Analysis of African cassava mosaic virus recombinants suggests strand nicking occurs within the conserved nonanucleotide motif during the initiation of rolling circle DNA replication.

Intact clones containing partial repeats of the genomic components of African cassava mosaic (ACMV DNAs A and B) are infectious when mechanically coinoculated onto Nicotiana benthamiana. Monomeric genomic components may be generated either by homologous recombination or, when two copies of the origin of replication (ori) are present, by a modified rolling circle replication mechanism in which nascent single-stranded DNA is resolved by the introduction of nicks at both oris. DNA B partial repeats with duplicated common region sequences containing combinations of wild-type sequences and nonlethal mutations at nucleotides 151 and 155 within the putative stem-loop region have been constructed and introduced into plants in the presence of DNA A. Analysis of progeny indicates that monomers are generated by DNA strand nicking preferentially between nucleotides 151 and 155, suggesting a nonrandom replicative release mechanism involving the ubiquitous TAATATTAC motif (nucleotides 146-154). Viable ACMV DNA A deletion mutants are known to revert to wild-type size during systemic infection by generating tandem repeats. The recombination point in one such revertant has been mapped between nucleotides 152 and 153. Just as ori-nicking enzymes mediate recombinational events during prokaryotic rolling circle DNA replication, the result suggests that a nick has been introduced in the virion-sense strand within the nonanucleotide motif (TAATATT decreases AC) during the initiation of ACMV DNA replication.