we also constructed two mutant reporters: Smad3 binding elements (SBEs)-substituted short-GLα (short-GLα mSBE) and
Runx3 binding elements (RBEs)-substituted short-GLα (short-GLα mRBE) promoter reporters. Promoter activities of the two mutant
reporters to RA were comparable to that of wild type reporter, while those of the two mutant reporters to TGF-β1 were markedly
diminished as compared to that of WT short-GLα. Finally, RA-induced GLα transcription was virtually disappeared by LE540,
an antagonist of RA receptor (RAR). Taken together, these results suggest that RA induces GLα transcription mainly through
RAR pathway, where neither Smad3/4 nor Runx3 is involved.
KeywordsRetinoic acid–TGF-β1–GLα promoter–IgA–Smad–RA receptor