Abstract
Deregulation of the cell cycle is a common strategy employed by many DNA and RNA viruses to trap and exploit the host cell machinery toward their own benefit. In many coronaviruses, the nucleocapsid protein (N protein) has been shown to inhibit cell cycle progression although the mechanism behind this is poorly understood. The N protein of severe acute respiratory syndrome-coronavirus (SARS-CoV) bears signature motifs for binding to cyclin and phosphorylation by cyclin-dependent kinase (CDK) and has recently been reported by us to get phosphorylated by the cyclin-CDK complex (Surjit, M., Kumar, R., Mishra, R. N., Reddy, M. K., Chow, V. T., and Lal, S. K. (2005) J. Virol. 79, 11476–11486). In the present study, we prove that the N protein of SARS-CoV can inhibit S phase progression in mammalian cell lines. N protein expression was found to directly inhibit the activity of the cyclin-CDK complex, resulting in hypophosphorylation of retinoblastoma protein with a concomitant down-regulation in E2F1-mediated transactivation. Coexpression of E2F1 under such conditions could restore the expression of S phase genes. Analysis of RXL and CDK phosphorylation mutant N protein identified the mechanism of inhibition of CDK4 and CDK2 activity to be different. Whereas N protein could directly bind to cyclin D and inhibit the activity of CDK4-cyclin D complex; inhibition of CDK2 activity appeared to be achieved in two different ways: indirectly by down-regulation of protein levels of CDK2, cyclin E, and cyclin A and by direct binding of N protein to CDK2-cyclin complex. Down-regulation of E2F1 targets was also observed in SARS-CoV-infected VeroE6 cells. These data suggest that the S phase inhibitory activity of the N protein may have major significance during viral pathogenesis.
Highlights
Mitotic cells undergo repeated cycles of division in order to produce daughter cells, a process essential for the maintenance of tissue homeostasis
N of severe acute respiratory syndrome (SARS)-CoV Inhibits Cy-cyclin-dependent kinases (CDKs) and Blocks S Phase Progression recently shown that the nucleocapsid protein (N protein) of the SARSCoV localizes to the nucleus as well as cytoplasm and bears signature motifs for binding to the cyclin box and phosphorylation by the CDK, and we have experimentally demonstrated it to be a substrate of the cyclin-CDK complex [11]
We asked whether the possession of signature motifs and phosphorylation by the CDK is a strategy employed by the N protein to manipulate the host cell cycle machinery toward its benefit by acting as a competitive inhibitor to in vivo substrates of the cyclin-CDK complex
Summary
Mitotic cells undergo repeated cycles of division in order to produce daughter cells, a process essential for the maintenance of tissue homeostasis. Rb phosphorylation too was effectively inhibited by lysate immunoprecipitated with CDK2 antibody from N protein-expressing cells (Fig. 1F).
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
