Abstract
It is unknown whether the mammalian cell cycle could impact the assembly of viruses maturing in the nucleus. We addressed this question using MVM, a reference member of the icosahedral ssDNA nuclear parvoviruses, which requires cell proliferation to infect by mechanisms partly understood. Constitutively expressed MVM capsid subunits (VPs) accumulated in the cytoplasm of mouse and human fibroblasts synchronized at G0, G1, and G1/S transition. Upon arrest release, VPs translocated to the nucleus as cells entered S phase, at efficiencies relying on cell origin and arrest method, and immediately assembled into capsids. In synchronously infected cells, the consecutive virus life cycle steps (gene expression, proteins nuclear translocation, capsid assembly, genome replication and encapsidation) proceeded tightly coupled to cell cycle progression from G0/G1 through S into G2 phase. However, a DNA synthesis stress caused by thymidine irreversibly disrupted virus life cycle, as VPs became increasingly retained in the cytoplasm hours post-stress, forming empty capsids in mouse fibroblasts, thereby impairing encapsidation of the nuclear viral DNA replicative intermediates. Synchronously infected cells subjected to density-arrest signals while traversing early S phase also blocked VPs transport, resulting in a similar misplaced cytoplasmic capsid assembly in mouse fibroblasts. In contrast, thymidine and density arrest signals deregulating virus assembly neither perturbed nuclear translocation of the NS1 protein nor viral genome replication occurring under S/G2 cycle arrest. An underlying mechanism of cell cycle control was identified in the nuclear translocation of phosphorylated VPs trimeric assembly intermediates, which accessed a non-conserved route distinct from the importin α2/β1 and transportin pathways. The exquisite cell cycle-dependence of parvovirus nuclear capsid assembly conforms a novel paradigm of time and functional coupling between cellular and virus life cycles. This junction may determine the characteristic parvovirus tropism for proliferative and cancer cells, and its disturbance could critically contribute to persistence in host tissues.
Highlights
Viruses infecting eukaryotes encounter a dynamic regulatory network of proteins expression, posttranslational modifications and signaling, ensuring fine-tuned control of the cell cycle progression and checkpoints [1,2,3]
We found that the nuclear translocation of parvovirus Minute Virus of Mice (MVM) capsid subunits (VPs) was highly dependent on physiological cell cycle regulations in mammalian fibroblasts, including: quiescence, progression through G1/S boundary, DNA synthesis, and cell to cell contacts
To address this fundamental issue in a mammalian system, we investigated the effects of cell cycle regulations on the nuclear assembly of the parvovirus Minute Virus of Mice (MVM) infecting mouse and human fibroblasts
Summary
Viruses infecting eukaryotes encounter a dynamic regulatory network of proteins expression, posttranslational modifications and signaling, ensuring fine-tuned control of the cell cycle progression and checkpoints [1,2,3] These processes include the still poorly understood cell cycledependent vast macromolecular traffic across the central aqueous channel of the nuclear pore complex (NPC; [4,5]), which requires soluble receptors (karyopherins) for the recognition of cargos carrying nuclear transport signals [6,7]. All these cell cycle regulations affect the multiplication of many eukaryotic viruses, which must either adapt their life cycles to these factors networks, or perturb them for their benefit. In spite of the important body of knowledge provided by these valuable studies, little is known on how cell cycle regulations and their perturbations are connected with the assembly and maturation of viruses
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