The U3-LTR region of leukemia viruses transactivates cancer-related signaling pathways through the production of a non-coding RNA transcript although the role of this transcript in virus infection remains unknown. In this study we demonstrate for the first time that an long terminal repeat (LTR)-specific small non-coding RNA is produced from a feline leukemia virus (FeLV)-infected feline cell line. RNA cloning identified this as a 104 base transcript that originates from the U3-LTR region. We also demonstrate that in in vitro assays this LTR-RNA transcript activates NF kappaB signaling. Taken together, our findings suggest a possible role for this LTR transcript in FeLV pathogenesis.