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Assembly of the coronavirus envelope: homotypic interactions between the M proteins
C A de Haan1, H Vennema, P J Rottier
1Institute of Virology, Department of Infectious Diseases and Immunology, Faculty of Veterinary Medicine, and Institute of Biomembranes, Utrecht University, 3584 CL Utrecht, The Netherlands.
Abstract:
The viral membrane proteins M and E are the minimal requirements for the budding of coronavirus particles. Since the E protein occurs in particles only in trace amounts, the lateral interactions between the M proteins apparently generate the major driving force for envelope formation. By using coimmunoprecipitation and envelope incorporation assays, we provide extensive evidence for the existence of such M-M interactions. In addition, we determined which domains of the M protein are involved in this homotypic association, using a mutagenetic approach. Mutant M proteins which were not able to assemble into viruslike particles (VLPs) by themselves (C. A. M. de Haan, L. Kuo, P. S. Masters, H. Vennema, and P. J. M. Rottier, J. Virol. 72:6838-6850, 1998) were tested for the ability to associate with other M proteins and to be rescued into VLPs formed by assembly-competent M proteins. We found that M proteins lacking parts of the transmembrane cluster, of the amphipathic domain, or of the hydrophilic carboxy-terminal tail, or M proteins that had their luminal domain replaced by heterologous ectodomains, were still able to associate with assembly-competent M proteins, resulting in their coincorporation into VLPs. Only a mutant M protein in which all three transmembrane domains had been replaced lost this ability. The results indicate that M protein molecules interact with each other through multiple contact sites, particularly at the transmembrane level. Finally, we tested the stringency with which membrane proteins are selected for incorporation into the coronavirus envelope by probing the coassembly of some foreign proteins. The observed efficient exclusion from budding of the vesicular stomatitis virus G protein and the equine arteritis virus M protein indicates that envelope assembly is indeed a highly selective sorting process. The low but detectable incorporation of CD8 molecules, however, demonstrated that this process is not perfect.
Insights
Coronavirus envelope formation relies on M protein interactions. These M-M interactions, particularly at the transmembrane level, drive viral budding and selective protein incorporation into new virus particles.
Area of Science:
- Virology
- Molecular Biology
- Structural Biology
Background:
- Coronavirus particle budding requires viral membrane proteins M and E.
- The M protein appears to be the primary driver of envelope formation due to its abundance and self-interaction.
- Understanding M protein interactions is crucial for comprehending coronavirus assembly.
Purpose of the Study:
- To investigate the homotypic interactions between coronavirus M proteins.
- To identify the specific domains of the M protein involved in self-assembly.
- To assess the selectivity of protein incorporation into the viral envelope.
Main Methods:
- Coimmunoprecipitation assays to detect M protein associations.
- Envelope incorporation assays using virus-like particles (VLPs).
- Mutagenetic analysis of M protein domains and testing of mutant protein assembly and incorporation.
Main Results:
- Extensive evidence supports M-M interactions as the driving force for coronavirus envelope formation.
- M protein association with itself is mediated by multiple contact sites, especially within the transmembrane regions.
- Mutations in transmembrane domains disrupt M protein self-assembly and VLP incorporation.
- The coronavirus envelope assembly process demonstrates high selectivity, efficiently excluding foreign membrane proteins like VSV-G and EAV-M, with minor exceptions like CD8.
Conclusions:
- Coronavirus M proteins interact homotypically through multiple sites, primarily at the transmembrane level, to drive envelope formation.
- The M protein's transmembrane domains are critical for its self-assembly and incorporation into virus particles.
- Coronavirus envelope assembly is a highly selective process, ensuring the proper composition of viral particles.