Messenger RNAs transcribed from yeast linear cytoplasmic plasmids possess unconventional 5’ and 3’ UTRs and suggest a novel mechanism of translation

Václav Vopálenský; Michal Sýkora; Tomáš Mašek; Martin Pospíšek

These results also indirectly strongly support the prediction that K2Orf3p either alone or as a part of a protein complex functions as a regular capping enzyme, exhibiting all three essential enzymatic activities necessary for complete conventional m 7 G cap synthesis. (Table S1A) . Mapping the 5' ends of pGKL transcripts helped us to classify genes encoded by pGKL plasmids into two major groups: genes transcribed to frequently 5'-capped mRNAs with low average number of 5' non-template adenosine nucleotides (K2ORF2, K2ORF3, K2ORF4 and K2ORF8) and a larger group of genes producing transcripts less frequently 5'-capped but with a higher degree of 5' non-template polyadenylation (Fig. 3C ). UCR sequences may themselves contain signals determining the 5' end formation of the pGKL transcripts. We took advantage of K. lactis strains containing . CC-BY-NC-ND 4.0 International license author/funder. It is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the . https://doi.org/10.1101/325316 doi: bioRxiv preprint altered sets of linear plasmids as described in Fig. 2 . We investigated selected transcripts from the K. lactis IFO1267[pRKL2-1] strain ( Fig. 2C and D) by 5' RACE. The pRKL2-1 linear plasmid is a pGKL2 derivative that contains a G418 resistance marker under the control of K1UCR2 and a K2ORF2 artificially controlled by K1UCR1 (Fig. 2 ) Indeed, we found that the presence of a cap at the 5' end of pRKL2-1 mRNAs, as well as the presence and length of a 5' poly(A) leader, are directly influenced by the UCR sequence used for expression.