IFN-γ Down-Regulates TGF-β1-Induced IgA Expression through Stat1 and p300 Signaling

박석래,Mee-Hyeun Jung,Seong-Hyun Jeon,Mi-Hee Park,Kyoung-Hoon Park,Mi-Ra Lee,Pyeung-Hyeun Kim 한국분자세포생물학회 2010 Molecules and cells Vol.29 No.1

IFN-γ has been shown to either up- or down-regulate the expression of specific TGF-β1-induced target genes. We investigated the effect of IFN-γ on TGF-β1-induced IgA isotype expression. We found that IFN-γ inhibited not only TGF-β1-induced germ-line (GL) α transcription, but also IgA secretion by TGF-β1-stimulated murine B cells. Overexpression of Stat1 diminished TGF-β1-induced, Smad3/4-and Runx3-mediated GLα 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 GLα transcription through binding to p300. Further, overexpression of SOCS1, a JAK inhibitor,diminished the antagonistic effect of IFN-γ on TGF-β1-induced GLα transcription and IgA secretion. These results indicate that JAK/Stat1-mediated IFN-γ signaling antagonizes TGF-β1-induced GLα transcription, mainly through deprivation of p300 from Smad3, resulting in decreased IgA synthesis.

Cloning and analysis of promoter region of mouse immunoglobulin germline c3 transcripts

박석래,서범석,윤희경,신종대,박하얀,이상훈,이주언,유영춘,이정림,김평현 한국유전학회 2016 Genes & Genomics Vol.38 No.11

The germline c3 transcripts (GLTc3) is transcribed by mouse B cells following stimulation with lipopolysaccharide (LPS). GLTc3 transcription is a prerequisite for IgG3 class switching. However, to date, the promoter activity of GLTc3 has not been investigated. In this study, we constructed GLTc3 promoter reporters and tested their promoter activities following LPS stimulation in B cells. Among the reporters, pGL3-c3(F4) had the highest basal activity and was optimally activated by LPS. Deletion of the first and second conserved sequences (CS1 and CS2) in the GLTc3 promoter abrogated LPS-induced promoter activity. In addition, DNase I-hypersensitive site 1,2 (HS1,2) further enhanced GLTc3 promoter activity. These findings indicate that CS1 and CS2 of the GLTc3 promoter are essential for LPS-induced GLTc3 transcription and that HS1,2 enhances transcription.

Activation-induced Cytidine Deaminase in B Cell Immunity and Cancers

박석래 대한면역학회 2012 Immune Network Vol.12 No.6

Activation-induced cytidine deaminase (AID) is an enzyme that is predominantly expressed in germinal center B cells and plays a pivotal role in immunoglobulin class switch recombination and somatic hypermutation for antibody (Ab)maturation. These two genetic processes endow Abs with protective functions against a multitude of antigens (pathogens)during humoral immune responses. In B cells, AID expression is regulated at the level of either transcriptional activation on AID gene loci or post-transcriptional suppression of AID mRNA. Furthermore, AID stabilization and targeting are determined by post-translational modifications and interactions with other cellular/nuclear factors. On the other hand, aberrant expression of AID causes B cell leukemias and lymphomas, including Burkitt’s lymphoma caused by c-myc/IgH translocation. AID is also ectopically expressed in T cells and non-immune cells, and triggers point mutations in relevant DNA loci, resulting in tumorigenesis. Here, I review the recent literatures on the function of AID, regulation of AID expression, stability and targeting in B cells, and AID-related tumor formation.

Kinetic Analysis of CpG-Induced Mouse B Cell Growth and Ig Production

김영하,박석래,이상훈,유영춘,이정림,박종환 대한면역학회 2012 Immune Network Vol.12 No.3

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.

TGF-$\beta$3는 마우스 IgA, IgG2b 항체의 선택적 유도작용

이은경,박석래,전계택,김평현,이세원,최의열 한국미생물학회 1999 미생물학회지 Vol.35 No.2

TGF-$\beta$1은 LPS 로 자극시킨 마우스의 spleen B cell 의 IgA와 IgG2b의 항체 합성을 선택적으로 증가시킨다고 알려져있다. 본 연구에서는 TGF-$\beta$1과 80%의 아미노산을 공유하는 TGF-$\beta$3가 마우스 spleen B cell 과 mesenteric lymph node B cell의 항체 합성에 미치는 영향을 IL-5와 함께 조사하였다. LPS로 활성화된 spleen B cell 에 TGF-$\beta$3만을 처리한 조건에서 IgA 항체합성이 약간 증가하였고, IL-5와 함께 넣어 준 배양조건에서는 IgA 항체가 현격히 증가하였다 IgG2b 합성의 증가는 TGF-$\beta$3 자극만으로도 가능하였고 IgA 와는 달리 IL-5 의 첨가 효과는 관찰되지 않았다. 한편, TGF-$\beta$3는 IgM 과 IgG1 항체 합성을 감소시켰고, IL-5와 함께 존재한 경우에도 의미있는 합성 증가는 볼 수 없었다. ELISPOT assay로 IgA 합성 세포수의 변화를 조사해본 결과, TGF-$\beta$3 단독으로도 IgA 합성세포수를 증가시켰으며, 이때 IL-5가 존재하였을 때 세포수가 조금 더 증가하였다. 이상의 결과는 TGF-$\beta$3가 약간의 차이는 있지만 TGF-$\beta$1과 유사하게 항체합성 패턴에 영향을 미침을 보여준다. 마지막으로, TGF-$\beta$3과 IL-5에 대한 MLN B cell 의 IgA와 IgG2b 항체합성 패턴은 spleen B cell 과 비슷하였다. 그러나 MLN B cell 의 IgG1 항체 합성은 spleen B cell과는 달리 TGF-$\beta$3에 의해 증가하였다. 본 실험의 결과는 전반적으로 TGF-$\beta$3가 TGF-$\beta$1과 비슷한 정도로 마우스 B cell의 항체합성에 영향을 미침을 보여준다. 그렇지만, 생체 내에서TGF-$\beta$3의 발현조절이 TGF-$\beta$1과 다를 것으로 예상됨으로 과연 TGF-$\beta$3가 B cell 분화에서 중요한 조절인자로 작용할지는 좀 더 연구되어야 할 것이다. TGF-$\beta$3 is among five TGF-$\beta$ isolorms and shows 80% sequence identity to TGF-$\beta$I, a prototype of TGF--$\beta$. It has been reported that TGF-$\beta$I, particularly in the presence of IL-2 or L-5, increases the pmduction of IgA and IgG2b isoiypes by LPS-actwated murine B cells. We examined the effect of TGF-P3 on Ig synlhesis by B cells from different lymphoid origins. IgA induction by TGP-$\beta$3 was mardnal in LPS-activated spleen B cell culture, while 1gA production was markedly enhanced in the culture shulated with TGF-$\beta$P3 and L-5. In addition, number of IgA secreting cells was increased by TGF-$\beta$P3. Under the same conditions, TGP-$\beta$3 alone was enough to increase IgG2b production but IgM and 1gGl. Sirmlar patiem of IgA and IgGZb enbancement by TGF-$\beta$3 and L-5 was observed in the cullures of mesenteric lymph node B cells. Thus, overall effect of TGF-$\beta$3 on Ig synthesis was quite similar to that of TGF-$\beta$I. Nonetheless, it remains to be underslood whether TGF-$\beta$3 is an important modulator in B cell differentiation since regulation of TGF-$\beta$3 expression is considered to differ from that of TGF-$\beta$I

Toll-like Receptor 1/2 Agonist Pam3CSK4 Suppresses Lipopolysaccharide-driven IgG1 Production while Enhancing IgG2a Production by B Cells

이상훈,박석래 대한면역학회 2018 Immune Network Vol.18 No.1

Interaction between pathogen-associated molecular patterns and pattern recognition receptors triggers innate and adaptive immune responses. Several studies have reported that toll-like receptors (TLRs) are involved in B cell proliferation, differentiation, and Ig class switch recombination (CSR). However, roles of TLRs in B cell activation and differentiation are not completely understood. In this study, we investigated the direct effect of stimulation of TLR1/2 agonist Pam3CSK4 on mouse B cell viability, proliferation, activation, Ig production, and Ig CSR in vitro. Treatment with 0.5 μg/ml of Pam3CSK4 only barely induced IgG1 production although it enhanced B cell viability. In addition, high-dosage Pam3CSK4 diminished IgG1 production in a dose-dependent manner, whereas the production of other Igs, cell viability, and proliferation increased. Pam3CSK4 additively increased TLR4 agonist lipopolysaccharide (LPS)-induced mouse B cell growth and activation. However, interestingly, Pam3CSK4 abrogated LPS-induced IgG1 production but enhanced LPS-induced IgG2a production. Further, Pam3CSK4 decreased LPS-induced germline γ1 transcripts (GLTγ1)/GLTε expression but increased GLTγ2a expression. On the other hand, Pam3CSK4 had no effect on LPS-induced plasma cell differentiation. Taken together, these results suggest that TLR1/2 agonist Pam3CSK4 acts as a potent mouse B cell mitogen in combination with TLR4 agonist LPS, but these 2 different TLR agonists play diverse roles in regulating the Ig CSR of each isotype, particularly IgG1/IgE and IgG2a.

TGF-β3는 마우스 IgA, IgG2b 항체의 선택적 유도작용

이은경,박석래,이세원,전계택,최의열,김평현 강원대학교 기초과학연구소 1999 기초과학연구 Vol.10 No.-

TGF-β1은 LPS로 자극시킨 마우스의 spleen B cell의 IgA와 IgG2b의 항체 합성을 선택적으로 증가시킨다고 알려져 있다. 본 연구에서는 TGF-β1과 80%의 아미노산을 공유하는 TGF-β3가 마우스 spleen B cell과 mesenteric lymph node (MLN) B cell의 항체 합성에 미치는 영향을 IL-5와 함께 조사하였다. LPS로 활성화된 spleen B cell에 TGF-β3만을 처리한 조건에서 IgA 항체합성이 약간 증가하였고, IL-5와 함께 넣어 준 배양 조건에서는 IgA 항체가 현격히 증가하였다. IgG2b 합성의 증가는 TGF-β3 자극만으로도 가능하였고 IgA와는 달리 IL-5의 첨가 효과는 관찰되지 않았다. 한편, TGF-β3는 IgM과 IgG1 항체 합성을 감소시켰고, IL-5와 함께 존재한 경우에도 의미있는 합성 증가는 볼 수 없었다. ELISPOT assay로 IgA 합성 세포수의 변화를 조사해본 결과, TGF-β3 단독으로도 IgA 합성세포수를 증가시켰으며, 이 때 IL-5가 존재하였을 때, 세포수가 조금 더 증가하였다. 이상의 결과는 TGF-β3와 IL-5에 대한 MLN B cell의 IgA와 IgG2b 항체합성 패턴은 spleen B cell과 비슷하였다. 그러나 MLN B cell의 IgG1 항체 합성은 spleen B cell과는 달리 TGF-β3에 의해 증가하였다. 본 실험의 결과는 전반적으로 TGF-β3가 TGF-β1과 비슷한 정도로 마우스 B cell의 항체합성에 영향을 미침을 보여준다. 그렇지만 생체 내에서의 TGF-β3의 발현 조절이 TGF-β1과 다를 것으로 예상됨으로 과연 TGF-β3가 B cell 분화에서 중요한 조절인자로 작용할지는 좀더 연구되어야 할 것이다. Key Words : TGF-β1, TGF-β3, IL-5, B lymphocyte, IgA, IgG2b

Anti-viral effect of ginsenoside-Rb2 from Red ginseng on Sendai virus infection in mice

유영춘,이정림,박석래,남기열,최재을 한국실험동물학회 2012 한국실험동물학회 학술발표대회 논문집 Vol.2012 No.8

IgA Isotype Switching 연구를 위한 마우스 B Lymphoma Cell (CH12F3-2A)의 특성 연구

장영생,최서현,박석래,김현아,박재봉,김평현 대한면역학회 2004 Immune Network Vol.4 No.4

Background: It is well known that IgA isotype switching is induced by TGF-β1. LPS-activated mouse normal B cells well differentiate into IgA secreting plasma cells under the influence of TGF-β1. Nevertheless, there are lots of difficulties in studying normal B cells in detail because it is not simple to obtain highly purified B cells, showing low reproducibility and transfection efficacy, moreover impossible to keep continuous culture. To overcome these obstacles, it is desperately needed to develop B cell line which acts like normal B cells. In the present study, we investigated whether CH12F3-2A lymphoma cells are appropriate for studying IgA isotype switching event. Methods: CH12F3-2A B cell line was treated with LPS and TGF-β1, then levels of germ-line (GL) transcripts were measured by RT-PCR, and GLα promoter activity was measured by luciferase assay. In addition, membrane IgA (mIgA) expression and IgA secretion were determined by FACS and ELISA, respectively. Results: TGF-β1, regardless of the presence of LPS, increased level of GLα transcripts but not GLγ2b transcripts. However, IgA secretion was increased dramatically by co-stimulation of LPS and TGF-β1. Both mIgA and IgA secretion in the presence of TGF-β1 were further increased by over-expression of Smad3/4. Finally, GLα promoter activity was increased by TGF-β1. Conclusion: CH12F3-2A cell line acts quite similarly to the normal B cells which have been previously reported regarding IgA expression. Thus, CH12F3-2A lymphoma cell line appears to be adequate for the investigation of the mechanism(s) of IgA isotype switching at the cellular and molecular levels. (Immune Network 2004;4(4):216-223)

Inhibitory Effect of the Extract of Aronia Fruits (AF-WE) on LPS-induced Inflammation in Raw 264.7 Macrophages

강은주,이정림,박석래,유영춘 한국실험동물학회 2014 한국실험동물학회 학술발표대회 논문집 Vol.2014 No.2