Many proteins make light work: the role of minor structural proteins in virus replication

Each of the eight simian hemorrhagic fever virus minor structural proteins is functionally important

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Simian hemorrhagic fever virus (SHFV) is a plus-strand RNA virus in the family Arteriviridae. The SHFV genome is unique in encoding two sets of four minor structural protein open reading frames (ORFs) instead of one. For other arteriviruses, the four minor structural proteins form a complex located on the virion surface that facilitates virion attachment and entry. It was previously hypothesized that the second set of SHFV minor structural proteins was generated by recombination and that orthologs in the two sets are functionally redundant. To analyze the functional relevance of each of the minor structural proteins, a full-length SHFV cDNA infectious clone was constructed and then a set of mutated clones, each with the expression of one of the minor structural proteins knocked out, were made. All eight of the minor structural proteins were found to be required for the production of extracellular, infectious virions.

Construction of the wild-type, full-length infectious cDNA clone was challenging due to the use of the 15.7 kb RNA SHFV (LVR) genome sequence obtained by our lab in the early 1990s as the reference. An infectious clone with this sequence was stable but viral RNA transcribed from it was not infectious. Resequencing of the viral genome RNA identified 18 nucleotides that differed from the reference sequence. RNA transcribed from a new infectious clone assembled from fragments containing these changes produced virus with replication kinetics comparable to those of parental virus.

The eight minor structural protein ORFs are conserved among divergent SHFV genomes. The different order of the first two minor structural ORFs in the duplicated sets indicates that the second set was not generated by a simple duplication event. This was an initial clue that all of the orthologs might not be functionally redundant. We were surprised to find that none of the mutant clones produced infectious virus indicating that each of the eight proteins has a critical function and that glycoproteins from each set were required for virion infectivity. Detection of a subgenomic mRNA not found in a previous study for one of the minor structural proteins led to the interesting observation that each set of orthologs is expressed from subgenomic mRNAs of similar abundance indicating conservation of expression regulation. Based on what is known about the minor structural proteins of other arteriviruses, discovery of the possible involvement of some of the SHFV minor structural proteins in virion production was surprising.

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Introducing the authors

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About the research

Each of the eight simian hemorrhagic fever virus minor structural proteins is functionally important
Virology, Volumes 462–463, August 2014, Pages 351–362
Heather A. Vatter, Han Di, Eric F. Donaldson, Ralph S. Baric, Margo A. Brinton

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