Fig. 1.

Umbravirus 3′ UTRs contain a BTE. (A) Representative predicted (Mfold) secondary structure of the BTE in the 3′ UTR of TBTV genomic RNA, and relative translation activities of selected mutants. 17 nt CS is in bold italics. The shaded bases were deleted in mutants dm1 and dm2 as indicated. In mutants sm1 and sm2, bases in dashed boxes were replaced by bases in solid boxes (dashed arrows). The luciferase activity generated by translation of uncapped mutant RNAs, as a percentage of wild type (100%) are indicated. (B) Genome organization of TBTV RNA and maps of translation reporter constructs containing TBTV UTRs. TUlucTU contains both complete UTRs of TBTV flanking the firefly luciferase coding sequence (fLUC). TUluc has only the TBTV 5′UTR. TUlucTUBF differs from TUlucTU by only a 4 nt insertion (underlined) in the BamH I site. Bases are numbered according to their position in the TBTV genome. (C) Relative translation activities of capped and uncapped reporter mRNAs in wheat germ extract. Luciferase activities obtained from the indicated RNAs are normalized to uncapped TUlucTU (defined as 100%) and shown as RLU (relative luciferase units). Error bars indicate standard error. (D) Predicted secondary structure of the GRV BTE. The 17 nt CS (italics) is in the dashed box, with bases that deviate from consensus indicated in bold italics. Mutant sequence (CS) which is identical to the consensus 17 nt CS is indicated at right by dashed arrow. (E) Map of reporter RNA GlucG. GlucG contains the GRV 5′UTR and a chimeric 3′UTR consisting of the TBTV 3′ UTR with the putative GRV BTE in place of the TBTV BTE. Positions of bases from each virus are indicated. (F) Relative translation activities of capped and uncapped reporter mRNAs in wheat germ extract.