NSP9 of SARS-CoV-2 attenuates nuclear transport by hampering nucleoporin 62 dynamics and functions in host cells

ABSTRACTRust fungi are plant pathogens that secrete an arsenal of effector proteins interfering with plant functions and promoting parasitic infection. Effectors are often species-specific, evolve rapidly, and display low sequence similarities with known proteins or domains. How rust fungal effectors function in host cells remains elusive, and biochemical and structural approaches have been scarcely used to tackle this question. In this study, we used a strategy based on recombinant protein production in Escherichia coli to study eleven candidate effectors of the leaf rust fungus Melampsora larici-populina. We successfully purified and solved the three-dimensional structure of two proteins, MLP124266 and MLP124017, using NMR spectroscopy. Although both proteins show no sequence similarity with known proteins, they exhibit structural similarities to knottin and nuclear transport factor 2-like proteins, respectively. Altogether, our findings show that sequence-unrelated effectors can adopt folds similar to known proteins, and encourage the use of biochemical and structural approaches to functionally characterize rust effector candidates.

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Granzyme K inhibits replication of influenza virus through cleaving the nuclear transport complex importin α1/β dimer of infected host cells

Cell Death and Differentiation ◽

10.1038/cdd.2011.178 ◽

2011 ◽

Vol 19 (5) ◽

pp. 882-890 ◽

Cited By ~ 17

Author(s):

C Zhong ◽

C Li ◽

X Wang ◽

T Toyoda ◽

G Gao ◽

...

Keyword(s):

Influenza Virus ◽

Nuclear Transport ◽

Host Cells ◽

Infected Host ◽

Transport Complex

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Effect of Ivermectin and Atorvastatin on Nuclear Localization of Importin Alpha and Drug Target Expression Profiling in Host Cells from Nasopharyngeal Swabs of SARS-CoV-2- Positive Patients

Viruses ◽

10.3390/v13102084 ◽

2021 ◽

Vol 13 (10) ◽

pp. 2084

Author(s):

Valeria Inés Segatori ◽

Juan Garona ◽

Lorena Grisel Caligiuri ◽

Juan Bizzotto ◽

Rosario Lavignolle ◽

...

Keyword(s):

Gene Expression ◽

Nuclear Localization ◽

Rho Gtpases ◽

Nuclear Transport ◽

Host Cells ◽

Poly I:C ◽

Nuclear Accumulation ◽

Synthetic Analog ◽

Importin Α

Nuclear transport and vesicle trafficking are key cellular functions involved in the pathogenesis of RNA viruses. Among other pleiotropic effects on virus-infected host cells, ivermectin (IVM) inhibits nuclear transport mechanisms mediated by importins and atorvastatin (ATV) affects actin cytoskeleton-dependent trafficking controlled by Rho GTPases signaling. In this work, we first analyzed the response to infection in nasopharyngeal swabs from SARS-CoV-2-positive and -negative patients by assessing the gene expression of the respective host cell drug targets importins and Rho GTPases. COVID-19 patients showed alterations in KPNA3, KPNA5, KPNA7, KPNB1, RHOA, and CDC42 expression compared with non-COVID-19 patients. An in vitro model of infection with Poly(I:C), a synthetic analog of viral double-stranded RNA, triggered NF-κB activation, an effect that was halted by IVM and ATV treatment. Importin and Rho GTPases gene expression was also impaired by these drugs. Furthermore, through confocal microscopy, we analyzed the effects of IVM and ATV on nuclear to cytoplasmic importin α distribution, alone or in combination. Results showed a significant inhibition of importin α nuclear accumulation under IVM and ATV treatments. These findings confirm transcriptional alterations in importins and Rho GTPases upon SARS-CoV-2 infection and point to IVM and ATV as valid drugs to impair nuclear localization of importin α when used at clinically-relevant concentrations.

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Maize Streak Virus Coat Protein Is Karyophyllic and Facilitates Nuclear Transport of Viral DNA

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi.1999.12.10.894 ◽

1999 ◽

Vol 12 (10) ◽

pp. 894-900 ◽

Cited By ~ 50

Author(s):

H. Liu ◽

M. I. Boulton ◽

C. L. Thomas ◽

D. A. M. Prior ◽

K. J. Oparka ◽

...

Keyword(s):

Coat Protein ◽

Nuclear Transport ◽

Transient Gene Expression ◽

Maize Streak Virus ◽

Host Cells ◽

Immunofluorescent Staining ◽

Vector System ◽

Viral Gene ◽

Streak Virus ◽

Viral Dna

Transport of maize streak virus (MSV) DNA into the nucleus of host cells is essential for virus replication and the presence of virus particles in the nuclei of infected cells implies that coat protein (CP) must enter the nucleus. To see if CP is imported into the nucleus in the absence of other viral gene products, the MSV CP gene was expressed in insect cells with a baculovirus vector system, and also in tobacco protoplasts with a cauliflower mosaic virus (CaMV) 35S promoter-driven transient gene expression vector. Immunofluorescent staining showed that the CP accumulated in the nuclei of both insect and tobacco cells. Mutagenesis of a potential nuclear localization signal in the CP resulted in cytoplasmic accumulation of the mutant protein. We have shown previously that the CP binds to single-stranded (ss) and double-stranded (ds) viral DNA. To investigate if CP might also be involved in viral DNA nuclear transport, Escherichia coli-expressed CP, together with TOTO-1-labeled viral ss or ds DNA, was microinjected into maize and tobacco epidermal cells. Both ss and ds DNA moved into the nucleus when co-injected with the CP but not with E. coli proteins alone. These results suggest that, in addition to entering the nucleus where it is required for encapsidation of the viral ss DNA, the MSV CP facilitates the rapid transport of viral (ss or ds) DNA into the nucleus.

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High Voltage Electron Microscopy of Viral Inclusion Bodies

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s1431927600002117 ◽

1980 ◽

Vol 38 ◽

pp. 486-487 ◽

Cited By ~ 1

Author(s):

H.M. Mazzone ◽

W.F. Engler ◽

G. Wray ◽

A. Szirmae ◽

J. Conroy ◽

...

Keyword(s):

New York ◽

Environmental Protection Agency ◽

Inclusion Bodies ◽

Host Cells ◽

High Voltage Electron Microscopy ◽

Voltage Electron ◽

Viral Particles ◽

Pest Insects ◽

Viral Inclusion ◽

Insect Gut

Viral inclusion bodies isolated from infected pest insects are being evaluated by the U.S. Dept. of Agriculture as biological insecticides against their hosts. Our research on these inclusion bodies constitutes part of an effort to support their approval by the Environmental Protection Agency as insect control agents. The inclusion bodies in this study are polyhedral in shape and contain rod-shaped viral particles. When ingested by pest insects, the inclusion bodies are broken down in the insect gut and release the viral particles which infect and multiply in the nuclei of host cells. These viruses are termed nucleopolyhedrosis viruses (NPV) and are representatives of the baculoviruses (Wildy, P. 1971 IN J.L. Melnick, ed., Monographs in Virology, vol. 5, S.Karger, New York).

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