Termini of all mRNA species of Marburg virus: sequence and secondary structure

The 3' and 5' ends of Marburg virus (MBG)-specific mRNA species have been determined using reverse transcription-PCR, rapid amplification of cDNA ends, or the reverse ligation-mediated PCR procedure after removal of cap structures with tobacco acid pyrophosphatase. The polyadenylation sites of all MBG-specific mRNAs were strictly conserved and corresponded to the predicted transcriptional stop signals of genomic RNA. Determination of the 5' ends of the mRNA species showed that mRNA synthesis started precisely at the first nucleotide of a highly conserved transcriptional start site. The 5' ends of the mRNA species can build a stable secondary structure with the conserved nucleotides always located in the stem region of a hairpin. Nucleotide substitutions in the conserved 5' regions are accompanied by compensatory mutations of the complementary nucleotide thus leading to a conservation of the secondary structures. Compensatory mutations were also found when 5' ends of mRNA of MBG strain Musoke were compared with MBG strain Popp or the closely related Ebola virus, indicating that the secondary structures will be conserved even if the sequence is altered.