CAPSID CODING SEQUENCE IS REQUIRED FOR EFFICIENT REPLICATION OF HUMANRHINOVIRUS-14 RNA

Mechanisms by which the plus-sense RNA genomes of picornaviruses are r eplicated remain poorly defined, but existing models do not suggest a role for sequences encoding the capsid proteins. However, candidate RN A replicons (Delta P1 beta gal and Delta P1Luc), representing the sequ ence of human rhinovirus 14 virus (HRV-14) with reporter protein seque nces (P-galactosidase or luciferase, respectively) replacing most of t he P1 capsid-coding region, failed to replicate in transfected H1-HeLa cells despite efficient primary cleavage of the polyprotein. To deter mine which P1 sequences might be required for RNA replication, HRV-14 mutants in which segments of the P1 region were removed in frame from the genome were constructed. Mutants with deletions involving the 5'-p roximal 1,489 nucleotides of the P1 region replicated efficiently, whi le those with deletions involving the 3' 1,079 nucleotides did not. Re introduction of the 3' P1 sequence into the nonreplicating Delta P1Luc construct resulted in a new candidate replicon, Delta P1Luc/VP3, whic h replicated well and expressed luciferase efficiently. Capsid protein s provided in trans by helper virus failed to rescue the nonreplicatin g Delta P1Luc genome but were able to package the larger-than-genome-l ength Delta P1Luc/VP3 replicon. Thus, a 3'-distal P1 capsid-coding seq uence has a previously unrecognized cis-active function related to rep lication of HRV-14 RNA.