RNA interference (RNAi) is a process of double-stranded RNA-dependent post-transcriptional gene silencing. It has become one of the most powerful and widely used strategies for genetic analysis based on the highly specific and efficient silencing of target genes. RNAi library screens have gained much attention from virologists resulting in the publication of several whole-genome short interfering RNA (siRNA) screens that identified numerous new host genes impacting RNA-virus replication. Although extremely powerful, one major shortcoming of current siRNA screens is the inherent risk of off-target effects, leading to high false positive rates. This proposal is geared towards the development of ‘next-generation’ lentiviral shRNA library screens that solve many problems existing in current RNAi screens and to exploit these libraries to identify host-factors critical for the infection cycle of human herpesviruses and for virus-induced ER-associated degradation (ERAD). Besides development of powerful whole genome shRNA library screens, these efforts will lead to new insights into the biology of human herpesvirus infections and boost research to explore the underlying cellular biology of these processes. Additionally, the identified virus-host interactions could be exploited to develop new antiviral therapies by specifically targeting host genes critically involved in virus infections.