Distribution Patterns of and Gene Clusters and New Insights into the Formation of the Architecture of pJP4, a Canonical 2,4-dichlorophenoxyacetic Acid (2,4-D) Degradation Plasmid.

Currently, pJP4 is one of the best-known plasmids for the biodegradation of xenobiotics that mediate the degradation of 2,4-dichlorophenoxyacetic acid (2,4-D), which is associated with serious health and environmental risks. Although the sequencing and proposed theory of pJP4 formation occurred almost 20 years ago (2004), pJP4 is still the model object of many studies focused on the biodegradation of 2,4-D. The uniqueness of this plasmid is due to the presence of two evolutionarily distinct gene clusters, and , controlling the degradation of 2,4-D. Recent advances in plasmid biology, especially those concerning the characterization of new IncP-1 plasmids and the systematization of gene cluster findings, serve as a basis for proposing new insights into the formation of the clusters' architecture of the canonical plasmid, pJP4, and their distribution among other plasmids. In the present work, a comparative genomic and phylogenetic in silico study of plasmids with and clusters was carried out. The possible initial distribution patterns of clusters among plasmids of different incompatibility groups (non-IncP-1) and clusters among IncP-1 plasmids using the IS-based composite transposon were revealed. A new theory on the formation of the architecture of the and clusters of pJP4 through sequential internal rearrangements, recombination, and IS insertion, is proposed. In addition, small gene clusters resulting from internal rearrangements of pJP4 ( and ORF31/32) served as fingerprints for exploring the distribution of and clusters. The revealed patterns and formulated theory extend the frontiers of plasmid biology and will be beneficial for understanding the role of plasmids in bacterial adaptation to xenobiotic-contaminated environments.