Disassociation of the SV40 Genome from Capsid Proteins Prior to Nuclear Entry

ABSTRACT This report describes the distribution of conventional nuclear localization sequences (NLS) and of a beta-stranded so-called nuclear localization motif (NLM) in the two proteins (VP1, 82 kDa; VP2, 63 kDa) forming the T=1 icosahedral capsid of the parvovirus minute virus of mice (MVM) and their functions in viral biogenesis and the onset of infection. The approximately 10 VP1 molecules assembled in the MVM particle harbor in its 142-amino-acid (aa) N-terminal-specific region four clusters of basic amino acids, here called BC1 (aa 6 to 10), BC2 (aa 87 to 90), BC3 (aa 109 to 115), and BC4 (aa 126 to 130), that fit consensus NLS and an NLM placed toward the opposite end of the polypeptide (aa 670 to 680) found to be necessary for VP2 nuclear uptake. Deletions and site-directed mutations constructed in an infectious MVM plasmid showed that BC1, BC2, and NLM are cooperative nuclear transport sequences in singly expressed VP1 subunits and that they conferred nuclear targeting competence on the VP1/VP2 oligomers arising in normal infection, while BC3 and BC4 did not display nuclear transport activity. Notably, VP1 proteins mutated at BC1 and -2, and particularly with BC1 to -4 sequences deleted, induced nuclear and cytoplasmic foci of colocalizing conjugated ubiquitin that could be rescued from the ubiquitin-proteasome degradation pathway by the coexpression of VP2 and NS2 isoforms. These results suggest a role for VP2 in viral morphogenesis by assisting cytoplasmic folding of VP1/VP2 subviral complexes, which is further supported by the capacity of NLM-bearing transport-competent VP2 subunits to recruit VP1 into the nuclear capsid assembly pathway regardless of the BC composition. Instead, all four BC sequences, which are located in the interior of the capsid, were absolutely required by the incoming infectious MVM particle for the onset of infection, suggesting either an important conformational change or a disassembly of the coat for nuclear entry of a VP1-associated viral genome. Therefore, the evolutionarily conserved BC sequences and NLM domains provide complementary nuclear transport functions to distinct supramolecular complexes of capsid proteins during the autonomous parvovirus life cycle.

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Minor Capsid Proteins of Simian Virus 40 Are Dispensable for Nucleocapsid Assembly and Cell Entry but Are Required for Nuclear Entry of the Viral Genome

Journal of Virology ◽

10.1128/jvi.02664-06 ◽

2007 ◽

Vol 81 (8) ◽

pp. 3778-3785 ◽

Cited By ~ 33

Author(s):

Akira Nakanishi ◽

Noriko Itoh ◽

Peggy P. Li ◽

Hiroshi Handa ◽

Robert C. Liddington ◽

...

Keyword(s):

Deletion Mutant ◽

Cell Attachment ◽

Simian Virus 40 ◽

Simian Virus ◽

T Antigen ◽

Capsid Proteins ◽

Nuclear Entry ◽

Viral Dna ◽

Viral Dnas ◽

Stable Particles

ABSTRACT We investigated the roles of simian virus 40 capsid proteins in the viral life cycle by analyzing point mutants in Vp1 and Vp2/3, as well as a deletion mutant lacking the Vp2/3 coding sequence. The Vp1 mutants (V243E and L245E) and the Vp2/3 mutants (F157E-I158E and P164R-G165E-G166R) were previously shown to be defective in Vp1-Vp2/3 interaction and to be noninfectious or poorly infectious, respectively. Here, we show that all these point mutants form stable particles following DNA transfection into cells. The Vp2/3-mutant particles contained very low levels of Vp2/3, whereas the Vp1 mutant particles contained no detectable Vp2/3. As expected, the deletion mutant also formed particles that were noninfectious. We further characterized the two Vp1 point mutants and the deletion mutant. All three mutant particles comprised Vp1 and histone-associated viral DNA, and all were able to enter cells. However, the mutant complexes failed to associate with host importins (owing to the loss of the Vp2/3 nuclear localization signal), and the mutant viral DNAs prematurely dissociated from the Vp1s, suggesting that the nucleocapsids did not enter the nucleus. Consistently, all three mutant particles failed to express large T antigen. Together, our results demonstrate unequivocally that Vp2/3 is dispensable for the formation of nucleocapsids. Further, the nucleocapsids' ability to enter cells implies that Vp1 contains the major determinants for cell attachment and entry. We propose that the major role of Vp2/3 in infectivity is to mediate the nuclear entry of viral DNA.

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Role of a nuclear localization signal on the minor capsid Proteins VP2 and VP3 in BKPyV nuclear entry

Virology ◽

10.1016/j.virol.2014.10.013 ◽

2015 ◽

Vol 474 ◽

pp. 110-116 ◽

Cited By ~ 32

Author(s):

Shauna M. Bennett ◽

Linbo Zhao ◽

Catherine Bosard ◽

Michael J. Imperiale

Keyword(s):

Nuclear Localization ◽

Nuclear Localization Signal ◽

Capsid Proteins ◽

Nuclear Entry ◽

Localization Signal

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The concentration of Ca2+ that solubilizes outer capsid proteins from rotavirus particles is dependent on the strain.

Journal of Virology ◽

10.1128/jvi.70.8.4877-4883.1996 ◽

1996 ◽

Vol 70 (8) ◽

pp. 4877-4883 ◽

Cited By ~ 36

Author(s):

M C Ruiz ◽

A Charpilienne ◽

F Liprandi ◽

R Gajardo ◽

F Michelangeli ◽

...

Keyword(s):

Capsid Proteins ◽

Outer Capsid

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Asymmetric reconstruction of mammalian reovirus reveals interactions among RNA, transcriptional factor µ2 and capsid proteins

Nature Communications ◽

10.1038/s41467-021-24455-4 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Muchen Pan ◽

Ana L. Alvarez-Cabrera ◽

Joon S. Kang ◽

Lihua Wang ◽

Chunhai Fan ◽

...

Keyword(s):

Rna Polymerase ◽

Molecular Interactions ◽

Messenger Rna ◽

Rna Binding ◽

Capsid Proteins ◽

Rna Transcription ◽

Infectious Particle ◽

Virion Assembly ◽

Rna Capping

AbstractMammalian reovirus (MRV) is the prototypical member of genus Orthoreovirus of family Reoviridae. However, lacking high-resolution structures of its RNA polymerase cofactor μ2 and infectious particle, limits understanding of molecular interactions among proteins and RNA, and their contributions to virion assembly and RNA transcription. Here, we report the 3.3 Å-resolution asymmetric reconstruction of transcribing MRV and in situ atomic models of its capsid proteins, the asymmetrically attached RNA-dependent RNA polymerase (RdRp) λ3, and RdRp-bound nucleoside triphosphatase μ2 with a unique RNA-binding domain. We reveal molecular interactions among virion proteins and genomic and messenger RNA. Polymerase complexes in three Spinoreovirinae subfamily members are organized with different pseudo-D3d symmetries to engage their highly diversified genomes. The above interactions and those between symmetry-mismatched receptor-binding σ1 trimers and RNA-capping λ2 pentamers balance competing needs of capsid assembly, external protein removal, and allosteric triggering of endogenous RNA transcription, before, during and after infection, respectively.

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The Role of Capsid in HIV-1 Nuclear Entry

Viruses ◽

10.3390/v13081425 ◽

2021 ◽

Vol 13 (8) ◽

pp. 1425

Author(s):

Anabel Guedán ◽

Eve R. Caroe ◽

Genevieve C. R. Barr ◽

Kate N. Bishop

Keyword(s):

Nuclear Pore ◽

Outer Face ◽

Nuclear Entry ◽

Nuclear Compartment ◽

Critical Step ◽

Technological Advances ◽

Dividing Cells ◽

Hiv 1 ◽

Gain Access

HIV-1 can infect non-dividing cells. The nuclear envelope therefore represents a barrier that HIV-1 must traverse in order to gain access to the host cell chromatin for integration. Hence, nuclear entry is a critical step in the early stages of HIV-1 replication. Following membrane fusion, the viral capsid (CA) lattice, which forms the outer face of the retroviral core, makes numerous interactions with cellular proteins that orchestrate the progress of HIV-1 through the replication cycle. The ability of CA to interact with nuclear pore proteins and other host factors around the nuclear pore determines whether nuclear entry occurs. Uncoating, the process by which the CA lattice opens and/or disassembles, is another critical step that must occur prior to integration. Both early and delayed uncoating have detrimental effects on viral infectivity. How uncoating relates to nuclear entry is currently hotly debated. Recent technological advances have led to intense discussions about the timing, location, and requirements for uncoating and have prompted the field to consider alternative uncoating scenarios that presently focus on uncoating at the nuclear pore and within the nuclear compartment. This review describes recent advances in the study of HIV-1 nuclear entry, outlines the interactions of the retroviral CA protein, and discusses the challenges of investigating HIV-1 uncoating.

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Rotaviruses and Rotavirus Vaccines

Pathogens ◽

10.3390/pathogens10080959 ◽

2021 ◽

Vol 10 (8) ◽

pp. 959

Author(s):

Celeste M. Donato ◽

Julie E. Bines

Keyword(s):

Capsid Proteins ◽

Virus Family ◽

Rotavirus Vaccines ◽

Group A Rotaviruses ◽

Group A ◽

Outer Capsid

Group A rotaviruses belong to the Reoviridae virus family and are classified into G and P genotypes based on the outer capsid proteins VP7 and VP4, respectively [...]

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Targeting novel LSD1-dependent ACE2 demethylation domains inhibits SARS-CoV-2 replication

Cell Discovery ◽

10.1038/s41421-021-00279-w ◽

2021 ◽

Vol 7 (1) ◽

Author(s):

Wen Juan Tu ◽

Robert D. McCuaig ◽

Michelle Melino ◽

Daniel J. Rawle ◽

Thuy T. Le ◽

...

Keyword(s):

Viral Replication ◽

Treatment Options ◽

Critical Role ◽

Post Exposure Prophylaxis ◽

Nuclear Entry ◽

Importin Α ◽

Active Transcription ◽

Nuclear Shuttling ◽

Asymptomatic Phase ◽

Exposure Prophylaxis

AbstractTreatment options for COVID-19 remain limited, especially during the early or asymptomatic phase. Here, we report a novel SARS-CoV-2 viral replication mechanism mediated by interactions between ACE2 and the epigenetic eraser enzyme LSD1, and its interplay with the nuclear shuttling importin pathway. Recent studies have shown a critical role for the importin pathway in SARS-CoV-2 infection, and many RNA viruses hijack this axis to re-direct host cell transcription. LSD1 colocalized with ACE2 at the cell surface to maintain demethylated SARS-CoV-2 spike receptor-binding domain lysine 31 to promote virus–ACE2 interactions. Two newly developed peptide inhibitors competitively inhibited virus–ACE2 interactions, and demethylase access to significantly inhibit viral replication. Similar to some other predominantly plasma membrane proteins, ACE2 had a novel nuclear function: its cytoplasmic domain harbors a nuclear shuttling domain, which when demethylated by LSD1 promoted importin-α-dependent nuclear ACE2 entry following infection to regulate active transcription. A novel, cell permeable ACE2 peptide inhibitor prevented ACE2 nuclear entry, significantly inhibiting viral replication in SARS-CoV-2-infected cell lines, outperforming other LSD1 inhibitors. These data raise the prospect of post-exposure prophylaxis for SARS-CoV-2, either through repurposed LSD1 inhibitors or new, nuclear-specific ACE2 inhibitors.

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