Preliminary Research On Structure And Function Of SARS-CoV 7a Protein

A novel coronavirus was identified as the etiological agent of recent severe acute respiratory syndrome epidemic. The genome of severe acute respiratory syndrome-associated coronavirus (SARS-CoV) contains four structural genes encoding M, E, S and N that are homologous to genes found in other coronaviruses, and also contains six subgroup-specific open reading frames (ORFs) encoding larger than 50 amino acids. Analysis of bioimformation shows that there are no homologous genes available about these ORFs, inferring some of which might be related to the pathogenesis of SARS-CoV. It was reported that ORF7a, one of these subgroup specific genes, was identified in virus infected cells and expression of ORF7a can result in apoptosis via a caspase-dependent pathway, so we predict that 7a may participate in pathogenesis induced by SARS-CoV. This study is proposed to investigate the structure and function of 7a protein, in order to give a clue to revealing the molecular mechanism. The cellular location and topology of 7a were determined by immunocytochemistry and we found that 7a protein was located at Golgi, with its N-terminus facing the cytoplasm and its C-terminus facing Golgi matrix. Meanwhile, we have noticed that HEK293 cells transfected with 7a grow slower than control cells, indicating expression of 7a protein may inhibit cell proliferation. Here, we observed that transient expression of ORF7a protein fused with myc or GFP tags at its N or C terminus inhibited cell growth and prevented BrdU incorporation in different cultural cells, suggesting that ORF7a expression may regulate cell cycle progression. Analysis by flow cytometry demonstrated that ORF7a expression was associated with blockage of cell cycle progression at G0/G1 phase in HEK 293 cells after 24 to 60h post-transfection. Similar results were observed in COS-7 and Vero cells.