Splicing of Cauliflower mosaic virus 35S RNA serves to downregulate a toxic gene product

Abstract

Alternative splicing usually leads to an increase in the number of gene products that can be derived from a single transcript. Here, a different and novel use of alternative splicing – as a means to control the amount of a potentially toxic gene product in the plant pararetrovirus Cauliflower mosaic virus (CaMV) – is reported. About 70 % of the CaMV 35S RNA, which serves as a substrate for both reverse transcription and polycistronic mRNA, is spliced into four additional RNA species. Splicing occurs between four donor sites – one in the 5′ untranslated region and three within open reading frame (ORF) I – and one unique acceptor site at position 1508 in ORF II. A previous study revealed that the acceptor site is vital for CaMV infectivity and expression of ORFs III and IV from one of the spliced RNA species suggested that splicing may facilitate expression of downstream CaMV ORFs. However, it is shown here that deleting the splice acceptor site and replacing ORF II with a cargo ORF that lacks splice acceptor sites does not interfere with virus proliferation. Furthermore, it is demonstrated that whenever P2 cannot accumulate in infected tissues, the splice acceptor site at position 1508 is no longer vital and has little effect on virus replication. This suggests that the vital role of splicing in CaMV is regulation of P2 expression and that P2 exhibits biological properties that, whilst indispensable for virus–vector interactions, can block in planta virus infection if this regulation is abolished.