Functional features of the bovine enterovirus 5 '-non-translated region

The bovine enterovirus (BEV) serotypes exhibit unique features of the non-t ranslated regions (NTRs) which separate them from the other enteroviruses, Their most remarkable property is an additional genome region of 110 nt loc ated between the 5'-cloverleaf and the internal ribosome entry site (IRES), This genome region has the potential to form an additional cloverleaf stru cture (domain I*) separated from the 5'-cloverleaf (domain I) by a small st em-loop (domain I**). Other characteristics involve the putative IRES domai ns III and VI, In order to investigate the features of the 5'-NTR, several full-length coxsackievirus B3 (CVB3) cDNA plasmids with hybrid 5'-NTRs were engineered. After exchange of the CVB3 cloverleaf with the BEV1 genome reg ion representing both cloverleafs, a viable virus chimera was generated, De letion of domain I** within the exchanged region also yielded viable virus albeit with reduced growth capacity. Deletion of sequences encoding either the first or the second BEV cloverleaf resulted in non-infectious construct s. Hybrid plasmids with exchanges of the IRES-encoding sequence or the comp lete 5'-NTR were non-infectious. Transfection experiments with SP6 transcri pts containing 5'-NTRs fused to the luciferase message indicated that IRES- driven translation is enhanced by the presence of the CVB3 cloverleaf and b oth BEV1 cloverleaf structures, respectively. Deletion of either the first or the second BEV cloverleaf domain reduced but did not abolish enhanced lu ciferase expression. These results suggest that the substitution of two put ative BEV cloverleaf structures for the putative coxsackieviral cloverleaf yields viable virus, while BEV sequences encoding the IRES fail to function ally replace CVB3 IRES-encoding sequences.