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Hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs) is a key component of the endosomal sorting complexes required for transport and has been demonstrated to play a regulatory role in endocytosis/exocytosis and the accumulation of inteal vesicles in multivesicular bodies.Citron kinase is a Ser/The kinase that we previously reported to enhance human immunodeficiency virus type 1 (HIV-1) virion production.However,the relationship between Hrs and citron kinase in HIV-1 production remains elusive.Here,we report that Hrs interacts with citron kinase via its FYVE domain.Overexpression of Hrs or the FYVE domain resulted in a significant decrease in HIV-1 virion production.Depletion of Hrs by RNA interference in HEK293T cells increased HIV-1 virion production and enhanced the activity of citron kinase.These data suggest that Hrs inhibits HIV-1 production by inhibiting citron kinase-mediated exocytosis.