The protein-protein interactions between viral proteins and host factors have crucial roles in plant virus infection and host defense. The potyviral nuclear inclusion protein a protease (Nia-Pro) is involved in various steps of viral infection. In this study,we constructed a Carica papaya L. Plant cDNA library to investigate the host proteins interacting with Papaya ringspot virus (PRSV) Nia-Pro using a Sos recruitment two-hybrid system (SRS). We confirmed that the full-length EIF3G , FBPA1, FK.506BP, GTPBP , MSRB1 and MTL from papaya can interact specifically with PRSV Nia-Pro in yeast,resepctively. These proteins play important roles in plant proteins translation, biotic and abiotic stress, energy metabolism and signal transduction. The possible functions of the interactions between these host proteins and Nia-Pro in PRSV infection and host defense are discussed.