03 March 2016

Virus to virus wolf

Giant viruses have found an antivirus system

Alexander Ershov, N+1

A scientific group of virologists led by Didier Raoul from the University of Aix-Marseille has discovered an analogue of the bacterial antiviral CRISPR/Cas system in mimiviruses. These giant parasitic amoeba viruses were discovered just a few years ago. It is known that they suffer from their own parasites – virophages – and, judging by new data, they can use the CRISPR/Cas analogue for self-defense. The study is published in Nature (Levasseur et al., MIMIVIRE is a defense system in mimivirus that confers resistance to virophage).

Mimiviruses are giant viruses by the standards of the microcosm, distantly related to the smallpox virus. They have a very large genome, exceeding one million base pairs in length (for comparison, the genome of E. coli is 4.6 million base pairs, mycobacteria – 580 thousand pairs). Virions (that is, particles) of mimiviruses are surrounded by a cell membrane, and large chunks of DNA "stolen" from their hosts are found in their genome, for example, ribosomal RNA genes that are very rare in viruses. Mimiviruses parasitize free-living freshwater amoebas of the genus Acanthamoeba, which themselves can act as facultative parasites, for example, cause encephalitis.

Following the discovery of mimiviruses, several other groups of similar giant cell-free organisms (megaviruses, marselviruses, pandoraviruses) were discovered. In 2008, one of them managed to find their own parasites, called virophages – by analogy with the "devouring bacteria" bacteriophages (the first virophages were Sputnik1, Sputni2, and so on).


Later it turned out that similar virophages infect other giant viruses. In addition, it was found that virophages themselves contain parasitic genetic elements (called transpovirons), which bring the nesting of parasitism to three levels.

The existence of virophages in giant viruses just prompted the authors of the new work to the need to find immunity, with which they could defend themselves against parasites.

The search was carried out as follows. As a test virophage, the scientists chose the recently discovered virophage Zamilon (Zamilon). Zamilon differs from the more well-known Sputnik virophages in that it is able to infect only two of the three lines of mimiviruses: B and C. All the mimiviruses of line A that were available in the researchers' reports turned out to be resistant to zamilon.

Then the scientists sequenced all available virus strains of all three lines and found several dozen more mimiviruses, bringing the total number of genomes read to 60. According to the scientists, the immunity of mimiviruses of line A, as in the case of bacterial CRISPR/Cas, should have been associated with the presence of viral DNA fragments in their genome "dossier". In the case of CRISPR/Cas, these fragments (spacers) are located in a special place of the genome (between repeating sequences) and are surrounded by genes responsible for the search and destruction of parasitic DNA.

Sequencing showed that although mimiviruses do not have repeats typical of CRISPR (otherwise they would have been found long ago), all representatives of line A have one short fragment completely identical to a piece of the genome of virophage zamilon. At the same time, in mimiviruses of other lines, this sequence (28 nucleotides) occurs only in a single case. More importantly, the 15-nucleotide part of this fragment was repeated four times in the genome of the "stable" first line, but there were no such repeats at all in the unstable lines B and C. Thus, repeats of a fragment of the genome of a potential parasite make mimiviruses resistant to its infection.

Analysis of the genomic environment of these repeats allowed the authors of the article to detect several proteins related to those traditionally associated with CRISPR repeats in bacteria. Among them were both DNA-destroying exonuclease enzymes and nucleic acid-untangling helicases. Interestingly, a decrease in the activity of these genes with the help of RNA interference led to a significant loss of immunity - the rate of replication of the virophage in a cell infected with mimivirus increased tenfold. Three of the 27 candidate genes located near 15-nucleotide repeats turned out to be key for immunity.

Thus, the antiviral system found by the authors in mimiviruses really protects them from virophages (at least one of them), although it is arranged significantly differently than bacterial CRISPR/Cas. That is why it has not yet been found by the methods that are now used to study genomes. Nevertheless, the mimivirus system relies on the same principle: the creation of an archive of DNA fragments of a "potential enemy" and their targeted destruction. The study of a new antiviral system opens up the possibility of searching for new directed nucleases by analogy with those that were discovered during the study of the CRISPR/Cas system. The main disadvantage of the Cas9 nuclease currently used is its size – it is very large for the currently existing viral delivery systems to cells. The use of a new, smaller mimivirus nuclease may help solve this problem.

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