Different architectures in the assembly of infectious bursal disease virus capsid proteins expressed in insect cells.
Infectious bursal disease virus (IBDV) capsid is formed by the processing of a large polyprotein and subsequent assembly of VPX/VP2 and VP3. To learn more about the processing of the polyprotein and factors affecting the correct assembly of the viral capsid in vitro, different constructs were made using two baculovirus transfer vectors, pFastBac and pAcYM1. Surprisingly, the expression of the capsid proteins gave rise to different types of particles in each system, as observed by electron microscopy and immunofluorescence. FastBac expression led to the production of only rigid tubular structures, similar to those described as type I in viral infection. Western blot analysis revealed that these rigid tubules are formed exclusively by VPX. These tubules revealed a hexagonal arrangement of units that are trimer clustered, similar to those observed in IBDV virions. In contrast, pAcYM1 expression led to the assembly of virus-like particles (VLPs), flexible tubules, and intermediate assembly products formed by icosahedral caps elongated in tubes, suggesting an aberrant morphogenesis. Processing of VPX to VP2 seems to be a crucial requirement for the proper morphogenesis and assembly of IBDV particles. After immunoelectron microscopy, VPX/VP2 was detected on the surface of tubules and VLPs. We also demonstrated that VP3 is found only on the inner surfaces of VLPs and caps of the tubular structures. In summary, assembly of VLPs requires the internal scaffolding of VP3, which seems to induce the closing of the tubular architecture into VLPs and, thereafter, the subsequent processing of VPX to VP2.
Martinez-Torrecuadrada
J L
JL
INGENASA, C/Hnos, Garcia Noblejas 41, 4 degrees, Madrid, 28037, Spain.
Castón
J R
JR
Castro
M
M
Carrascosa
J L
JL
Rodriguez
J F
JF
Casal
J I
JI
D000818:Animals; D016367:Baculoviridae; D002213:Capsid / Q000235:genetics / Q000302:isolation & purification / Q000648:ultrastructure*; D036022:Capsid Proteins; D002460:Cell Line; D005822:Genetic Vectors; D000882:Haplorhini; D007243:Infectious bursal disease virus / Q000235:genetics* / Q000648:ultrastructure; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000648:ultrastructure; D018411:Spodoptera; D014162:Transfection; D054334:Viral Regulatory and Accessory Proteins / Q000235:genetics / Q000302:isolation & purification / Q000648:ultrastructure
D036022:Capsid Proteins; D011994:Recombinant Proteins; D054334:Viral Regulatory and Accessory Proteins