The SARS coronavirus (SARS-CoV) open reading frame 7a (ORF 7a) encodes a 122 amino acid accessory protein. culture cells transiently expressing 7a and Ap4A-hydrolase tagged with EGFP and Ds-Red2 respectively show these proteins co-localize in the cytoplasm. Background Severe acute respiratory syndrome coronavirus (SARS-CoV) has been shown to be the etiological agent for the global SARS outbreak in the winter 2002/2003 that affected about 30 countries [1]. SARS-CoV is an enveloped, positive-sense RNA computer virus with ~30 kb genome. It contains 14 potential ORFs. Some of these ORFs encode proteins that are homologues to the structural proteins founded in other coronaviruses, namely the replicase (ORF 1a and 1b), membrane, nucleocapsid, envelope and spike proteins [2,3]. Other ORFs encode group-specific or accessory proteins which are unique to SARS-CoV. Accessory proteins are not necessary for viral replication in cell culture systems and in mice, but may be important for virus-host interactions and thus may contribute to viral strength and/or pathogenesis em in vivo /em [4-6]. Protein 7a (also known as ORF SKI-606 cost 8, U122 and X4 protein [2,3,7]), 122 amino acids in length, shows no significant similarity to any other viral or non-viral proteins. The ORF 7a gene is usually conserved in SKI-606 cost all SARS-CoV strains [8], and sequence analysis predicts that ORF 7a encodes a type I transmembrane protein. The crystal structure of the luminal domain of the 7a protein has been resolved, revealing a structure unexpectedly comparable in fold and topology to members of the immunoglobulin superfamily [9]. It has been exhibited that 7a is usually incorporated into SARS-CoV particles by interacting with viral structural proteins E and M [10,11]. In addition, 7a interacts with the viral proteins TFRC 3a and S [10,12], and these proteins may form a complex during viral contamination. Recombinant mutant SARS-CoV lacking the 7a gene is completely viable in cultural cells and mice [4]; therefore, 7a protein is usually dispensable for computer virus growth and replication but may play role in virus-host interactions. The 7a protein seems to have diverse biological functions in cultured cells. Over-expression of ORF 7a induces apoptosis via the caspase-dependent pathway [13] and inhibits cellular protein synthesis by activation of p38 MAPK [14]. The induction of apoptosis by the 7a SKI-606 cost protein is dependent on its conversation with the Bcl-XL protein and other pro-survival proteins (Bcl-2, Bcl-w, Mcl-1 and A1) [15]. In addition, 7a can block cell cycle progression at the G0/G1 phase via the cyclin D3/pRb pathway [16]. Also, conversation between 7a and hSGT (human small glutamine-rich tetricopeptide repeat containing protein) has been exhibited although the biological significance of this interaction needs to be further elucidated [17]. Taken together, these observations suggest that the 7a protein interacts with several host cell proteins and may play a role in the SARS-CoV pathogenesis. We performed a yeast-two-hybrid screening using a commercially prepared human lung cDNA library as the source of the “prey” cDNAs and using a full-length ORF 7a as the “bait”. Among the potential novel 7a interacting partners, Ap4A-hydrolase was identified. Its conversation with 7a was confirmed by co-immunoprecipitation and co-localization experiments in transiently transfected cultured human cells. Ap4A-hydrolase belongs to the Nudix (nucleoside diphosphate linked to x) hydrolases, which are a superfamily of enzymes required for maintenance of physiological homeostasis by metabolizing signaling molecules and potentially toxic substances. Ap4A-hydrolase is found in all higher eukaryotes and contributes to regulation of the intracellular level of “allarmone” nucleotide Ap4A [18,19]. It is an asymmetrically-cleaving enzyme, catalyzing the reaction (Ap4ATP+AMP). The intracellular concentration of Ap4A has been shown to increase in cells after heat, oxidative, nutritional or DNA damage stresses [20]. A recent study exhibited that Ap4A-hydrolase belongs to the transcriptional SKI-606 cost regulation network.