Genetic Analysis of the Major Capsid Protein of the Archaeal Fusellovirus SSV1: Mutational Flexibility and Conformational Change

Abstract

Viruses with spindle or lemon-shaped virions are rare in the world of viruses, but are common in viruses of archaeal extremophiles, possibly due to the extreme conditions in which they thrive. However, the structural and genetic basis for the unique spindle shape is unknown. The best-studied spindle-shaped virus, Sulfolobus Spindle-shaped Virus 1 (SSV1), is composed mostly of the major capsid protein VP1. Similar to many other viruses, proteolytic cleavage of VP1 is thought to be critical for virion formation. Unlike half of the genes in SSV1, including the minor capsid protein gene VP3, the VP1 gene does not tolerate deletion or transposon insertion. To determine the role of the VP1 gene and its proteolysis for virus function, we developed techniques for site-directed mutagenesis of the SSV1 genome and complemented deletion mutants with VP1 genes from other SSVs. By analyzing these mutants, we demonstrate that the N-terminus of the VP1 protein is required, but the N-terminus, or entire SSV1 VP1 protein, can be exchanged with VP1s from other SSVs. However, the conserved glutamate at the cleavage site is not essential for infectivity. Interestingly, viruses containing point mutations at this position generate mostly abnormal virions.

Archaeal Viruses from High-Temperature Environments

“…These tools include viral (SSV1) and plasmid-based shuttle systems, as well as CRISPR based gene deletion, mutagenesis, and silencing techniques [ 79 ]. Genetic systems have also been developed for some archaeal viruses, for example, the SSV1 genetic system has provided insightful into the function of archaeal viral genes [ 80 , 81 , 82 ]. Both random and targeted mutagenesis of the SSV1 viral genome surprisingly revealed that only half of the viral genes encoded by the SSV1 genome (16/35 viral genes) are essential for infectivity [ 81 ].…”

“…Genetic systems have also been developed for some archaeal viruses, for example, the SSV1 genetic system has provided insightful into the function of archaeal viral genes [ 80 , 81 , 82 ]. Both random and targeted mutagenesis of the SSV1 viral genome surprisingly revealed that only half of the viral genes encoded by the SSV1 genome (16/35 viral genes) are essential for infectivity [ 81 ]. Furthermore, the requirement for a particular viral gene correlates well with its degree of conservation among the Fuselloviridae .…”

Genomics, Transcriptomics, and Proteomics of SSV1 and Related Fusellovirus: A Minireview

“…Transcripts T 4/7/8 , T 9 , and T 1/2 contain most of the core-genome ORFs, while the shell and variable portions of the genome are identifiable in transcripts T 5 and T 6 . The latter are highly flexible regions in the fuselloviruses and contain most of the non-essential genes identified in previous functional genomics studies [ 11 , 34 , 35 ]. Among the genes in the soft-core genome, nine are present in all genomes and constitute the absolute-core (VP1, VP4, D355-integrase, A82, A92, B251-AAA ATPase, C166, A154, and B115), while four genes are present in over 90% of the genomes (VP3, B129, B277, and B78).…”

“…A transcriptional activity has been detected also for d335 , encoding the SSV1 integrase. Then, the carrier state of SSV1 is sustained by the expression of a few genes, among which only those encoding the structural capsid proteins (VP1, VP2, and VP3) [ 34 , 52 ] and the integrase have been functionally characterized [ 22 ]. Moreover, a recent study has revealed that most of the lysogenic genes ( vp1, vp2, d355, and a291 ) are apparently essential for SSV1 infectivity, except for c124 and vp3 [ 16 ].…”

“…All these virus mutants were tested for their ability to infect the permissive host S. solfataricus . By doing so, the authors have shown that nearly half of the SSV1 genes (16 out of 35 ORFs) were dispensable for the SSV1 virions infectivity and that most of essential genes are highly conserved within the Fuselloviridae family and constitute the so-called “fusellovirus core” [ 34 ]. Whereas almost the entire T5 early transcript was shown to be dispensable (7 out of 10 ORFs), many proteins encoded by the T6 early transcript were essential, which is in line with the abundance of well-conserved fusellovirus core genes in this transcript.…”