Cullin-RING E3 ubiquitin ligases (CRLs) are a major family of ubiquitin ligases. CRLs are formed by a scaffold protein (Cullin) that forms a multimeric complex with an E2 ubiquitin conjugating enzyme on one end and a substrate receptor protein recruiting target substrate on the other end. F-box proteins are the adaptor proteins for CRL1 ligases which recognizes a large repertoire of proteins for ubiquitination, resulting in their proteasomal degradation or a modification that alters their function. The transcription factor NF-κB plays a crucial role in the host response to infection and is targeted by many viruses including vaccinia virus (VACV). NF-κB is highly regulated by ubiquitination and the ubiquitin-proteasome system, with the paradigm being the degradation of the inhibitor of κB (IκB) by the F-box protein β-TrCP. A previous screen performed in the laboratory identified FBXO22 as a putative positive regulator of NF-κB activation. FBXO22 is an F-box protein known to coordinate cytoskeleton framework, senescence, hormone signalling, and metastasis regulating proteins. However, only a few physiological substrates of FBXO22 have been characterized till now and the role of FBXO22 in innate immunity and infection is uncharacterized. This project aims to determine the role of FBXO22 in antiviral immunity and how this protein is antagonised by orthopoxviruses. My work demonstrates that depletion of FBXO22 in A549 cells, either by shRNA knock-down or in gRNA knock-out cells, results in a significant reduction of NF-κB activation induced by cytokines or virus infection, measured by a consensus κB-luciferase reporter system or qPCR of endogenous cytokines. Conversely, overexpression of FBXO22 resulted in enhanced NF-κB activation. FBXO22 activity was mapped at, or downstream of, NF-κB translocation to the nucleus. My results also show that FBXO22 is being degraded by VACV in a proteasome- and neddylation-dependent manner. By constructing various mutants of FBXO22, I show that the F-box domain is required for VACV to target FBXO22. Furthermore, I confirmed that whilst cowpox virus, ectromelia virus and the highly attenuated strain of VACV, MVA, degrade FBXO22, another attenuated strain termed vv811 as well as, surprisingly, monkeypox virus, did not. This comparative analysis allowed the identification of the viral protein responsible for FBXO22 degradation, which was the historical 68k-ANK also known as B18 or OPG203. In conclusion, here I have discovered that FBXO22 is a positive regulator of innate responses, restricts orthopoxviruses and is antagonized by protein B18. Further work will aim to understand the exact mechanism of action of FBXO22 in innate immunity and the diverse reasons this multifunctional protein is targeted by orthopoxviruses. The outcomes of this project provide novel insights onto cell responses to stress and infection and viral hijacking of CRLs, some of which may have potential as therapeutic strategies.