A vaccinia virus lacking A10L: viral core proteins accumulate on structures derived from the endoplasmic reticulum.
The assembly of the intracellular mature virus (IMV) of vaccinia virus (VV), the prototype member of the poxviridae, is poorly understood and controversial. We have previously proposed that the IMV is composed of a continuous double-membraned cisterna derived from the smooth ER, whereby the genome-containing core is enwrapped by a part of this cisterna. In the present study we characterize a mutant virus in which the synthesis of the major core protein A10L can be conditionally expressed. Without A10L, IMVs are not made; immature viruses (IVs) and regularly stacked membrane structures that contain viral DNA, accumulate instead. By immunolabelling of thawed cryo-sections these stacks contain most of the viral core proteins and low levels of viral membrane proteins. Importantly, the stacked membranes could be labelled with antibodies to an ER marker protein, implying that they are derived from this cellular compartment. By electron tomography (ET) on semi-thin cryo-sections we show that the membranes of the stacks are continuous with the membranes of the IVs. Direct continuities with ER cisternae, to which the stacks are tightly apposed, were, however, not unequivocally seen. Finally, ET revealed how the IV membranes separated to become two-membrane profiles. Taken together, this study shows that VV core proteins and the viral DNA can coassemble onto ER-derived membranes that are continuous with the membranes of the IVs.
Centro Nacional de Biotecnología, Campus Universidad Autónoma, 28049 Madrid, Spain.
Dolores
D
Rodriguez
Montserrat
M
Bárcena
Wiebke
W
Möbius
Sibylle
S
Schleich
Mariano
M
Esteban
Willie J C
WJ
Geerts
Abraham J
AJ
Koster
Gareth
G
Griffiths
Jacomine Krijnse
JK
Locker
D000818:Animals; D002460:Cell Line; D004279:DNA, Viral / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism* / Q000648:ultrastructure*; D006801:Humans; D008854:Microscopy, Electron; D009154:Mutation; D014054:Tomography; D014616:Vaccinia virus / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D014758:Viral Core Proteins / Q000235:genetics / Q000378:metabolism*; D014771:Virion / Q000378:metabolism*; D019065:Virus Assembly
D004279:DNA, Viral; C069852:P4a protein, Vaccinia virus; D014758:Viral Core Proteins