Impaired genome encapsidation restricts the in vitro propagation of human parvovirus B19

ABSTRACT Human parvovirus B19 (B19V) is a member of the genus Erythrovirus in the family Parvoviridae. In vitro, autonomous B19V replication is limited to human erythroid progenitor cells and in a small number of erythropoietin-dependent human megakaryoblastoid and erythroid leukemic cell lines. Here we report that the failure of B19V DNA replication in nonpermissive 293 cells can be overcome by adenovirus infection. More specifically, the replication of B19V DNA in the 293 cells and the production of infectious progeny virus were made possible by the presence of the adenovirus E2a, E4orf6, and VA RNA genes that emerged during the transfection of the pHelper plasmid. Using this replication system, we identified the terminal resolution site and the nonstructural protein 1 (NS1) binding site on the right terminal palindrome of the viral genome, which is composed of a minimal origin of replication spanning 67 nucleotides. Plasmids or DNA fragments containing an NS1 expression cassette and this minimal origin were able to replicate in both pHelper-transfected 293 cells and B19V-semipermissive UT7/Epo-S1 cells. Our results have important implications for our understanding of native B19V infection.

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T Helper Cell–Mediated In Vitro Responses of Recently and Remotely Infected Subjects to a Candidate Recombinant Vaccine for Human Parvovirus B19

The Journal of Infectious Diseases ◽

10.1086/318819 ◽

2001 ◽

Vol 183 (5) ◽

pp. 805-809 ◽

Cited By ~ 39

Author(s):

Rauli Franssila ◽

Kati Hokynar ◽

Klaus Hedman

Keyword(s):

Parvovirus B19 ◽

Recombinant Vaccine ◽

T Helper Cell ◽

Helper Cell ◽

Human Parvovirus B19 ◽

T Helper

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In vitro propagation of parvovirus B19 in primary foetal liver culture

Journal of General Virology ◽

10.1099/0022-1317-72-3-741 ◽

1991 ◽

Vol 72 (3) ◽

pp. 741-745 ◽

Cited By ~ 35

Author(s):

K. E. Brown ◽

J. Mori ◽

B. J. Cohen ◽

A. M. Field

Keyword(s):

In Vitro Propagation ◽

Parvovirus B19 ◽

Liver Culture ◽

Foetal Liver

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In Vitro Encapsulation of Heterologous dsDNA Into Human Parvovirus B19 Virus-Like Particles

Molecular Biotechnology ◽

10.1007/s12033-014-9823-4 ◽

2014 ◽

Vol 57 (4) ◽

pp. 309-317 ◽

Cited By ~ 3

Author(s):

Sandra Paola Sánchez-Rodríguez ◽

Joana Valeria Enrriquez-Avila ◽

Juan Miguel Soto-Fajardo ◽

Carolina Peña-Montes ◽

Ismael Bustos-Jaimes

Keyword(s):

Parvovirus B19 ◽

Human Parvovirus B19 ◽

Virus Like Particles

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The 11-Kilodalton Nonstructural Protein of Human Parvovirus B19 Facilitates Viral DNA Replication by Interacting with Grb2 through Its Proline-Rich Motifs

Journal of Virology ◽

10.1128/jvi.01464-18 ◽

2018 ◽

Vol 93 (1) ◽

Cited By ~ 7

Author(s):

Peng Xu ◽

Aaron Yun Chen ◽

Safder S. Ganaie ◽

Fang Cheng ◽

Weiran Shen ◽

...

Keyword(s):

Dna Replication ◽

Nonstructural Protein ◽

Parvovirus B19 ◽

Growth Factor Receptor ◽

Cellular Growth ◽

Human Parvovirus B19 ◽

Nonstructural Proteins ◽

Viral Dna ◽

Viral Dna Replication

ABSTRACTLytic infection of human parvovirus B19 (B19V) takes place exclusively in human erythroid progenitor cells of bone marrow and fetal liver, which disrupts erythropoiesis. During infection, B19V expresses three nonstructural proteins (NS1, 11-kDa, and 7.5-kDa) and two structural proteins (VP1 and VP2). While NS1 is essential for B19V DNA replication, 11-kDa enhances viral DNA replication significantly. In this study, we confirmed the enhancement role of 11-kDa in viral DNA replication and elucidated the underlying mechanism. We found that 11-kDa specially interacts with cellular growth factor receptor-bound protein 2 (Grb2) during virus infection andin vitro. We determined a high affinity interaction between 11-kDa and Grb2 that has an equilibrium dissociation constant (KD) value of 18.13 nM.In vitro, one proline-rich motif was sufficient for 11-kDa to sustain a strong interaction with Grb2. In consistence,in vivoduring infection, one proline-rich motif was enough for 11-kDa to significantly reduce phosphorylation of extracellular signal-regulated kinase (ERK). Mutations of all three proline-rich motifs of 11-kDa abolished its capability to reduce ERK activity and, accordingly, decreased viral DNA replication. Transduction of a lentiviral vector encoding a short hairpin RNA (shRNA) targetingGrb2decreased the expression of Grb2 as well as the level of ERK phosphorylation, which resulted in an increase of B19V replication. These results, in concert, indicate that the B19V 11-kDa protein interacts with cellular Grb2 to downregulate ERK activity, which upregulates viral DNA replication.IMPORTANCEHuman parvovirus B19 (B19V) infection causes hematological disorders and is the leading cause of nonimmunological fetal hydrops during pregnancy. During infection, B19V expresses two structural proteins, VP1 and VP2, and three nonstructural proteins, NS1, 11-kDa, and 7.5-kDa. While NS1 is essential, 11-kDa plays an enhancing role in viral DNA replication. Here, we elucidated a mechanism underlying 11-kDa protein-regulated B19V DNA replication. 11-kDa is tightly associated with cellular growth factor receptor-bound protein 2 (Grb2) during infection.In vitro, 11-kDa interacts with Grb2 with high affinity through three proline-rich motifs, of which at least one is indispensable for the regulation of viral DNA replication. 11-kDa and Grb2 interaction disrupts extracellular signal-regulated kinase (ERK) signaling, which mediates upregulation of B19V replication. Thus, our study reveals a novel mechanism of how a parvoviral small nonstructural protein regulates viral DNA replication by interacting with a host protein that is predominately expressed in the cytoplasm.

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Human parvovirus B19-induced epidemic acute red cell aplasia in patients with hereditary hemolytic anemia

Blood ◽

10.1182/blood.v67.5.1411.1411 ◽

1986 ◽

Vol 67 (5) ◽

pp. 1411-1417 ◽

Cited By ~ 124

Author(s):

UM Saarinen ◽

TL Chorba ◽

P Tattersall ◽

NS Young ◽

LJ Anderson ◽

...

Keyword(s):

Hemolytic Anemia ◽

Acute Phase ◽

Parvovirus B19 ◽

Beta Thalassemia ◽

Red Cell ◽

Human Parvovirus B19 ◽

High Concentration ◽

Red Cell Aplasia ◽

Abdominal Symptoms

Abstract From March to August 1984, 26 patients with hereditary hemolytic anemia in northeastern Ohio developed acute, profound red cell aplasia. The patients included 14 males and 12 females 2 to 23 years old, with sickle cell anemia (20 cases), hemoglobin SC-disease (4 cases), sickle- beta-thalassemia (1 case), or hereditary spherocytosis (1 case). All had an acute onset of severe reticulocytopenia and anemia and prodromal symptoms of illness including fever, abdominal symptoms, headache, and arthralgias. Twenty-two received transfusions. Reticulocytosis occurred spontaneously within 2 to 14 days of presentation. In five acute-phase sera, 10(8) to 10(12) viral particles/mL were detected by electron microscopy. Human parvovirus B19 DNA was demonstrated in high concentration by hybridization in the same five acute-phase sera and in low concentration in sera of eight additional patients. The five highly viremic sera inhibited erythroid colony formation in vitro. B19- specific IgM was detected in sera of 24/26 patients, and B19-specific IgG in 21 of 22 patients tested. Our results indicate that human parvovirus B19 was the etiologic agent in this large epidemic of life- threatening acute red cell aplasia in patients with hereditary hemolytic anemia.

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