Discovery of uncapped and 5 polyadenylated mRNAs in poxviruses and yeast virus-like elements comparison and similarities of both models

Abstract

Cytoplasmic double-stranded DNA virus-like elements (VLEs) are present in yeast species across nine genera. VLE archetypes are linear DNA elements pGKL1/2 from the yeast Kluyveromyces lactis. They exhibit unique features, including cytoplasmic localization, terminally linked proteins, and compact genomes carrying altogether 15 genes coding, besides others, for a killer toxin, DNA and RNA polymerases, a helicase, and a capping enzyme. Functions of many pGKL1/2genes remain unclear. However, proteins that are essential for cytoplasmic transcription of pGKL1/2-encoded genes have been characterised in more detail and demonstrated substantial similarity to their functional counterparts in vaccinia virus (VACV). We found recently that although VLEs encode their putative capping enzyme, most of their mRNAs are uncapped and lack 3'-polyadenylation. Unexpectedly, many VLE mRNAs contain short 5' poly(A) sequences which are not complementary to their genomic DNA. Considering similarities in transcriptional apparatuses of VLEs and poxviruses, we further analysed also poxviral mRNAs. VACV, a prototypical poxvirus that was crucial for the smallpox eradication, played a key role in discovering the eukaryotic 5' mRNA cap and mechanism of its synthesis. Curiously, our analysis of individual VACV mRNAs revealed that postreplicative VACV transcripts in addition to presence of non-templated 5' poly(A) leaders lack a 5' cap. We found that cap occurrence in viral mRNAs decreases from early to late viral transcripts and similarly as in VLE mRNAs correlates inversely with increasing length of the 5' non-templated poly(A) leaders. Furthermore, the sequence of transcriptional initiator (INR) influences capping frequency and poly(A) leader formation by similar mechanism in yeast VLEs and VACV. Our findings also suggest that 5' mRNA polyadenylation may induce its cap-independent protein synthesis via an unknown mechanism.Our results indicate that switch in synthesis of capped to non-capped but 5'-polyadenylated viral mRNAs may facilitate transition from cap-dependent to cap-independent synthesis of VACV proteins and thus my help the virus to take over the host translation machinery in the course of the viral infection.