TY - JOUR
T1 - Cells that express all five proteins of vesicular stomatitis virus from cloned cDNAs support replication, assembly, and budding of defective interfering particles
AU - Pattnaik, Asit K.
AU - Wertz, Gail W.
PY - 1991
Y1 - 1991
N2 - An alternative approach to structure-function analysis of vesicular stomatitis virus (VSV) gene products and their interactions with one another during each phase of the viral life cycle is described. We showed previously by using the vaccinia virus-T7 RNA polymerase expression system that when cells expressing the nucleocapsid protein (N), the phosphoprotein (NS), and the large polymerase protein (L) of VSV were superinfected with defective interfering (DI) particles, rapid and efficient replication and amplification of DI particle RNA occurred. Here, we demonstrate that all five VSV proteins can be expressed simultaneously when cells are cotransfected with plasmids containing the matrix protein (M) gene and the glycoprotein (G) gene of VSV in addition to plasmids containing the genes for the N, NS, and L proteins. When cells coexpressing all five VSV proteins were superinfected with DI particles, which because of their defectiveness are unable to express any viral proteins or to replicate, DI particle replication, assembly, and budding were observed and infectious DI particles were released into the culture fluids. Omission of either the M or G protein expression resulted in no DI particle budding. The vector-supported DI particles were similar in size and morphology to the authentic DI particles generated from cells coinfected with DI particles and helper VSV and their infectivity could be blocked by anti-VSV or anti-G antiserum. The successful replication, assembly, and budding of DI particles from cells expressing all five VSV proteins from cloned cDNAs provide a powerful approach for detailed structure-function analysis of the VSV gene products in each step of the replicative cycle of the virus.
AB - An alternative approach to structure-function analysis of vesicular stomatitis virus (VSV) gene products and their interactions with one another during each phase of the viral life cycle is described. We showed previously by using the vaccinia virus-T7 RNA polymerase expression system that when cells expressing the nucleocapsid protein (N), the phosphoprotein (NS), and the large polymerase protein (L) of VSV were superinfected with defective interfering (DI) particles, rapid and efficient replication and amplification of DI particle RNA occurred. Here, we demonstrate that all five VSV proteins can be expressed simultaneously when cells are cotransfected with plasmids containing the matrix protein (M) gene and the glycoprotein (G) gene of VSV in addition to plasmids containing the genes for the N, NS, and L proteins. When cells coexpressing all five VSV proteins were superinfected with DI particles, which because of their defectiveness are unable to express any viral proteins or to replicate, DI particle replication, assembly, and budding were observed and infectious DI particles were released into the culture fluids. Omission of either the M or G protein expression resulted in no DI particle budding. The vector-supported DI particles were similar in size and morphology to the authentic DI particles generated from cells coinfected with DI particles and helper VSV and their infectivity could be blocked by anti-VSV or anti-G antiserum. The successful replication, assembly, and budding of DI particles from cells expressing all five VSV proteins from cloned cDNAs provide a powerful approach for detailed structure-function analysis of the VSV gene products in each step of the replicative cycle of the virus.
KW - Coexpression
KW - Helper-independent budding
KW - Vaccinia virus-T7 RNA polymerase
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U2 - 10.1073/pnas.88.4.1379
DO - 10.1073/pnas.88.4.1379
M3 - Article
C2 - 1847519
AN - SCOPUS:0026031302
SN - 0027-8424
VL - 88
SP - 1379
EP - 1383
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 4
ER -