Interferon regulatory factor (IRF) family members have been implicated as critical transcription factors that function in immune response, hematopoietic differentiation and cell growth regulation. Activation of IRF-5 results in the production of pro-inflammatory cytokines such as TNFalpha, IL6 and IL12p40, as well as type I interferons.In this study, we identify a G202C (position relative to translation start codon) missense-mutation transcript of IRF-5 in transformed B and T cell lines, which were either infected or non-infected by viruses, and peripheral blood from ATL or CLL patients. The mutated transcript encodes a novel protein in which the sixty-eighth amino acid, Alanine, is substituted by Proline (IRF-5P68) in the DNA binding domain of IRF-5. IRF-5P68 phenotype results in a complete loss of its DNA-binding activity and functions as a dominant negative molecule through interacting with wild type IRF-5. Co-expression of IRF-5P68 inhibits MyD88-mediated IRF-5 transactivation. Moreover, Toll-like receptor (TLR)-dependent IL6 and IL12P40 production induced by lipopolysaccharide (LPS), R837 or CpG ODN 1826 was reduced in IRF-5 (P68) expressing cells as compared to the control cells.IRF-5P68 acts as a dominant negative regulator that interferes with IRF-5-mediated production of pro-inflammatory cytokines. The functional characterization of the novel IRF-5 mutant in transformed B and T cell lines and in ATL and CLL patients may lead to a better understanding of the role of these transcriptional regulators in hematopoietic malignancies.