Karsten Kretschmer

Cellular and molecular pathways of antigen-specific immunosuppression

Previous and current research

CD4+CD25+ regulatory T cells (Treg) expressing the transcription factor Foxp3 play an essential role in establishing dominant self-tolerance, controlling inflammatory responses and maintaining immune homeostasis in mice and men. In recent years, Treg have attracted considerable attention as promising gain-of-function targets in clinical settings of unwanted immune responses, such as organ-specific autoimmunity and immune rejection of transplanted hematopoietic stem cells. In this context, our research interests focus on molecular and cellular pathways of Foxp3+ Treg generation and suppressor function.

Promoting antigen-specific Treg in autoimmunity. Previous studies have indicated that selective delivery of non-self antigen to DEC-205+ dendritic cells (DCs) via recombinant anti-DEC-205 antibodies can extrathymically induce Foxp3+ Treg from initially naïve CD4+CD25–Foxp3– T cell receptor (TCR) transgenic T cells (Kretschmer et al., Nat Immunol. 2005; Nat Protoc. 2006). Global gene expression analysis of Foxp3+ Treg, purified from different anatomical locations or artificially generated by different means, revealed that Foxp3+ Treg generated in vivo by DEC-205+ DC targeting are unique in that they exhibited a distinct mRNA Treg signature, including many mRNAs encoding Treg effector molecules (Feuerer et al., PNAS 2010). In contrast to TGF-β-mediated in vitro generation of Foxp3+ cells, DEC-205+ DC-targeted Treg conversion in vivo resulted in efficient demethylation of conserved CpG motifs within the non-coding part of the Foxp3 gene and long-term stability of induced Foxp3 expression (Polansky*, Kretschmer* et al., Eur. J. Immunol. 2008). Furthermore, we have evaluated the concept of extrathymic Treg de novo generation in vivo for self-antigens and self-reactive CD4+ T cells in the NOD mouse model for type 1 diabetes. Proof-of-principle experiments indicated that anti-DEC-205-mediated targeting efficiently converted pancreatic beta-cell-reactive CD4+ T cells into long-lived Foxp3+ Treg and reduced the incidence of diabetes (Petzold*, Riewaldt* et. al., Rev Diabet Stud. 2010). Furthermore, DEC-205+ DC targeting ameliorated clinical symptoms in the PLP(139-151)-induced SJL model of experimental autoimmune encephalomyelitis by both recessive and dominant tolerance mechanisms (Stern et al., Proc Natl Acad Sci U S A. 2010).

Extrathymic Treg development in the steady state. Analysis of early events during DEC-205+ DC-targeted Treg generation from TCR transgenic CD4+ T cells allowed us to delineate extrathymic differentiation stages to Foxp3+ Treg with distinct surface markers. Correlating these findings with polyclonal non-TCR-transgenic T cells helped identifying a population of CD4+Foxp3– T cells in peripheral lymphoid organs of nonmanipulated mice that is precommitted to differentiate into stable Foxp3+ Treg (Schallenberg et al., J Exp Med. 2010). This study provided evidence that, under physiological conditions, extrathymic Treg generation contributes to the overall peripheral Treg pool in the steady state.

Treg in hematopoietic homeostasis. We have recently established a mouse model, in which recombination-activating gene 1 (Rag1) expression and thereby T and B cell development is prevented by targeted inversion of exon 2 of the Rag1 gene flanked by opposing loxP sites. In such Indu-Rag1fl/fl mice, B and T lymphopoiesis is initially prevented but can be induced by activatable Cre recombinase to restore a functional Rag1 transcription unit (Düber et al., Blood 2009). Currently, we employ this novel mouse model with inducible lymphopoiesis to study different aspects of development, homeostasis and function of T and B cells in the context of immune tolerance, autoimmunity and hematopoietic stem cell transplantation.

Foxp3-dependent Treg lineage specification. Previous studies combining genome-wide location (ChIP-on-Chip) and global mRNA expression analysis have identified a core set of approximately 1.100 promoters of protein-coding genes that are occupied by Foxp3 (Marson*, Kretschmer* et al., Nature 2007). Our ongoing studies, employing transcriptome-wide microRNA quantification, bioinformatics prediction of transcription factor DNA binding and ChIP focus on mechanisms of transcriptional and translational regulation that govern the generation and function of Foxp3+ Treg.

Future prospects and goals

We will continue studying molecular and cellular aspects of Foxp3+ Treg physiology, including their role in establishing dominant self-tolerance, controlling inflammatory responses and maintaining immune homeostasis. Specifically, we will continue studying

• Molecular and cellular pathways governing the generation and function of Treg and Th17 cells

• Mechanisms of immune regulation by Foxp3- and Foxp3+ Treg cells

• Molecular networks in immunity, tolerance and autoimmunity (including miRNAs)

• Antigen-specific Treg as gain-of-function targets in autoimmune diseases (autoimmune diabetes / pancreatic beta-cell replacement, multiple sclerosis and rheumatoid arthritis)

• Role of Treg in homeostatic hematopoiesis and hematopoietic stem cell transplantation

• Neuro-immune crosstalk in health and disease

About

Selected publications

Stern JN‡*, Keskin DB, Kato Z, Waldner H, Schallenberg S, Anderson A, von Boehmer H, Kretschmer K‡*, Strominger JL‡. (2010): Promoting tolerance to proteolipid protein-induced experimental autoimmune encephalomyelitis through targeting dendritic cells. Proc Natl Acad Sci U S A. 107(40):17280-5. ‡Corresponding and *equally contributing first authors.

Petzold C*, Riewaldt J*, Koenig T, Schallenberg S, Kretschmer K. Dendritic (2010): cell-targeted pancreatic beta-cell antigen leads to conversion of self-reactive CD4(+) T cells into regulatory T cells and promotes immunotolerance in NOD mice. Rev Diabet Stud. 7(1):47-61. *These authors contributed equally.

Schallenberg S, Tsai PY, Riewaldt J, Kretschmer K. (2010): Identification of an immediate Foxp3(-) precursor to Foxp3(+) regulatory T cells in peripheral lymphoid organs of nonmanipulated mice. J Exp Med. 207(7):1393-407.

Düber S, Hafner M, Krey M, Lienenklaus S, Roy B, Hobeika E, Reth M, Buch T, Waisman A, Kretschmer K*, Weiss S* (2009):  Induction of B-cell development in adult mice reveals the ability of bone marrow to produce B-1a cells. Blood. 14(24):4960-7. *Shared senior authorship.

Marson A*, Kretschmer K*, Frampton GM, Jacobsen ES, Polansky JK, MacIsaac KD, Levine SS, Fraenkel E, von Boehmer H, Young RA (2007): Foxp3 occupancy and regulation of key target genes during T-cell stimulation. Nature. 45(7130):931-5. *These authors contributed equally.

Kretschmer K, Heng TS, von Boehmer H.(2006): De novo production of antigen-specific suppressor cells in vivo. Nat Protoc. 1(2):653-61.

Kretschmer K, Apostolou I, Hawiger D, Khazaie K, Nussenzweig MC, von Boehmer H (2005): Inducing and expanding regulatory T cell populations by foreign antigen. Nat Immunol. 6(12):1219-27.

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