Giant Viruses
Exploring the Land of the Giants
by Antonia Evripioti & Stuart MacNeill
Viruses are the most abundant and widespread biological entity on Earth. The vast majority are small (much smaller than even the smallest bacterial cells), cannot be seen under the light microscope and encode only a small number of viral proteins, instead being reliant on the host cell to provide most of the materials required for their propagation.
This textbook view of virus biology has been shaken up in recent years, however, by a series of studies that have welcomed new additions to the virus world in the shape of giant viruses infecting amoeba and marine microorganisms.
Researchers in St Andrews are now turning their attention to understanding how these viral giants propagate their genetic material through the generations by embarking on a comprehensive biochemical analysis of the molecular machinery of viral DNA replication. This work will lead to insights into the lifestyle of the giant viruses and offer opportunities for the development of anti-viral agents.
Mimivirus, the first giant
The prototype of the giant viruses, named Mimivirus, was first identified infecting amoeba living in a water cooling tower in Yorkshire - in fact, Mimivirus particles, which are so large that they are visible under the light microscope, had initially been mistaken for bacterial cells until being identified as viral particles by the laboratory of Didier Raoult in Marseilles in 2003.
With a diameter of 750 nm, Mimivirus particles are larger than many bacteria, a completely unexpected result.
The next surprise came when the Mimivirus genetic material was sequenced, revealing a genome of over 1 million base pairs of DNA, more than twice the size of the largest previously sequenced viral genome, and the capacity to encode than 1000 proteins. (For comparison, the genome of the parasitic bacterium Mycoplasma genitalium spans 600,000 base pairs and encodes only around 500 proteins, whereas that of the free-living bacterium Pelagibacter ubique spans 1,300,00 base pairs and encodes 1300 proteins).
Since the discovery of Mimivirus a handful of other giant viruses have been identified, including the closely-related Megavirus, which boasts a 10% larger genome, encodes over 1200 proteins and also infects amoeba, and Cafeteria roenbergensis virus (CroV), which is 750,000 base pairs in length, encodes over 500 proteins and infects the abundant marine flagellate Cafeteria roenbergensis. Evidence is now emerging that many of these viral giants are members of the same family, tentatively termed the Megaviridiae. Debate continues to rage over the evolutionary origins of the Megaviridiae and in particular, whether they constitute a fourth domain of life on Earth.
Should we be afraid of the giants?
To date, none of the giant viruses has been demonstrated to be pathogenic to humans, despite some early indirect evidence linking Mimivirus to a case of pneumonia in an unfortunate laboratory technician. However, marine giant viruses may impact on the ocean ecosystems, causing population crashes in their host organisms. Understanding giant virus biology is potentially very important for future ecosystem management.
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Giant viruses
The viral giants are members of a large supergroup of eukaryotic viruses called the NCLDVs (nucleocytoplasmic large DNA viruses) that includes poxviruses (smallpox, cowpox, etc.), asfarviruses and iridoviruses amongst others. All have double-stranded DNA genomes, encode a conserved core set of proteins required for their replication and may be derived from a common ancestor. The three viruses under investigation in St Andrews are Mimivirus and CroV (both of which have recently been assigned to a proposed new sub-family of the NCLDVs, the Megaviridae) and Marseillevirus (which appears to be most closely related to the Iridoviridae sub-family).
First steps in the Land of the Giants
To begin to understand how the viral giants replicate their genetic material, Antonia Evripioti, a PhD student in the Chromosome replication and genome stability group has begun to explore the biology of what are likely to be key components of the giant virus replication machinery, molecules known as sliding clamps. Sliding clamps are evolutionarily conserved protein complexes that are able to fully or partially encircle the DNA double helix and which act as a stable platform onto which other replication factors assemble.
To determine the properties of the giant virus clamps, Antonia will first use synthetic genes to express the clamps encoded by Mimivirus, CroV and Marseillevirus individually in bacteria. The proteins will then be purified and biochemical analysis can begin. Computational analysis suggests that the three clamps chosen for this study are likely to have strikingly different molecular make-ups - the initial aim of Antonia's work will be to determine whether this is the case. Once the biology of the sliding clamps is established, the next phase of research will focus on two additional components of the core replication machinery: replication factor C (the five-subunit complex predicted to load the putative trimeric Mimivirus and CroV sliding clamps onto DNA) and the Mimivirus, CroV and Marseillevirus DNA polymerases (the enzymes that perform synthesis of new DNA strands).