The virion N protein of infectious bronchitis virus is more phosphorylated than the N protein from infected cell lysates

Because phosphorylation of the infectious bronchitis virus (IBV) nucleocapsid protein (N) may regulate its multiple roles in viral replication, the dynamics of N phosphorylation were examined. P-32-orthophosphate labeling and Western blot analyses confirmed that N was the only viral protein that was phosphorylated. Pulse labeling with P-32-orthophosphate indicated that the IBV N protein was phosphorylated in the virion, as well as at all times during infection in either chicken embryo kidney cells or Vero cells. Pulse-chase analyses followed by immunoprecipitation of IBV N proteins using rabbit anti-IBV N polyclonal antibody demonstrated that the phosphate on the N protein was stable for at least 1 h. Simultaneous labeling with P-32-orthophosphate and H-3-leucinc identified a 3.5-fold increase in the P-32:H-3 counts per minute (cpm) ratio of N in the virion as compared to the P-32:H-3 cpm ratio of N in the cell lysates from chicken embryo kidney cells, whereas in Vero cells the P-32:H-3 cpm ratio of N from the virion was 10.5-fold greater than the P-32:H-3 cpm ratio of N from the cell lysates. These studies are consistent with the phosphorylation of the IBV N playing a role in assembly or maturation of the viral particle. (C) 2005 Elsevier Inc. All rights reserved.