Human adult T cell leukaemia-derived factor (ADF) was first described as an IL-2 receptor α chain (IL-2R α, Tac) upregulating factor produced by cell lines from patients with adult T-cell leukemia. 1 The purification and cloning of human ADF showed that it represents a mammalian homologue of bacterial thiore- doxin (TRX). ADF/human TRX has an active thiol- group (-Cys-Gly-Pro-Cys-) and acts as an endogenous thiol-related reducing protein. 2,3 TRX functions as hydrogen donor mediating the conversion of ribo- nucleotides to deoxyribonucleotides, the degradation of insulin, and the stabilization of glucocorticoid re- ceptors. Furthermore, thioredoxin is involved in an electron-transfer system common to a variety of cells and scavanges free radicals. 4 Despite its intracellular function as disulfide reduc- ing enzyme and its lack of a signal sequence the protein has been found to exert several effects extracellularily. 5 It has been shown that TRX/ADF is actively secreted by a variety of normal and transformed human cells, including fibroblasts, airway epithelial cells, and acti- vated B and T cells. The secretory mechanism for ADF shares several features with but is not identical to the alternative pathway described for IL-1β. 6 For human ADF it has been demonstrated that sev- eral cytokine-like functions are related to this protein. For example, an autocrine growth factor produced by the EBV-transformed B-cell line 3B6 7 named 3B6-IL-1, and MP6-BCGF, a B-cell growth factor de- rived from the T-cell hybridoma MP6, 8 were shown to be identical to ADF. A factor enhancing cytotoxicity of eosinophils (eosinophil cytotoxicity-enhancing factor, ECEF), which is produced by activated U937 cells, is closely related or identical to ADF. 9 In line with this, purified recombinant ADF enhanced cytotoxicity and migration of human eosinophils induced by C5a. 10 In addition, recombinant ADF has been described to inhibit the cytotoxicity mediated by TNF and anti-Fas antibodies on U937 cells. 11 In contrast to the large array of studies analysing effects of human ADF, very little is known about the expression and regulating acivities of the murine homo- logue. We therefore expressed this molecule in E. coli, established a purification protocol and analysed the effect of murine ADF on the proliferation of murine T cells. 6 CYTOKINE, Vol. 8, No. 1 (January), 1996: pp 6–13 EXPRESSION AND CO-CYTOKINE FUNCTION OF MURINE THIOREDOXIN/ADULT T CELL LEUKAEMIA-DERIVED FACTOR (ADF) Horst Blum, Martin Röllinghoff, André Gessner Human ADF (adult T cell leukaemia-derived factor), an isoform of thioredoxin, promotes proliferation of certain human lymphoid cell lines and is involved in many thiol-dependent reducing reactions. To study functional aspects of the murine homologue, we established inducible overexpression of murine ADF in E. coli and a purification method which led to an apparently homogeneous 14 kDa protein. This recombinant ADF was tested in prolifer- ation assays with murine Th2 cells (D 10.G4.1) and CTLL-2 cells. In synergy with IL-2, IL-4, IL-7 and IL-9 A D F displayed co-cytokine activity. These proliferative effects were neutral- ized by an affinity-purified polyclonal rabbit anti-A D F antiserum. The effects of A D F were critically dependent on the presence of 2-mercaptoethanol. Bacterial thioredoxin had simi- lar effects on the proliferation of murine T cells. Thus, the thiol-related reducing capacity of these proteins is essential for their growth promoting activity. A s investigated at the levels of mRNA and protein in several murine cell clones and lines as well as in mouse tissues A D F is expressed ubiquitously. Finally it could be demonstrated by competitive PCR that in contrast to cytokine mRNAs (e.g. IL-4 and IL-13) the expression of ADF mRNA in murine Th2 clones and spleen cells is not influenced by stimulation of these cells through the T cell recep- tor complex. Murine ADF therefore represents a protein constitutively expressed in a wide variety of cells with the capacity to enhance the proliferative effect of several cytokines on murine T cells. © 1996 A cademic Press Limited From the Institut für Klinische Mikrobiologie und Immunologie der Universität Erlangen-Nürnberg, Wasserturmstr. 3, D-91054 Erlangen, Germany Correspondence to: André Gessner Received 3 April 1995; accepted for publication 18 August 1995 © 1996 Academic Press Limited 1043-4666/96/010006+8 $12.00/0 KEY WORDS: Thioredoxin/Adult T cell leukemia-derived factor/co-cytokine function/IL-2/IL-4/IL-7/IL-9