Canine parvovirus (CPV) is capable of inducing hemorrhagic gastroenteritis and myocarditis in dogs, which imposes financial burdens on dog breeding and pet industries. To elucidate the complex interactions between the CPV non-structural protein NS1 and host proteins during CPV infection, this study employed co-immunoprecipitation (Co-IP) approach coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) to screen 401 host proteins interacting with NS1 in F81 cells. Subsequent protein-protein interaction (PPI) network construction, Gene Ontology (GO) functional categorization, and Kyoto Encyclopedia of Genes and Genomes (KEGG) mapping analysis revealed that the NS1-interacting proteins are involved in metabolism, gene expression, and the cell cycle. Based on these findings, Co-IP and GST pulldown assays were used to verify the interactions of seven proteins (PCNA, FHL2, SEH1L, DHRS7B, RFC4, POLB, and FRMD6) that specifically bind to CPV NS1. Further investigation of the interaction with PCNA revealed that overexpression of PCNA reduced CPV replication, whereas knockdown or inhibition of PCNA increased CPV replication. Additionally, knockdown of PCNA during CPV infection resulted in an increased proportion of cells in the S phase, thereby modulating the cell cycle. These results establish a basis for comprehending the molecular processes involved in CPV-host interactions, and they could assist in pinpointing possible antiviral targets.

To Cite This Article: Zhou H, Su X, Li Q, Cui K, Xu F, Xiao B, Cheng H, Zhang J, Qi Q, Li S and Yang B, 2026. Identification of canine parvovirus NS1-interacting host proteins and functional characterization of PCNA in regulating viral replication. Pak Vet J, 46(8): 2074-2083. http://dx.doi.org/10.29261/pakvetj/2026.181