Importin α/β-dependent nuclear transport of human parvovirus B19 nonstructural protein 1 is essential for viral replication

• Published in: Antiviral research Vol. 213; p. 105588
• Main Authors: Alvisi, Gualtiero, Manaresi, Elisabetta, Cross, Emily M., Hoad, Mikayla, Akbari, Nasim, Pavan, Silvia, Ariawan, Daryl, Bua, Gloria, Petersen, Gayle F., Forwood, Jade, Gallinella, Giorgio
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.05.2023
• Subjects: Active Transport, Cell Nucleus; alpha Karyopherins - genetics; alpha Karyopherins - metabolism; Antiviral; B19V; beta Karyopherins - metabolism; cNLS; Humans; Importins; Ivermectin; Nuclear transport; Parvovirus B19, Human - genetics; Viral Nonstructural Proteins - genetics; Viral Nonstructural Proteins - metabolism; Virus Replication; Antiviral; B19V; cNLS; Importins; Nuclear transport; Ivermectin
• ISSN: 0166-3542; 1872-9096
• DOI: 10.1016/j.antiviral.2023.105588

Human parvovirus B19 (B19V) is a major human pathogen causing a variety of diseases, characterized by a selective tropism to human progenitor cells in bone marrow. In similar fashion to all Parvoviridae members, the B19V ssDNA genome is replicated within the nucleus of infected cells through a process which involves both cellular and viral proteins. Among the latter, a crucial role is played by non-structural protein (NS)1, a multifunctional protein involved in genome replication and transcription, as well as modulation of host gene expression and function. Despite the localization of NS1 within the host cell nucleus during infection, little is known regarding the mechanism of its nuclear transport pathway. In this study we undertake structural, biophysical, and cellular approaches to characterize this process. Quantitative confocal laser scanning microscopy (CLSM), gel mobility shift, fluorescence polarization and crystallographic analysis identified a short sequence of amino acids (GACHAKKPRIT-182) as the classical nuclear localization signal (cNLS) responsible for nuclear import, mediated in an energy and importin (IMP) α/β-dependent fashion. Structure-guided mutagenesis of key residue K177 strongly impaired IMPα binding, nuclear import, and viral gene expression in a minigenome system. Further, treatment with ivermectin, an antiparasitic drug interfering with the IMPα/β dependent nuclear import pathway, inhibited NS1 nuclear accumulation and viral replication in infected UT7/Epo-S1 cells. Thus, NS1 nuclear transport is a potential target of therapeutic intervention against B19V induced disease. •We characterized the nuclear import process of B19V NS1.•Import is dependent on the host cell IMPα/β heterodimer and a classical NLS on NS1.•Structural analysis of IMPα2-NS1 NLS complex suggested a key role for residue K177.•Inhibition of NS1 nuclear import impaired B19V replication in cell culture.