On the stability of sequences inserted into viral genomes

Willemsen, Anouk; Zwart, Mark P

The ssRNA(À) viruses are composed of genomes that range from 10 to 25.2 kbp in size. These viruses are particularly attractive candidates as viral vectors. While in ssRNA(þ) viruses inserts are subject to deletion, inserts in their ssRNA(À) counterparts appear more stable (Mebatsion et al.1996; Schnell et al. 1996) . One reason for this stability is that in general the genes in the ssRNA(À) viral genomes are non-overlapping and are expressed as separate mRNAs, thus consisting of a modular organization that can be easily manipulated for the insertion of foreign genes. If correctly engineered (e.g. without affecting any regulatory regions), one could expect that gene insertions are more stable in ssRNA(À) viruses as compared to ssRNA(þ) viruses, since the complexities surrounding correct processing of a polyprotein are not an issue here. Moreover, if expressed as a separate mRNA, the size of the insert is probably restricted only by the packaging limits of ssRNA(À) viruses. The low rate of homologous recombination in ssRNA(À) viruses can be another explanation for higher genomic stability (Chare, Gould, and Holmes 2003; Han and Worobey 2011) . Non-homologous recombination will probably rarely lead to variants with the insert deleted and other regions undisturbed, given it is less constrained than homologous recombination, and hence low homologous recombination rates could be a limiting factor on sequence evolution. However, genomic deletions that disrupt the inserted sequence will be subject to less constraints, as for example they can disrupt the reading frame of the insert without affecting the expression of virus genes.