Coordinate replication of alfalfa mosaic virus RNAs 1 and 2 involves cis- and trans-acting functions of the encoded helicase-like and polymerase-like domains.
RNAs 1 and 2 of the tripartite genome of alfalfa mosaic virus encode the replicase proteins P1 and P2, respectively, whereas RNA 3 encodes the movement protein and coat protein. Transient expression of wild-type (wt) and mutant viral RNAs and proteins by agroinfiltration of plant leaves was used to study cis- and trans-acting functions of the helicase-like domain in P1 and the polymerase-like domain in P2. Three mutations in conserved motifs of the helicase-like domain of P1 affected one or more steps leading to synthesis of minus-strand RNAs 1, 2, and 3. In leaves containing transiently expressed P1 and P2, replication of wt but not mutant RNA 1 was observed. Apparently, the transiently expressed P1 could not complement the defect in replication of the RNA 1 mutant. Moreover, the transiently expressed wt replicase supported replication of RNA 2, but this replication was blocked in trans by coexpression of mutant RNA 1. However, expression of mutant RNA 1 did not interfere with the replication of RNA 3 by the wt replicase. Similarly, a mutation in the GDD motif encoded by RNA 2 could not be complemented in trans and affected the replication of RNA 1 by a wt replicase, while replication of RNA 3 remained unaffected. In competition assays, the transient wt replicase preferentially replicated RNA 3 over RNAs 1 and 2. The results indicate that one or more functions of P1 and P2 act in cis and point to the existence of a mechanism that coordinates the replication of RNAs 1 and 2.
D017794:Alfalfa mosaic virus / Q000235:genetics* / Q000502:physiology; D000595:Amino Acid Sequence; D036022:Capsid Proteins / Q000235:genetics / Q000502:physiology; D008969:Molecular Sequence Data; D009154:Mutation; D053830:Plant Viral Movement Proteins; D020365:RNA Helicases / Q000235:genetics / Q000502:physiology*; D012367:RNA, Viral / Q000096:biosynthesis*; D012324:RNA-Dependent RNA Polymerase / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D014764:Viral Proteins / Q000235:genetics / Q000502:physiology; D014779:Virus Replication
D036022:Capsid Proteins; D053830:Plant Viral Movement Proteins; D012367:RNA, Viral; D014764:Viral Proteins; C095691:coat protein, Alfalfa mosaic virus; D012324:RNA-Dependent RNA Polymerase; D020365:RNA Helicases