Utilization of a mammalian cell-based RNA binding assay to characterize the RNA binding properties of picornavirus 3C proteinases.

Published

Journal Article

Using an assay capable of detecting sequence-specific RNA/protein interactions in mammalian cells, we demonstrate that the poliovirus and rhinovirus 3C proteinases are able to bind structured target RNA sequences derived from their respective 5' noncoding regions in vivo. Specific RNA binding by poliovirus 3C was found to be dependent on the integrity of stem-loop d of the RNA cloverleaf structure located at the 5' end of poliovirus genomic RNA. In contrast, mutation of stem-loop b did not prevent this in vivo interaction. However, mutation of stem-loop b, which serves as the RNA binding site for a cellular co-factor important for efficient poliovirus replication, did significantly attenuate the efficiency of 3C RNA binding in vivo and 3CD RNA binding in vitro. This in vivo protein:RNA binding assay was also used to identify several residues in 3C that are critical for RNA binding, but dispensable for 3C proteinase activity. The mammalian cell-based RNA binding assay described in this study may have considerable potential utility in the future detection or analysis of in vivo RNA/protein interactions unrelated to the 3C/RNA interaction described here.

Full Text

Duke Authors

Cited Authors

  • Blair, WS; Parsley, TB; Bogerd, HP; Towner, JS; Semler, BL; Cullen, BR

Published Date

  • February 1998

Published In

Volume / Issue

  • 4 / 2

Start / End Page

  • 215 - 225

PubMed ID

  • 9570321

Pubmed Central ID

  • 9570321

International Standard Serial Number (ISSN)

  • 1355-8382

Language

  • eng

Conference Location

  • United States