Accumulation of a 5′ proximal subgenomic RNA of Citrus tristeza virus is correlated with encapsidation by the minor coat protein

AbstractDuring replication, Citrus tristeza virus (CTV) produces large amounts of two unusual subgenomic (sg) RNAs that are positive-stranded and 5′ coterminal. Although these RNAs are produced in similar amounts and are similar in size, with LMT1 (∼750 nt) only slightly larger than LMT2 (∼650), we found that the similar sgRNAs are produced differently. We previously showed that the LMT1 RNA is produced by premature termination during genomic RNA synthesis. However, LMT2 production was found to correlate with virion assembly instead of RNA replication. The time course of accumulation of the LMT2 RNA occurred late, coinciding with virion accumulation. The long flexuous virions of CTV contain two coat proteins that encapsidate the virions in a polar manner. The major coat protein encapsidates ∼97% of the virion, while the minor capsid protein encapsidates the remainder of the genome beginning in the 5′ non-translated region with the transition zone at ∼630 nucleotides from the 5′ end. The section of the virion RNA that was encapsidated by CPm was identical in size to the LMT2 RNA, suggesting that the LMT2 RNA represented a portion of the viral RNA protected by CPm encapsidation. Mutations that abrogated encapsidation by CPm also abolished the accumulation of LMT2 RNA. Thus, these two unusual but similar RNAs are produced via different pathways, one from RNA replication and one processed by the virion assembly process. To our knowledge, this represents the first evidence of a viral RNA processed by the assembly mechanism