Foxp3+ Tregs express 4 class IIa Hdac enzymes (Hdac4, 5, 7 & 9) with little or no catalytic activity and yet when deleted, at least with Hdac7 and Hdac9, enhance Foxp3 expression and acetylation, and promote Treg production and suppressive function. In seeking how this might occur, we have examined the functions of the DNA binding Mef2 family of transcription factors, since these are well established partners of class IIa Hdacs.

[bull] Of the 4 Mef2 isoforms (Mef2a, b, c & d), Mef2c and Mef2d are highly expressed in Tregs compared to Mef2a or Mef2b (~10 fold higher levels). [bull] Conditional deletion of individual genes in Foxp3+ Treg cells led to partial but not complete overlap between the genes affected (microarray), and qPCR studies confirmed that Mef2c deletion led to decreased IL-10 and granzyme B (GrB) expression, whereas Mef2d deletion led to decreased Foxp3, Ctla4, IL-10, GrB and CD25 but increased IL-17 expression. [bull] ChIP assays showed that both Mef2c and Mef2d bound to the Foxp3 promoter, and the binding of Mef2d was increased >10-fold by p300. [bull] Western blotting confirmed that Mef2d was hyperacetylated and stabilized by p300. [bull] Luciferase assays showed that the basal activity of individual Mef2 isoforms at the Foxp3 promoter was low but was markedly increased by the presence of p65. Moreover, the activity of Mef2 isoforms at the Foxp3 promoter, even in the presence of p65, was impaired by Hdac7, Hdac9 or MITR (Hdac9 isoform lacking the catalytic domain), and the combination of Hdac7 and Hdac9 was especially suppressive. [bull] Deletion of Mef2c or Mef2d with in Tregs resulted in impairment of Treg suppressive function in vitro. [bull] Mice with Treg deletion of Mef2 or Mef2d showed increased activation of Tcon cells (CD44^hiCD62L^lo) within peripheral lymphoid tissues, and in contrast to their wild-type counterparts, developed rapid and acute rejection of BALB/c cardiac allografts despite rapamycin therapy (p<0.01).

There are no known data as to the presence and functions of Mef2 genes in Treg cells. Our studies highlight a key role of Mef2c and Mef2d in promoting Foxp3 expression and Treg function, and provide key insights into how and why targeting of Hdac7 and Hdac9 promotes Treg function. These data have recently led to identification of small molecules that disrupt the interactions of class IIa Hdac enzymes with Mef2 proteins. We have found that such drugs enhance murine and human Foxp3+ Treg function in vitro, and we are currently evaluating these compounds in vivo.

CITATION INFORMATION: Samanta A., Wang L., Han R., Hancock W. Central Role of Mef2 Transcription Factors in Regulation of Foxp3 Expression and Treg Function Am J Transplant. 2017;17 (suppl 3).

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