Analysis of phage Mu DNA transposition by whole-genome Escherichia coli tiling arrays reveals a complex relationship to distribution of target selection protein B, transcription and chromosome architectural elements

Abstract

Of all known transposable elements, phage Mu exhibits the highest transposition efficiency and the lowest target specificity. In vitro, MuB protein is responsible for target choice. In this work, we provide a comprehensive assessment of the genome-wide distribution of MuB and its relationship to Mu target selection using high-resolution Escherichia coli tiling DNA arrays. We have also assessed how MuB binding and Mu transposition are influenced by chromosome-organizing elements such as AT-rich DNA signatures, or the binding of the nucleoid-associated protein Fis, or processes such as transcription. The results confirm and extend previous biochemical and lower resolution in vivo data. Despite the generally random nature of Mu transposition and MuB binding, there were hot and cold insertion sites and MuB binding sites in the genome, and differences between the hottest and coldest sites were large. The new data also suggest that MuB distribution and subsequent Mu integration is responsive to DNA sequences that contribute to the structural organization of the chromosome.

Unlocking Tn3-family transposase activity in vitro unveils an asymetric pathway for transposome assembly

“…For Mu, the target adaptor MuB binds DNA with relatively low specificity, preferring A/T-rich DNA stretches (23,24,40). In contrast, TnsC is addressed to specific sites by the Tn7-encoded target selector proteins TnsD or TnsE.…”

“…In addition to mediating transposition, TnpA is responsible for target immunity, a long-range regulatory phenomenon whereby transposons avoid inserting more than once into the same DNA region (18,19). Other systems with target immunity, such as phage Mu or Tn7, require an adaptor protein (MuB and TnsC, respectively) that controls integration by directing the transpososome to appropriate targets (20)(21)(22)(23)(24). The Tn3-family TnpA is the only transposonencoded protein directly involved in immunity and is the only possible determinant to account for the specificity whereby each element confers immunity to itself but not to other family members (18,25).…”

Transposable Phage Mu

“…The Mu domain configuration is assisted by MuB and several nucleoid-associated proteins (NAP), and promotes low-level transcription from an early prophage promoter, which controls the expression of Mu A and B , as well as several genes not essential for phage growth, including a ligase and a Ku-like DNA repair function (86, 87). MuB might provide a NAP-like function (88). It is proposed that the Mu domain provides long-term survival benefits to both the prophage and the host: to the prophage in bestowing transposition-ready topological properties unique to the Mu reaction, and to the host in contributing extraneous DNA housekeeping functions (85).…”

“…A DNA microarray analysis of target site selection during the Mu lytic phase in both E. coli and Salmonella found hot- and cold-spots throughout the genome, reflecting >1,000-fold variation in target preference (93, 94). Transcription had a strong negative influence on transposition (93), although a direct relationship between transcription and transposition is unlikely (88). …”

“…A whole-genome E. coli tiling array revealed that there were hot and cold MuB binding sites in the genome, and that Mu transposition was in the vicinity rather than within MuB bound regions (88). MuB is an AAA+ ATPase and a non-specific DNA-binding protein (Fig.…”