Functional and molecular characterization of Pandoraviruses – Pandoravirus

After our discovery of the largest giant virus Megavirus chilensis in 2011, following that of Mimivirus in 2003, we thought we had reached the ultimate limits in virion size and genome complexity. These viruses infecting Acanthamoeba largely encroach on a territory previously thought to be restricted to bacteria, with their particles 0.7 µm in diameter and their 1Mb genome encoding 1000 proteins.
Our recent discovery of Pandoravirus salinus, a new type of giant virus with a 2.5Mb GC-rich DNA genome, demonstrated that the complexity of viruses could reach that of parasitic eukaryotes such as microsporidia. Beyond their record number of genes and particle size (1µm), detailed analyses of the first two Pandoraviruses (P. salinus, from Chile and P. dulcis, from Australia) brought in additional surprises. Although P. salinus and P. dulcis share a unique amphora-like morphology, drastically different from the icosahedral geometry of Mimiviridae, the two viruses are far from being identical. For instance, the genome of P. dulcis only encodes 1500 proteins compared to 2556 for P. salinus. Moreover, a third of their gene content is unique to each of them. Remarkably, only 6% of the Pandoravirus proteins exhibit detectable homologs in other viruses or cellular organisms. In this context of extreme novelty, the proteome analysis of the P. salinus particles was crucial to confirm that Pandoraviruses use the universal genetic code. Yet again, only 7% of the 200 virion proteins resemble known proteins. Finally, despite their huge gene content, the pandoraviruses lack the transcription machinery that would allow them to replicate in the cytoplasm, akin to the Mimiviridae. As small DNA viruses, they depend from host nuclear functions to complete their infectious cycle.
We reported this year the discovery of the first member of a third family of giant virus, Pithovirus sibericum. This virus was revived from a sample of 30,000-year old Siberian permafrost. Despite its particle shape globally similar to the one of Pandoraviruses, the infectious cycle and genome content of Pithovirus is totally different. Its surprisingly "small" 600 kb genome is AT-rich and only encodes 470 proteins. Once again, 68% do not have homologs. Finally, in contrast with Pandoraviruses, but like the Mimiviridae, Pithovirus particles are packing their own transcriptional machinery allowing them to replicate in the host cytoplasm, leaving the nucleus intact throughout the infectious cycle.
The above discoveries, strongly suggests that the viral world remains an untapped reservoir of unknown biochemical processes, cellular manipulation tools and enzymes. Our project aims at understanding how Pandoraviruses and Pithoviruses manipulate the standard biochemical building blocks of the living world using vastly different proteins. We therefore designed a systematic approach of Pandoraviruses and Pithoviruses physiologies through a multidisciplinary effort combining proteomic, transcriptomic, structural biology and bioinformatic studies. For this, we will first perform detailed proteomic and transcriptomic analyses of the viruses’ infectious cycles. A subset of the most promising viral proteins will be selected and characterized at the structural, functional and interactome levels. New methods to extract the content of these resilient virions to perform interaction studies in native conditions will be developed. Complementary in vivo studies will be performed to elucidate the initial infection processes, viral replication and the unique mechanisms by which the unique amphora-shaped particles are built. These combined approaches will provide clues on the biology of the viruses and on the way they interact with the host cellular machinery. The study of these giant viruses should provide many opportunities for fundamental discoveries in metabolism, cell biology, structural biology and molecular evolution, as well as unforeseen applications in biotechnology and medicine.