The giant mimivirus 1.2 Mb genome is elegantly organized into a 30-nm diameter helical protein shield

Abstract

Mimivirus is the prototype of the Mimiviridae family of giant dsDNA viruses. Little is known about the organization of the 1.2 Mb genome inside the membrane-limited nucleoid filling the ~0.5 µm icosahedral capsids. Cryo-electron microscopy, cryo-electron tomography and proteomics revealed that it is encased into a ~30 nm diameter helical protein shell surprisingly composed of two GMC-type oxidoreductases, which also form the glycosylated fibrils decorating the capsid. The genome is arranged in 5- or 6-start left-handed super-helices, with each DNA-strand lining the central channel. This luminal channel of the nucleoprotein fiber is wide enough to accommodate oxidative stress proteins and RNA polymerase subunits identified by proteomics. Such elegant supramolecular organization would represent a remarkable evolutionary strategy for packaging and protecting the genome, in a state ready for immediate transcription upon unwinding in the host cytoplasm. The parsimonious use of the same protein in two unrelated substructures of the virion is unexpected for a giant virus with thousand genes at its disposal.

DNA origami directed virus capsid polymorphism

“…1). 6HB fits inside the previously described, hollow tubes [18] (considering also the thickness of the CP layer [17]), and additionally, it closely mimicks the packaging of DNA observed in naturally occurring viral systems [29]. By optimizing the ratio between DNA origami and CPs, complexes with multiple protein layers were obtained and further characterized using single-particle cryo-electron microscopy (cryo-EM) reconstruction.…”

Evolution of giant pandoravirus from small icosahedral viruses revealed by CRISPR/Cas9

“…Expected PCR size: a+b: 837bp (wt), a+b 890bp (KO), a+c: (wt). A map of the construct used and a cartoon depicting the strategy of disruption is shown in Figure S1c-d 19 and pithovirus sibericum 18 ) and a nuclear version of Cas9 was used for the gene disruption of nuclear GVs (pandoravirus neocaledonia 7 and mollivirus Kamchatka 17 ). Guides off-target and on-target of the rbp1 gene for each virus were used.…”

“…The following viral strains have been used in this study: pandoravirus neocaledonia 7 , pandoravirus macleodensis 7 , pandoravirus kuranda (this study, ON887157), mollivirus kamchatka 17 , pithovirus sibericum 18 and mimivirus reunion 19 .…”

“…(g).Relative quantification of the number of viruses produced 24 hours post infection (hpi) of CRISPR/Cas9 expressing amoebas. A cytoplasmic version of Cas9 was used to attempt gene disruption of cytoplasmic GV (mimivirus reunion 19 and pithovirus sibericum 18 ) and a nuclear version of Cas9 was used for the gene disruption of nuclear GVs (pandoravirus neocaledonia 7 and mollivirus Kamchatka 17 ). Guides off-target and on-target of the rbp1 gene for each virus were used.…”

“…1f-g), while the cytosolic version of the protein (Fig. 1a) was unable to target the genome of pithovirus sibericum 18 or mimivirus reunion 19 (Fig. 1f-g).…”