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... of the possible N gene function(s) in TGEV replication. Using synthetic defective-interfering (DI) RNA genomes, some groups have reported that downstream ISs suppress transcription from upstream ISs =-=(40, 43)-=-. In addition, data suggest that the N gene IS is the strongest initiator of TGEV subgenomic RNA transcription (35, 38). In our recombinant virus and replicon RNAs, GFP subgenomic mRNA synthesis was i...

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... of sg RNA it produces. This observation is in accordance with earlier data showing that the amounts of sg RNA that are generated by an IS can depend on the flanking sequences, which are not specific =-=(8, 15)-=-. The influence of the flanking sequences could stem from the overall RNA secondary structure. Recently, Zhang and Lai found a correlation between the amounts of sg RNA produced by wt and mut MHV mRNA...

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...r the mutants, the molar amounts of sgRNA7 and sgRNA6 were reduced by roughly one-half and one-third, respectively, relative to wildtype levels. A similar effect has been observed with DI RNAs of MHV =-=(13, 45)-=- and bovine coronavirus (18) into which multiple IGSs had been engineered, although it should be noted that this phenomenon was not seen with one MHV DI RNA (1). In one case (45), the inhibitory effec...

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...arger mRNA if the two TASs were 23 nt apart. The inhibition decreased as the distance between the two TASs was increased, resulting in equal amounts of the mRNAs if the TASs were separated by 124 nt (=-=Joo & Makino, 1995-=-). The authors concluded that the TAS for the smaller mRNA had some inhibitory effect on the upstream TAS. Further studies on MHV D-RNAs containing combinations of up to three TASs separated by 361–76...

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..., was observed (Pasternak et al., 2004). This confirmed earlier theoretical work on the MHV genome (Konings et al., 1988) and studies with coronavirus DI RNA replicon systems (van Marle et al., 1995; =-=Joo & Makino, 1995-=-; Krishnan et al., 1996), which have shown that relative order and/or location of TRSs in the genome play an important role. An additional aspect that was investigated was the origin of the TRS that i...

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...e 9-nt sequence could serve as a potential transcription initiation site. Previous studies have also suggested that some neighboring sequences of the IG region could affect transcription efficiencies =-=(8, 9, 21)-=-. Therefore, to determine whether the 9-nt sequence located at the 59 end of the genomic RNA can initiate mRNA transcription, we examined the effect of the neighboring 59-terminal sequence on the tran...

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...rs indicate the position in the TGEV genome. VOL. 78, 2004 TRANSCRIPTION MECHANISM IN CORONAVIRUS 991so nsApril 20, 2014 by PENN STATE UNIV http://jvi.asm.org/ D ow nloaded from for other coronavirus =-=(17, 23)-=-, in arterivirus (24), and in nidovirus-derived expression systems (7, 11, 36). Nucleotide substitutions, not allowing base pairing with cCS-B, within the first 3 nt of the CS-L led to the inhibition ...

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...ere observed when a TRS was placed closer to the 3 end (2). Another conclusion suggested by various studies is that downstream TRSs have an attenuating effect on the transcription from upstream TRSs =-=(10, 18, 20, 23, 45, 48)-=-. The potential of coronaviruses as vectors has not been studied in any detail. Recent observations have, however, revealed that these viruses have a number of attractive features that might make them...