The human T-cell leukemia virus -2 (HTLV-2) antisense protein APH-2 interacts with p65 and inhibits NF-ĸB Tax-2 activation.

Human T-cell leukemia virus types 1 and 2 (HTLV-1 and 2) are genetically related retroviruses that cause persistent infections in vivo. HTLV-1 is the causative agent of adult T-cell leukemia (ATL) and myelopathy/tropical spastic paraparesis, while HTLV-2 is not associated with ATL-like disorders. HTLV genome codes for a transactivator protein, Tax, which is crucial for enhancing viral expression and driving cell transformation. The promoter located in the 3’ LTR in both HTLV-1 and 2 drives the transcription of antisense RNAs coding for regulatory proteins named HBZ (HTLV-1 b-Zip factor) for HTLV-1 and APH-2 (antisense protein of HTLV-2) for HTLV-2. Both proteins contain basic residues required for nuclear localization, but differ in their N- and C- terminus. They negatively regulate HTLV transcription, inhibiting Tax-dependent LTR activation, although not in a similar extent. HBZ does not bind Tax-1, whereas APH-2 binds to Tax-2. We therefore aim to investigate the presence of APH-2 in complexes that recruit Tax-2 and the effect of APH-2 expression on NF-ĸB activation. We found that APH-2 interacts with p65, as well as with Tax-2 and Tax-1, indicating that protein structural differences between APH-2 and HBZ are responsible for Tax proteins interaction. We performed promoter activated luciferase assays showing that when co-expressed with p65 or Tax-2, increased amounts of APH-2 inhibited the mediated p65 and Tax-2 NF-ĸB activation. Immunofluorescence analyses indicated that APH-2, unlike HBZ, localized in both nucleus and cytoplasm. Altogether, our results demonstrate that APH-2 shares with HBZ the affinity with p65 and the inhibition of p65 induced NF-ĸB activation and highlight the difference between APH-2 and HBZ in the selectivity for the interaction that may explain their diversity in cellular localization. The different role of these proteins in altering cellular pathways will be further investigated. This study is funded by AIRC-Cariverona.