Pestivirus translation initiation occurs by internal ribosome entry.
The role of the 385 nucleotide 5' noncoding region (NCR) in the translation of the pestivirus genome was investigated. In vitro translation of an RNA transcript containing the 5' NCR of the bovine viral diarrhea virus (BVDV) genome followed by the coding sequence of the first gene product (p20) of the BVDV large open reading frame resulted in the synthesis of a 20-kDa polypeptide. Results from hybrid-arrest translation studies identified a region involving a predicted RNA stem-loop structure spanning nucleotides 154-261 within the 5' NCR that was important for p20 synthesis. An additional inhibitory oligonucleotide was complementary to the sequence at the base of this stem-loop and encompassed the initiating AUG at nucleotide 386. Antisense oligonucleotides both upstream and downstream of those that were inhibitory had no effect on p20 translation. RNA from a dicistronic expression vector in which the BVDV 5' NCR was inserted between two reporter genes, CAT and LUC, showed strong expression of the second (LUC) cistron upon in vitro translation. This expression was dramatically reduced in an analogous construct in which nucleotides 173-236 of the 5' NCR were deleted. Similar results were obtained when RNA from these same vectors was evaluated for expression after transfection into BHK cells. These results suggest that the BVDV 5' NCR contains an internal ribosome entry site for translation initiation. This translational mechanism is similar to that shown for hepatitis C virus, further demonstrating the close relationship between viruses of these two genera within the family Flaviviridae.
Oak Ridge National Laboratory, Biology Division, Tennessee 37831.
T L
TL
Poole
C
C
Wang
R A
RA
Popp
L N
LN
Potgieter
A
A
Siddiqui
M S
MS
Collett
D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D006224:Cricetinae; D001908:Diarrhea Viruses, Bovine Viral / Q000235:genetics* / Q000502:physiology; D008969:Molecular Sequence Data; D010442:Peptide Chain Initiation, Translational; D012367:RNA, Viral / Q000235:genetics; D012270:Ribosomes / Q000821:virology*
D012367:RNA, Viral