Cowpea mosaic virus (CPMV) replication occurs in close association with small membranous vesicles in the host cell. The CPMV RNA1-encoded 60 kDa nucleotide-binding protein ('60K') plays a role in the formation of these vesicles. In this study, five cellular proteins were identified that interacted with different domains of 60K using a yeast two-hybrid search of an Arabidopsis cDNA library. Two of these host proteins (termed VAP27-1 and VAP27-2), with high homology to the VAP33 family of SNARE-like proteins from animals, interacted specifically with the C-terminal domain of 60K and upon transient expression colocalized with 60K in CPMV-infected cowpea protoplasts. eEF1-beta, picked up using the central domain of 60K, was also found to colocalize with 60K. The possible role of these host proteins in the viral replicative cycle is discussed.