Molecular Counting in Moloney Murine Leukemia Virus Assembly.

[摘要] The Moloney murine leukemia virus (MoMLV) particle is composed of roughly 2,500 Gag polyproteins, 125 Gag-Pol polyproteins, and precisely two copies of genomic RNA (gRNA) in the form of a dimer. The Gag polyprotein is responsible for directing MoMLV assembly and for recruiting both Gag-Pol and the gRNA dimer for incorporation during assembly. Each of the thousands of Gag molecules in a MoMLV particle contains the domains required to bind both Gag-Pol and gRNA. The mechanisms that account for the conserved asymmetrical numerical proportions of these molecules are unknown. The goal of this dissertation was to examine the parameters that define the molecular proportions of Gag, Gag-Pol, and the gRNA dimer in the MoMLV particle.In order to test the hypothesis that only a small number of Gag molecules within the MoMLV particle are required to recruit gRNA for incorporation during MoMLV assembly, MoMLV particles were produced that contained variable proportions of two types of Gag molecules: one that was able to participate in high-affinity gRNA packaging interactions and one that was not. Utilizing these mixed particles, results showed that MoMLV gRNA was packaged in the absence of a full complement of packaging competent Gag.To test the hypothesis that the molecular counting mechanism responsible for incorporating MoMLV Gag-Pol and the gRNA dimer are distinct, an experimental system was constructed that allowed the proportion of Gag to Gag-Pol and Gag to gRNA to be manipulated. Using this system, our results showed that the incorporation of Gag-Pol into MoMLV is a stochastic process that is dependent on intracellular expression ratios. In contrast, incorporation of gRNA was shown to be a non-random and saturable, thus not strictly dependent of intracellular expression ratios. These results support a model whereby the proportions of Gag and Gag-Pol co-assembled are determined by a distinct mechanism from that which defines Gag and gRNA ratios.

[发布机构] University of Michigan

[效力级别] Microbiology and Immunology

[关键词] Molecular Counting in Moloney Murine Leukemia Virus Assembly;Microbiology and Immunology;Science;Microbiology and Immunology