Impaired hyperphosphorylation of rotavirus NSP5 in cells depleted of casein kinase 1alpha is associated with the formation of viroplasms with altered morphology and a moderate decrease in virus replication.
The rotavirus (RV) non-structural protein 5, NSP5, is encoded by the smallest of the 11 genomic segments and localizes in 'viroplasms', cytoplasmic inclusion bodies in which viral RNA replication and packaging take place. NSP5 is essential for the replicative cycle of the virus because, in its absence, viroplasms are not formed and viral RNA replication and transcription do not occur. NSP5 is produced early in infection and undergoes a complex hyperphosphorylation process, leading to the formation of proteins differing in electrophoretic mobility. The role of hyperphosphorylation of NSP5 in the replicative cycle of rotavirus is unknown. Previous in vitro studies have suggested that the cellular kinase CK1alpha is responsible for the NSP5 hyperphosphorylation process. Here it is shown, by means of specific RNA interference, that in vivo, CK1alpha is the enzyme that initiates phosphorylation of NSP5. Lack of NSP5 hyperphosphorylation affected neither its interaction with the virus VP1 and NSP2 proteins normally found in viroplasms, nor the production of viral proteins. In contrast, the morphology of viroplasms was altered markedly in cells in which CK1alpha was depleted and a moderate decrease in the production of double-stranded RNA and infectious virus was observed. These data show that CK1alpha is the kinase that phosphorylates NSP5 in virus-infected cells and contribute to further understanding of the role of NSP5 in RV infection.
Campagna
Michela
M
International Centre for Genetic Engineering and Biotechnology, Padriciano 99, 34011 Trieste, Italy.
Budini
Mauricio
M
Instituto de Ciencias Biomedicas, Programa de Biologia Celular y Molecular, Facultad de Medicina, Universidad de Chile, Independencia 1027, 8380453 Santiago, Chile.
Arnoldi
Francesca
F
International Centre for Genetic Engineering and Biotechnology, Padriciano 99, 34011 Trieste, Italy.
Desselberger
Ulrich
U
International Centre for Genetic Engineering and Biotechnology, Padriciano 99, 34011 Trieste, Italy.
Allende
Jorge E
JE
Instituto de Ciencias Biomedicas, Programa de Biologia Celular y Molecular, Facultad de Medicina, Universidad de Chile, Independencia 1027, 8380453 Santiago, Chile.
Burrone
Oscar R
OR
International Centre for Genetic Engineering and Biotechnology, Padriciano 99, 34011 Trieste, Italy.
D048128:Casein Kinase Ialpha / Q000172:deficiency*; D017930:Genes, Reporter; D010766:Phosphorylation; D010957:Plasmids; D034741:RNA, Small Interfering / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics; D012401:Rotavirus / Q000201:enzymology* / Q000235:genetics / Q000502:physiology / Q000648:ultrastructure; D014162:Transfection; D017361:Viral Nonstructural Proteins / Q000378:metabolism; D014764:Viral Proteins / Q000235:genetics; D014779:Virus Replication / Q000502:physiology*
D034741:RNA, Small Interfering; D012367:RNA, Viral; D017361:Viral Nonstructural Proteins; D014764:Viral Proteins; D048128:Casein Kinase Ialpha