332회 Roles of Ets and Smad proteins in regulating induction of IgA iso…

332회

연사 : 김 평 현 , 강원대학교 생명과학부

제목:  Roles of Ets and Smad proteins in regulating induction of IgA isotype switching by TGF-β1

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), C(H) region genes. 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 GLα 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 study we demonstrate that Ets site in the mouse GLα promoter bind the transcription factor Elf-1. In addition, Binding of Elf-1 to the GLα promoter was inducible by lipopolysaccharide in nuclear extracts from splenic B cells. However, Overexpressed Elf-1 did not increase the TGF-β1-inducible GLα promoter activity as anticipated. Nonetheless, the dominant negative Ets2 expression plasmid inhibited the promoter activity in a dose-responsive manner, suggesting that endogenous Elf-1 is important in GLα transcription. We next examined the involvement of Smad proteins in TGF-β1-induced IgA expression. First, we found that TGF-β1 significantly increases endogenous germline (GL) α transcripts by LPS-stimulated CH12.LX.4933 (μ+) B lymphoma cells. To investigate its signaling mechanisms, the lymphoma cell line was transfected with pFL3 that contains the TGF-β-responsive element of the GLα promoter, and stimulated with TGF-β1. Similar to endogenous GLα transcripts, TGF-β1 induces GLα promoter activity and overexpression of Smad3 markedly enhances the promoter activity. This activity is further augmented by cotransfected Smad4. On the other hand, Smad7 substantially abrogates the synergistic effect of Smad3/4 on GLα promoter activity. In addition, overexpression of Smad3/4 enhances TGF-β1-induced endogenous GLα transcripts in normal spleen B cells. Finally, in the presence of TGF-β1, overexpression of Smad3/4 selectively increases both surface IgA expression and IgA production. The results from the present study indicate that Smad3, Smad4, and Smad7, at least in part, serve as mediators linking TGF-β1 to transcriptional regulation of IgA switching related gene and regulation of IgA class switching.