PROJECT SUMMARY Over the past several decades, the traditional approach to combating viral infectious diseases has been to target the virus itself, in most cases by either blocking virus-encoded enzymes that are required for viral replication, or by preventing the virus from entering host cells. One of the major caveats of these approaches has been the ability of the virus to readily mutate and thereby become resistant to these classical types of antiviral therapies. In fact, this is a serious problem for the therapy of RNA virus infections, such as HIV or influenza virus, which are known to rapidly mutate and thereby escape antiviral drugs. Additionally, traditional antiviral approaches are designed to target a specific virus, and therefore are ineffective against any new virus that may emerge in the future, and it is impossible to predict what virus will cause the next outbreak or pandemic. Therefore, there is the urgent need to develop new ways for targeting viral pathogens, which will require creative and innovative research. Like human proteins, viral proteins robustly undergo posttranslational modifications (PTMs) for their regulation and proper functioning in the virus life cycle. In most cases, viral PTMs are dynamically regulated by human enzymes, such as kinases/phosphatases, ubiquitin E3 ligases/deubiquitinating enzymes, or acetyl transferases/deacetylases. Thus, cellular enzymes play an important role in controlling the ability of the virus to replicate and to cause disease. This proposal’s overarching goal is to comprehensively map the ‘viral PTMome’ to identify the PTMs that are essential for virus replication and pathogenesis. We will combine proteomics screens and molecular virology approaches including reverse genetics techniques with cutting- edge molecular, biochemical and biophysical studies. This will allow us to identify and characterize viral PTMs and the responsible host modifying enzymes, as well as to determine their roles for effective viral replication and pathogenesis. This powerful approach, combined with collaborative studies to design and test chemical inhibitors to block the enzymes that regulate critical viral PTMs, will not only provide unique mechanistic insight into host control of virus replication but will also lay the groundwork for developing new antivirals for a range of emerging viral infectious diseases.