Central role of a serine phosphorylation site within duck hepatitis B virus core protein for capsid trafficking and genome release.
Viral nucleocapsids compartmentalize and protect viral genomes during assembly while they mediate targeted genome release during viral infection. This dual role of the capsid in the viral life cycle must be tightly regulated to ensure efficient virus spread. Here, we used the duck hepatitis B virus (DHBV) infection model to analyze the effects of capsid phosphorylation and hydrogen bond formation. The potential key phosphorylation site at serine 245 within the core protein, the building block of DHBV capsids, was substituted by alanine (S245A), aspartic acid (S245D) and asparagine (S245N), respectively. Mutant capsids were analyzed for replication competence, stability, nuclear transport, and infectivity. All mutants formed DHBV DNA-containing nucleocapsids. Wild-type and S245N but not S245A and S245D fully protected capsid-associated mature viral DNA from nuclease action. A negative ionic charge as contributed by phosphorylated serine or aspartic acid-supported nuclear localization of the viral capsid and generation of nuclear superhelical DNA. Finally, wild-type and S245D but not S245N virions were infectious in primary duck hepatocytes. These results suggest that hydrogen bonds formed by non-phosphorylated serine 245 stabilize the quarterny structure of DHBV nucleocapsids during viral assembly, while serine phosphorylation plays an important role in nuclear targeting and DNA release from capsids during viral infection.
Department of Medicine II, University Hospital, 79106 Freiburg, Germany.
Josef
J
Kock
Michael
M
Kann
Gerhard
G
Putz
Hubert E
HE
Blum
Fritz
F
Von Weizsacker
D000409:Alanine / Q000737:chemistry; D000818:Animals; D001216:Asparagine / Q000737:chemistry; D001224:Aspartic Acid / Q000737:chemistry; D001665:Binding Sites; D015153:Blotting, Western; D002213:Capsid / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D002645:Chickens; D004270:DNA, Circular; D004279:DNA, Viral / Q000378:metabolism; D015611:Hepatitis B Virus, Duck / Q000378:metabolism*; D006860:Hydrogen Bonding; D018613:Microscopy, Confocal; D009154:Mutation; D010766:Phosphorylation; D011233:Precipitin Tests; D012694:Serine / Q000737:chemistry* / Q000378:metabolism; D014162:Transfection
D004270:DNA, Circular; D004279:DNA, Viral; D001224:Aspartic Acid; D012694:Serine; D001216:Asparagine; D000409:Alanine