Periodic genome sequences facilitate packaging in a single-stranded DNA virus

Icosahedral viruses organize and compact their genomes within volumetrically constrained capsids. Double-stranded (ds) DNA viral genomes form ordered spool-like structures when packaged into preformed capsids. By contrast, single-stranded (ss) RNA viral coat proteins recognize genomic sequences or structures, nucleating assembly around folded genomes. A similar mechanism may occur in some ssDNA systems; however, in parvo- and microviruses, the ss genome is concurrently synthesized and packaged into a preformed shell. In the øX174 X-ray virion structure, only ~12% of the genome is ordered. Consequently, the mechanism by which the genome is accommodated within the constrained capsid remains obscure. Sequence motifs within the øX174 genome produce a periodic segmentation pattern consistent with T = 1 icosahedral symmetry. To determine the function of these motifs, a subset within the first five packaged segments was altered. No regulatory elements or encoded amino acids were changed. The resulting mutant, øX ^DO5 , displayed a phenotype consistent with those of previously characterized packaging mutants. To further understand this phenomenon, the DNA replication-packaging pathway was characterized in cells co-infected with wild-type and øX ^DO5 . Each genome could both be separately tracked and distinguished throughout the pathway. Early, pre-packaging øX ^DO5 genome replication was comparable to the wild-type. However, øX ^DO5 genomes were severely diminished within virions, suggesting a defect in the transition from packaging intermediates to mature virions.