Interferon (IFN) regulatory factors (IRFs) are a family of transcription mediators involved in the regulation of type I IFN (IFN-alpha/beta) transcription. Among the nine already identified IRFs, IRF-3 is a constitutively and ubiquitously expressed and plays a critical role in the transcriptional activation of type I IFN and IFN-stimulated genes including IFN-alpha1, IFN-beta and RANTES. In the present study, we report the identification of a novel alternatively spliced transcript of murine Irf-3 gene (mIrf-3) designated mIRF-3a. mIRF-3a is ubiquitously present in mouse tissues along with mIRF-3 and could be translocated into the nucleus in uninfected L929 cells. EMSA and reporter assay demonstrated that mIRF-3a binds to ISRE sequences in murine Ifn-beta promoter and represses Ifn-beta promoter activation induced by Newcastle disease virus infection. These results suggest that mIRF-3a may act as a modulator of mIRF-3 functions in vivo.