Analyses of the Role of Structural Changes in the Regulation of Uncoating and Assembly of Alphavirus Cores

In the late stages of alphavirus multiplication virus cores accumulate in the cytoplasm as stable structures, whereas they are unstable during the early stages of infection. Three types of explanation can be put forward to understand this phenomenon: (1) A cellular uncoating process is active early which is inactivated later, (2) the core structure differs between stable cores which accumulate and labile cores which are dissociated, (3) both mechanisms cooperate. A model based exclusively on the first principle involving core disassembly by cellular ribosomes has been proposed, but structural changes of the core may cooperate with the cellular uncoating process. We have therefore isolated cores of the Sindbis alphavirus from the cellular cytoplasm, from virus particles (vi-cores), and from low-pH-treated virus particles. Comparative analyses of the structure of these cores, using crosslinking and proteolytic digestion, and of the stability of the cores in the presence of ribosomesin vitrowere performed. The structural comparisons indicate that the interactions between the molecular components of these cores are very similar and probably identical. In the presence of ribosomes vi-cores were slightly more stable than the two other types of cores. Evidence for a cellular uncoating mechanism is furnished by experiments which analyze the stability of cores in the presence of postmitochondrial cytoplasm or of 60 S ribosomal subunits derived from either uninfected or Sindbis virus-infected cells. The results obtained indicate that structural alterations of the core do not play a role in the regulation of disassembly and assembly of alphavirus cores.