Fermentation genes in a giant algal virus

TetV infected Tetraselmis

TetV infected Tetraselmis. Arrow indicates virus particle. Inset, single particle. Image credit.

The latest giant virus discovery is Tetraselmis virus 1, which infects green algae. It is unusual because it encodes enzymes involved in fermentation. Green beer, anyone?

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TWiV 484: Float like a mimivirus STING like a bat

The TWiVumvirate discuss the giant Tupanvirus, with the longest tail in the known virosphere, and dampened STING dependent interferon activation in bats.

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Only the ribosome is lacking

tupanvirusIf you know anything about me, you know that I’m mad about viruses. Although this madness extends to everything viral, I have a peculiar fondness for giant viruses. A new giant virus has been found that not only looks different from all the others, but has an amazing set of genes.

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TWiV 474: Call me fish meal

The TWiVanguardians take on Bodo saltans virus, a leviathan which infects an abundant flagellated eukaryote in Earth’s waters.

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Bodo saltans virus, an abundant giant aquatic Mimivirus

Bodo saltans

Bodo saltans

The discovery of Mimivirus in a French cooling tower amazed virologists and changed our view of the biology and evolution of giant viruses. Since then, many other giant viruses have been identified, and with three exceptions, they all appear to infect species of Acanthamoeba. Now a new member of the Mimivirus family has been discovered that infects the flagellated eukaryote Bodo saltans (pictured: image credit).

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TWiV 440: I hardly noumeavirus

No problem not being nice to Dickson in this episode, because he’s absent for a discussion of a new giant virus that replicates in the cytoplasm yet transiently accesses the nucleus to bootstrap infection.

You can find TWiV #440 at microbe.tv/twiv, or listen below.

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Forget the fourth domain of life

three domains of lifeWhen giant viruses were discovered – with genomes much larger than any previously seen – some suggested that they had descended from a fourth domain of life (the current three are bacteria, archaea, and eukaryotes). Part of the reason for such a claim was the finding of homologs of bacterial and eukaryotic genes, including molecules involved in translation. Analysis of new giant viruses encoding even more components of the translation machinery has thrown cold water on the fourth domain hypothesis.

Klosneuvirus, with a 1.57 million base pair DNA genome, was discovered in a wastewater treatment plant in Austria, and three related viruses – Indivirus, Hokovirus, and Catovirus – were found in environmental samples.  Sequence analyses suggests that these viruses should be classified in a subfamily of the Mimiviridae.

The Klosneuviruses encode far more components of the translational machinery than do mimiviruses – 25 tRNAs, 19 aminoacyl tRNA synthetases, 11 initiation and elongation proteins, a chain release factor, and tRNA modifying enzymes.

Phylogenomic analyses demonstrate that the aminoacyl tRNA synthetase and translation factor genes are likely derived from protists. This finding is not compatible with the hypothesis that these viruses are derived from a fourth domain of life. It is more likely that smaller ancestors of giant viruses acquired these genes from known eukaryotes.

Why these components of the translational system have been maintained in these giant virus genomes is an excellent question. They might confer some advantage to the viruses, for example when host translation is shut off as a viral defense. Having components of the translational apparatus might allow viral protein synthesis to proceed.

Note that genes encoding ribosomal RNAs or proteins have not been found in any virus. In fact no virus encodes a complete protein synthesis machinery. Maybe they have yet to be discovered? Or perhaps these energetically costly activities are best left to the cell?

 

TWiV 419: The selfless gene

The TWiVrific gang reveal how integration of a virophage into the nuclear genome of a marine protozoan enhances host survival after infection with a giant virus.

You can find TWiV #419 at microbe.tv/twiv, or listen below.

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Altruistic viruses

Cafeteria roenbergensisVirophages (the name means virus eater) were first discovered to replicate only in amoeba infected with the giant mimiviruses or mamaviruses.  They reduce yields of the giant viruses, and also decrease killing of the host cell. Another virophage called mavirus has been found to integrate into the genome of its host and behaves like an inducible antiviral defense system (link to paper).

The host cell of the virophage mavirus is Cafeteria roenbergensis, Cro (pictured), a marine phagotropic flagellate, that is infected with the giant virus CroV (Cafeteria roenbergensis virus). When Cro cells are infected with a mixture of mavirus and CroV, the virophage integrates into the host cell genome. There it remains silent; the cells survive, and no virophage particles are produced. Such cells can be called lysogens, a name applied to bacteria containing integrated bacteriophage genomes, or prophages.

How does the mavirus genome integrate into the Cro cell? The viral genome encodes an integrase, an enzyme that cuts host DNA and inserts a copy of the viral genome. Retroviruses achieve the same feat via an integrase.

When Cro-mavirus lysogens are infected with CroV, the integrated mavirus genome is transcribed to RNA, the viral DNA replicates, and new virus particles are formed. These virophages inhibit the replication of CroV by 100-1000 fold. As a consequence, the host cell population survives.

These findings suggest that the virophage mavirus is altruistic: induction of the integrated genome leads to killing of the host cell, but other members of the cell population are protected. Altruism is not unknown in Nature, but how it evolved is an intriguing question.

All this work was done in a laboratory. It will be necessary to determine if integration of mavirus into Cro cells in the wild has any influence on the ecology of these organisms.

Do giant viruses have a CRISPR-like immune system or a protein restriction factor?

Zamilon virophageA battle is brewing between two research groups in Marseille, France that are involved in the discovery and study of giant viruses. Didier Raoul and colleagues believe that they have discovered a CRISPR-like, DNA based defense system in mimivirus that confers resistance to virophage (paper link). Claverie and Abergel disagree: they think that the defense system involves proteins, not nucleic acids (paper link).

Virophages are DNA viruses that can only replicate in cells infected by giant viruses like mimivirus. Their name, which means ‘virus eater’, comes from the observation that they inhibit mimivirus replication. A specific virophage called Zamilon was discovered that can inhibit the replication of lineage B and C mimivirus but not lineage A.

Examination of the DNA sequences of 60 different mimivirus strains revealed that the genomes of lineage A contained a 28 nucleotide sequence identical to Zamilon virophage. This sequence was not found in any lineage B mimivirus and in only one out of 19 lineage C mimiviruses. In addition, a 15 nucleotide subset of this sequence is repeated four times in the lineage B and C mimivirus genomes.

Near the 15 nucleotide Zamilon-derived repeated sequences in lineage B and C mimivirus genomes are genes encoding several proteins related to components of the bacterial CRISPR-Cas system. These include a nuclease, an RNAse, and an ATP-dependent DNA helicase.

The CRISPR system provides defense against invading DNA. When a foreign DNA, such as a bacteriophage genome, enters a bacterial cell, some is fragmented and integrated into the CRISPR locus as a ’spacer’ (sequences in the foreign DNA are called ‘protospacers’). Following transcription, CRISPR RNAs (crRNA) are processed by a multiprotein complex to produce ~60 nucleotide RNAs. When the spacer of a crRNA base pairs with a complementary sequence in an invading DNA molecule, CRISPR-associated endonucleases cleave the DNA. The integration of the sequences of the invading DNA into the host cell genome, from which they can be mobilized in the form of crRNAs, provides a form of “memory” and acquired immunity. It should be noted that there are six known types of CRISPR systems that differ in their components and mechanisms.

Because the CRISPR-Cas system is an adaptive immune system that protects bacteria and Archaea from virus infections and invasion of foreign DNA, the authors propose that they have discovered a new adaptive immune system that protects mimiviruses from virophage infection. They call this system mimivirus virophage resistance element, or MIMIVIRE.

The authors provide experimental support for their hypothesis by showing that silencing the genes encoding the endonuclease, the helicase, and the repeated insert using siRNA allows Zamilon replication in mimivirus-infected cells.

Claverie and Abergel think that Raoult and colleagues are wrong (paper link). They provide three reasons to dispute their findings, and ‘propose a simpler protein-based interaction model that explains the observed phenomena without having to extend the realm of adaptive immunity to the world of eukaryotic viruses, a revolutionary step that would require stronger experimental evidences.’

The first problem is that mimivirus and Zamilon virophage replicate in the same location in the infected cell, making a CRISPR-like defense system difficult to conceptualize. In contrast, CRISPR sequences reside in the bacterial genome, from which RNAs are produced that target the destruction of invading DNAs elsewhere in the cell.

The second problem is that the Zamilon sequences in the mimivirus genome are not regularly spaced or flanked by recognizable repeats, a hallmark of the CRISPR system (the name stands for ‘clustered regularly interspersed short palindromic repeats). However it should be noted that type VI CRISPR systems have no CRISPR locus and likely function via mechanisms that are different from other CRISPR systems.

Finally, Claverie and Abergel argue that there is no way for the proposed nucleic acid defense system to distinguish between the virophage and the virophage sequences in the mimivirus genome. In some CRISPR systems this discrimination is achieved by protospacer adjacent motifs (PAMs), short (2-5 nt) sequences next to the invader protospacer sequences that are recognized by the endonuclease complex guided by the crRNA. PAMs are not present in the bacterial genome, sparing it from endonucleolytic cleavage. Nevertheless, non PAM-based mechanisms of discriminating invader from host are known, for example, in the type III CRISPR system.

If Raoult and colleagues have not discovered a CRISPR-like mimivirus defense system, then why would silencing the genes encoding CRISPR-like proteins allow Zamilon replication? Claverie and Abergel think that it is not the 15 nucleotide Zamilon repeats that are important to mimivirus, but the encoded amino acids: Asp-Asn-Glu-Ser (DNES in one letter code). They believe that DNES is a motif present in proteins that block Zamilon replication by as yet unidentified mechanisms.

Many cellular proteins have been identified that interfere with virus replication, such as those encoded by interferon induced genes (ISGs). DNES containing proteins that inhibit Zamilon replication would be conceptually analogous, except that they are encoded by a virus, not the host.

Claverie and Abergele appear to have a strong case that mimivirus defense against Zamilon virophage is mediated by protein, not nucleic acid, but further experimentation is certainly needed to support their position. Nevertheless they recognize that the discovery by Raoult and colleagues “remains fascinating even if it falls short of demonstrating the existence of a CRISPR-Cas-like adaptive immune system”.