Human IL4I1 is a secreted l-phenylalanine oxidase expressed by mature dendritic cells that inhibits T-lymphocyte proliferation

Abstract

Interleukin-4-induced gene 1 (IL4I1) was first described as a B-cell IL4-inducible gene and is highly expressed in primary mediastinal B-cell lymphomas. We established stable HEK293 clones expressing human and mouse IL4I1 to examine their biochemical properties and function. Both proteins were secreted into the culture medium, and we observed the secretion of endogenous human IL4I1 (hIL4I1) protein in a mediastinal lymphoma B-cell line, MedB-1. We showed that IL4I1 has L-amino acid oxidase activity, optimal at physiological pH and primarily directed toward phenylalanine. Immunohistochemical analysis of secondary lymphoid organs showed staining of germinal center macrophages and inflammatory myeloid cells. In vitro, functional enzyme was highest in mature dendritic cells (DCs), suggesting a role in antigenpresenting cell/T-lymphocyte cross-talk. Indeed, hIL4I1 inhibited the proliferation of CD3-stimulated T lymphocytes with a similar effect on CD4 ؉ and CD8 ؉ T cells. IntroductionInterleukin-4 (IL4) is the master cytokine for T-helper type 2 (Th2) lymphocyte differentiation and function. Along with its important role in B-lymphocyte stimulation by T-helper cells in germinal centers of secondary lymphoid organs, it also regulates the growth and function of many different cell types of the immune system. 1 IL4 transcriptional effects are primarily mediated by the signalization and transcription factor 6 (STAT6), which is responsible for the induction and repression of multiple target genes. 2 An immediate-early IL4-inducible gene called interleukin fourinduced gene 1 (FIG1/IL4I1) has first been described in the mouse 3 and subsequently characterized in human B cells. 4 IL4I1 mRNA expression is restricted to lymphoid tissues, with the highest levels found in lymph nodes and spleen. 4,5 IL4-activated murine B cells express IL4I1 within 2 hours of stimulation. We have demonstrated that high levels of expression of this gene are characteristic of primary mediastinal large B-cell lymphoma (PMBL), a specific subtype of diffuse large B-cell lymphoma. 5 The high expression of IL4I1 by PMBL may result from the constitutive activation of STAT6 in these tumors. 6 Both human and mouse IL4I1 mRNA sequences encode a protein containing a putative signal peptide indicative of potential secretion and a large central domain, highly homologous to flavin-containing amino acid oxidases. 3,4 While sharing high similarity over the majority of the sequence (547 amino acids of the 567 in the human sequence), the human and mouse proteins diverge substantially at their C-terminal region. 4 The IL4I1 protein shares the highest similarity with proteins presenting L-amino acid oxidase (LAAO) activity, such as snake venom LAAO,7,8 and mouse milk LAAO. 9 Recently, Mason et al have demonstrated that mouse IL4I1 (mIL4I1) possesses an LAAO activity toward aromatic amino acids. The enzyme was mostly active at acidic pH, in contrast to other known members of the LAAO family, and its expression was restricted primarily to lysosomes. 10 In this work, w...

Rarity of Human T Helper 17 Cells Is due to Retinoic Acid Orphan Receptor-Dependent Mechanisms that Limit Their Expansion

“…IL4I1 encodes a secreted L-amino-acid oxidase that inhibits human T lymphocyte proliferation in vitro by inducing a temporary decrease in CD3z chain expression resulting from the enzymatic production of H 2 O 2 (Boulland et al, 2007). We found that IL4I1 mRNA expression was virtually undetectable on either resting or activated Th1 cell clones, but it was highly expressed on resting Th17 cell clones and further increased by anti-CD3-CD28 stimulation ( Figure 4A).…”

“…Whereas the reduced IL-2 responsiveness was explained by the lack of PI3K-AKT-mTORC1 axis activation, the defective IL-2 production by Th17 cells appeared to be related to their reduced c-Fos, c-Jun, and nuclear factor of activated T cells (NF-AT) activity. The reduced activity of these transcription factors associated with high expression of the IL-4-induced gene 1 (IL4I1) mRNA, which encodes for a L-phenylalanine oxidase that has been shown to downregulate CD3z expression in T cells (Boulland et al, 2007). IL4I1 mRNA expression was strictly RORC dependent and was already detectable in Th17 cell precursors from UBC and newborn thymus.…”

The novel immunosuppressive enzyme IL4I1 is expressed by neoplastic cells of several B-cell lymphomas and by tumor-associated macrophages

“…Moreover, we have observed high IL4I1 expression in antigen-presenting cells of myeloid origin (macrophages and dendritic cells) from normal and inflammatory lymphoid tissue. 2 These observations highlight similarities between IL4I1 and other amino-acid-catabolizing enzymes, such as indoleamine-2,3-dioxygenase (IDO), arginase-1 and inducible nitric oxide synthase, all associated with the inhibition of T-cell responses. The expression of these enzymes has been demonstrated in several tumor types, both in the malignant cell and/or the microenvironment.…”

Human spermatozoa possess an IL4I1 l-amino acid oxidase with a potential role in sperm function

“…A survey of potential substrates for this enzyme demonstrated that it was optimally active following exposure to the aromatic amino acids phenylalanine and tryptophan (Fig. 2) in keeping with the known activity of this enzyme in the immune system (Boulland et al 2007). The localization of this enzyme in the sperm acrosome is also consistent with the observation that this enzyme is maximally active at acidic pH and is preferentially targeted to lysosomes (Mason et al 2004); the acrosomal vesicle is known to exhibit a low intraacrosomal pH and shares many features with cellular lysosomes (Working & Meizel 1983, Moreno & Alvarado 2006.…”