Vaccinia virus K1 ankyrin repeat protein inhibits NF-κB activation by preventing RelA acetylation

The vaccinia virus (VACV) K1 protein inhibits dsRNA-dependent protein kinase (PKR) activation. A consequence of this function is that K1 inhibits PKR-induced NF-kappa B activation during VACV infection. However, transient expression of K1 also inhibits Toll-like receptor (TLR)-induced NF-kappa B activation. This suggests that K1 has a second NF-kappa B inhibitory mechanism that is PKR-independent. This possibility was explored by expressing K1 independently of infection and stimulating NF-kappa B under conditions that minimized or excluded PKR activation. K1 inhibited both TNF- and phorbol 12-myristate 13-acetate (PMA)-induced NF-kB activation, as detected by transcription of synthetic (e.g. luciferase) and natural (e.g. CXCL8) genes controlled by NF-kappa B. K1 also inhibited NF-kappa B activity in PKRkd cells, cells that have greatly decreased amounts of PKR. K1 no longer prevented I kappa B alpha degradation or NF-kappa B nuclear translocation in the absence of PKR, suggesting that K1 acted on a nuclear event. Indeed, K1 was present in the nucleus and cytoplasm of stimulated and unstimulated cells. K1 inhibited acetylation of the RelA (p65) subunit of NF-kappa B, a nuclear event known to be required for NF-kappa B activation. Moreover, p65-CBP (CREB-binding protein) interactions were blocked in the presence of K1. However, K1 did not preclude NF-kappa B binding to oligonucleotides containing kappa B-binding sites. The current interpretation of these data is that NF-kappa B-promoter interactions still occur in the presence of K1, but NF-kappa B cannot properly trigger transcriptional activation because K1 antagonizes acetylation of RelA. Thus, in comparison to all known VACV NF-kappa B inhibitory proteins, K1 acts at one of the most downstream events of NF-kappa B activation.