The nucleocapsid protein is significant in the formation of viral RNA of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), accounting for the largest proportion of viral structural proteins. Here, we report for the first time that the 11S proteasomal activator PA28 gamma regulates the intracellular abundance of the SARS-CoV-2 N protein (nCoV N). Furthermore, we have identified proteasome activator PA28 gamma as a nCoV N binding protein by co-immunoprecipitation assay. As a result of their interaction, nCoV N could be degraded by PA28 gamma-20S in vitro degradation assay. This was also demonstrated by blocking de novo protein synthesis with cycloheximide. The stability of nCoV N in PA28 gamma-knockout cells was greater than in PA28 gamma-wildtype cells. Notably, immunofluorescence staining revealed that knockout of the PA28 gamma gene in cells led to the transport of nCoV N from the nucleus to the cytoplasm. Overexpression of PA28 gamma enhanced proteolysis of nCoV N compared to that in PA28 gamma-N151Y cells containing a dominant-negative PA28y mutation, which reduced this process. These results suggest that PA28 gamma binding is important in regulating 20S proteasome activity, which in turn regulates levels of the critical nCoV N nucleocapsid protein of SARS-CoV-2, furthering our understanding of the pathogenesis of COVID-19. (C) 2020 Elsevier Inc. All rights reserved.