Short communication Molecular characterisation of porcine Forkhead-box p3 (Foxp3) Kerstin Bolzer 1 , Tobias Ka ¨ser 1 , Armin Saalmu ¨ ller, Sabine E. Hammer * Institute of Immunology, Department of Pathobiology, University of Veterinary Medicine, Veterina ¨rplatz 1, 1210 Vienna, Austria 1. Introduction The transcription factor forkhead-box p3 (Foxp3), a 49– 55 kDa protein, is a member of the P subfamily of transcriptional regulators that are characterised by the presence of a highly conserved winged-helix/forkhead DNA binding domain (DBD) together with other functional domains, including a C 2 H 2 zinc finger and a coiled/coiled leucine zipper (Lopes et al., 2006). Mutation of Foxp3 in ‘scurfy’ mice and Foxp3 in humans with IPEX (Immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome) results in fatal, early onset autoimmune disease and demonstrates the critical role of Foxp3 in maintaining immune homeostasis (Chatila, 2005). The central role of the Fox family transcription factors in embryonic pattern- ing, development, and metabolism has been replenished by their decisive mode of operation in thymic development (Foxn1), lineage commitment (Foxp3), and function of lymphocytes (Foxj1, Foxo3, and Foxd1) (Carlsson and Mahlapuu, 2002; Coffer and Burgering, 2004; Lin and Peng, 2006; Marson et al., 2007). Foxp3 has been shown to be both necessary and sufficient for the development and function of naturally occurring CD4 + CD25 + regulatory T cells in mice (Ziegler, 2006). Constitutive high expression of Foxp3 mRNA has been shown in CD4 + CD25 + regulatory T cells (Tregs), and ectopic expression of Foxp3 in CD4 + CD25 À cells imparts a Treg phenotype in these cells (Hori et al., 2003; Khattri et al., 2003; Fontenot et al., 2005). The number and performance of Foxp3 + CD4 + CD25 + Tregs are essential for the prevention of autoimmunity (Saka- guchi et al., 1995; Baecher-Allan and Hafler, 2006). They have an attenuated cytokine response to T-cell receptor Veterinary Immunology and Immunopathology 132 (2009) 275–281 ARTICLE INFO Article history: Received 20 February 2009 Received in revised form 21 May 2009 Accepted 27 May 2009 Keywords: Foxp3 Porcine Sus scrofa Swine Regulatory T cells ABSTRACT In swine the phenotypical identification of regulatory T cells (Tregs) was limited so far to the surface expression of CD4 and CD25. However, with the discovery of the Treg-specific transcription factor forkhead-box p3 (Foxp3) in mice and humans a powerful marker for the identification of Tregs is available. Recently, we published data on a murine anti- mouse/rat Foxp3 antibody (FJK-16s) showing cross-reactivity with the putative porcine Foxp3 protein in lymphoid cells but the final proof for the specific cross-reactivity of this antibody was missing. By performing RACE-experiments, we have sequenced the entire porcine Foxp3 cDNA which is 1296 nucleotides in length and codes for a polypeptide of 432 amino acids. The porcine Foxp3 nucleotide and amino acid sequences show high homology to all known orthologues from other mammals, with the greatest homology with the bovine sequence. To demonstrate the specificity of the FJK-16s antibody for the porcine Foxp3 protein, HEK293T cells were transfected with porcine Foxp3 containing the FJK-16s- specific binding region and the expression of the epitope was identified by immuno- staining. In conclusion, this study represents the final proof for the specificity of the murine FJK-16s antibody for the porcine Foxp3 homologue and therefore strengthens future work on porcine Tregs. ß 2009 Elsevier B.V. All rights reserved. * Corresponding author. Tel.: +43 1 25077 2752; fax: +43 1 25077 2791. E-mail address: sabine.hammer@phylo-dat.net (S.E. Hammer). 1 These authors contributed equally to the work and should be regarded as joint first authors. Contents lists available at ScienceDirect Veterinary Immunology and Immunopathology journal homepage: www.elsevier.com/locate/vetimm 0165-2427/$ – see front matter ß 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.vetimm.2009.05.014