The Large Nonstructural Protein (NS1) of Human Bocavirus 1 Directly Interacts with Ku70, Which Plays an Important Role in Virus Replication in Human Airway Epithelia

• Published in: Journal of virology Vol. 96; no. 4; p. e0184021
• Main Authors: Shao, Liting, Ning, Kang, Wang, Jianke, Cheng, Fang, Wang, Shengqi, Qiu, Jianming
• Format: Journal Article
• Language: English
• Published: United States American Society for Microbiology 23.02.2022
• Subjects: DNA Damage; DNA Replication; DNA, Viral - biosynthesis; Genome, Viral; HEK293 Cells; Human bocavirus - metabolism; Human bocavirus - physiology; Humans; Ku Autoantigen - genetics; Ku Autoantigen - metabolism; Protein Binding; Protein Domains; Respiratory Mucosa - metabolism; Respiratory Mucosa - virology; Viral Nonstructural Proteins - genetics; Viral Nonstructural Proteins - metabolism; Viral Replication Compartments - metabolism; Virology; Virus Replication; Virus-Cell Interactions; human airway epithelia; DNA replication; parvovirus; DNA damage
• ISSN: 0022-538X; 1098-5514
• DOI: 10.1128/JVI.01840-21

Human bocavirus 1 (HBoV1), an autonomous human parvovirus, causes acute respiratory tract infections in young children. HBoV1 infects well-differentiated (polarized) human airway epithelium cultured at an air-liquid interface (HAE-ALI). HBoV1 expresses a large nonstructural protein, NS1, that is essential for viral DNA replication. HBoV1 infection of polarized human airway epithelial cells induces a DNA damage response (DDR) that is critical to viral DNA replication involving DNA repair with error-free Y-family DNA polymerases. HBoV1 NS1 or the isoform NS1-70 induces a DDR. In this study, using the second-generation proximity-dependent biotin identification (BioID2) approach, we identified that Ku70 is associated with the NS1-BioID2 pulldown complex through a direct interaction with NS1. Biolayer interferometry (BLI) assay determined a high binding affinity of NS1 with Ku70, which has an equilibrium dissociation constant ( ) value of 0.16 μM and processes the strongest interaction at the C-terminal domain. The association of Ku70 with NS1 was also revealed during HBoV1 infection of HAE-ALI. Knockdown of Ku70 and overexpression of the C-terminal domain of Ku70 significantly decreased HBoV1 replication in HAE-ALI. Thus, our study provides, for the first time, a direct interaction of parvovirus large nonstructural protein NS1 with Ku70. Parvovirus infection induces a DNA damage response (DDR) that plays a pivotal role in viral DNA replication. The DDR includes activation of ATM (ataxia telangiectasia mutated), ATR (ATM- and RAD3-related), and DNA-PKcs (DNA-dependent protein kinase catalytic subunit). The large nonstructural protein (NS1) often plays a role in the induction of DDR; however, how the DDR is induced during parvovirus infection or simply by the NS1 is not well studied. Activation of DNA-PKcs has been shown as one of the key DDR pathways in DNA replication of HBoV1. We identified that HBoV1 NS1 directly interacts with Ku70, but not Ku80, of the Ku70/Ku80 heterodimer at high affinity. This interaction is also important for HBoV1 replication in HAE-ALI. We propose that the interaction of NS1 with Ku70 recruits the Ku70/Ku80 complex to the viral DNA replication center, which activates DNA-PKcs and facilitates viral DNA replication.