IDENTIFICATION OF A TRANSCRIPTION ENHANCER IN CORONAVIRUS

Centro Nacional de Biotecnología, CSIC. Department of Molecular and Cell Biology. C/Darwin 3, Cantoblanco. 28049 Madrid, Spain

^* To whom correspondence should be addressed. Email: L.Enjuanes{at}cnb.uam.es.

	Abstract

Coronavirus (CoV) transcription includes a discontinuous mechanism during the synthesis of subgenome-length minus strand RNAs leading to a collection of mRNAs in which the 5' terminal leader sequence is fused to contiguous genome sequences. It has been previously shown that transcription-regulating sequences (TRSs) preceding each gene regulate transcription. Base pairing between the leader TRS (TRS-L) and the complement of the body TRS (cTRS-B) in the nascent RNA is a determinant factor during CoV transcription. In fact, in transmissible gastroenteritis CoV, a good correlation has been observed between subgenomic mRNA (sg mRNA) levels and the free energy (G) of TRS-L and cTRS-B duplex formation. The only exception was sg mRNA N, the most abundant during viral infection in spite of its minimum G associated to duplex formation. We postulated that additional factors should regulate transcription of sg mRNA N. In this report, we have described a novel transcription regulation mechanism operating in CoV by which a 9-nt sequence located 449 nucleotides upstream of N gene TRS core sequence (CS-N) interacts with a complementary sequence just upstream of CS-N, specifically increasing the accumulation of sg mRNA N. Alteration of this complementarity in mutant replicon genomes showed a correlation between the predicted stability of the base-pairing between 9-nt sequences and the accumulation of sg mRNA N. This interaction is exclusively conserved in group 1a CoVs, the only CoV subgroup in which N gene is not the most 3' end in the viral genome. This is the first time that a long distance RNA-RNA interaction regulating transcriptional activity specifically enhancing the transcription of one gene has been described in CoVs.