STUDIES INTO THE MECHANISM OF MHV TRANSCRIPTION Ralph S. Baric l , Chien Kou Shieh 2 , Michael M.C. Lai 2 2 Stephen A. Stohlman , and luniversity of North Carolina School of Public Health Department of Parasitology and Laboratory Practice 348 Rosenau Hall, 201H Chapel Hill, North Carolina 27514 2University of Southern California School of Medicine Department of Microbiology and Neurology 2025 Zonal Avenue Los Angeles, California 90033 INTRODUCTION Coronaviruses are enveloped, plus-polarity RNA viruses which repli- b . d f RNA .. 1,2,3 p. d·· cate y a mo e 0 stu our laboratory have indicated that RNA recombination occurs ".t very high frequency during mixed infection with two heterologous strains of MHV 4 ,5 These data, coupled with the presence of discrete larger leader-contain- ing RNAs which range from 47 to 1000 nucleotides in length in MHV-infect- 6 ed cells, suggest that discrete RNA intermediates are synthesized during transcription which may dissociate and reassort between viral RNA 1 b·· b h . h· 4 temp ates to generate recom y a copy-c mec Therefore, the larger leader-containing RNAs in MHV-infected cells may represent functional intermediates of RNA transcription and recombina- tion. In this paper, we have analyzed the origin, structure, and prob- able mechanism of synthesis of these RNAs. These data provide evidence that MHV RNA transcription pauses at sites corresponding to hairpin loops in the RNA template or product strands and that these RNA intermediates may dissociate and reassociate with the RNA template intermittently during the course of transcription. We shall also present a new approach to identify the sequences encoded at the intergenic start sites for transcription which function in subgenomic mRNA synthesis. This approach utilizes a murine retrovirus 137 M. M. C. Lai et al. (eds.), Coronaviruses © Plenum Press, New York 1987