Immunolocalization of Picornavirus RNA in infected cells with antibodies to Tyr-pUp, the covalent linkage unit between VPg and RNA.

Abstract

The genomic RNA of picornaviruses is attached to a small protein (VPg) via a covalent bond between a tyrosine and a 5'-terminal uridine phosphate. The same structure is present in potyvirus and calicivirus families. VPgs play a key role in initiation of viral replication by acting as primers for RNA synthesis. The model compound [N(Ac),CO(NHMe)]Tyr-(5'P→O)Up-O-(CH(2))(6)NH(2) (mCLU), mimicking this 'covalent linkage unit' (CLU) and containing Tyr-pUp was synthesized in solution following the phosphoramidite scheme and used to raise antibodies for studying picornavirus infection. The antibodies recognized CLU-containing mengovirus RNA and showed minimal cross-reactivity with RNAs lacking CLU. Immunofluorescence staining of cells infected with a human rhinovirus demonstrated co-localization of the signals from anti-mCLU and from anti-VPg antibodies. Efficient synthesis of mCLU and anti-mCLU antibodies might be of great utility for investigating viral replication and identifying yet unknown viral and cellular CLU-containing RNA-protein complexes.

• Authors: Elena S Gavryushina, Sofia A Bryantseva, Elena S Nadezhdina, Timofei S Zatsepin, Ilya Yu Toropygin, Angela Pickl-Herk, Dieter Blaas, Yuri F Drygin
• Journal: Journal of virological methods (J Virol Methods) Vol. 171 Issue 1 Pg. 206-11 (Jan 2011) ISSN: 1879-0984 [Electronic] Netherlands
• PMID: 21056058 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2010 Elsevier B.V. All rights reserved.

Chemical References

• Antibodies, Viral
• Oligoribonucleotides
• RNA, Viral
• VPg-pUpU

Topics

• Animals
• Antibodies, Viral (isolation & purification)
• HeLa Cells
• Humans
• Microscopy, Fluorescence (methods)
• Oligoribonucleotides (chemical synthesis, immunology)
• Picornaviridae (chemistry, growth & development)
• RNA, Viral (analysis)
• Rabbits
• Virology (methods)