A proline-rich region in the coxsackievirus 3A protein is required for the protein to inhibit endoplasmic reticulum-to-golgi transport.
The ability of the 3A protein of coxsackievirus B (CVB) to inhibit protein secretion was investigated for this study. Here we show that the ectopic expression of CVB 3A blocked the transport of both the glycoprotein of vesicular stomatitis virus, a membrane-bound secretory marker, and the alpha-1 protease inhibitor, a luminal secretory protein, at a step between the endoplasmic reticulum (ER) and the Golgi complex. CVB 3A contains a conserved proline-rich region in its N terminus. The importance of this proline-rich region was investigated by introducing Pro-to-Ala substitutions. The mutation of Pro19 completely abolished the ability of 3A to inhibit ER-to-Golgi transport. The mutation of Pro14, Pro17, or Pro20 also impaired this ability, but to a lesser extent. The mutation of Pro18 had no effect. We also investigated the possible importance of this proline-rich region for the function of 3A in viral RNA replication. To this end, we introduced the Pro-to-Ala mutations into an infectious cDNA clone of CVB3. The transfection of cells with in vitro-transcribed RNAs of these clones gave rise to mutant viruses that replicated with wild-type characteristics. We concluded that the proline-rich region in CVB 3A is required for its ability to inhibit ER-to-Golgi transport, but not for its function in viral RNA replication. The functional relevance of the proline-rich region is discussed in light of the proposed structural model of 3A.
Wessels
Els
E
Department of Medical Microbiology, Nijmegen Center for Molecular Life Sciences, University Medical Center Nijmegen, P.O. Box 9101, 6500 HB Nijmegen, The Netherlands.
Duijsings
Daniël
D
Notebaart
Richard A
RA
Melchers
Willem J G
WJ
van Kuppeveld
Frank J M
FJ
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002522:Chlorocebus aethiops; D017931:DNA Primers; D019281:Dimerization; D004721:Endoplasmic Reticulum / Q000502:physiology*; D029822:Enterovirus B, Human / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D006056:Golgi Apparatus / Q000502:physiology*; D007668:Kidney; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D011392:Proline; D011487:Protein Conformation; D021381:Protein Transport; D012367:RNA, Viral / Q000235:genetics; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
C500427:3A protein, coxsackievirus B; D017931:DNA Primers; D012367:RNA, Viral; D014764:Viral Proteins; D011392:Proline