Enhancer-independent Mu transposition from two topologically distinct synapses.

Abstract

Transposition of Mu is strictly dependent on a specific orientation of the left (L) and right (R) ends of Mu and a distant enhancer site (E) located on supercoiled DNA. Five DNA crossings are trapped in the three-site synapse, two of which are contributed by the interwrapping of L and R. To determine the contribution of E to the interwrapping of Mu ends, we examined the topology of the LR synapse under two different enhancer-independent reaction conditions. One of these conditions, which also alleviates the requirement for a specific orientation of Mu ends, revealed two topologically distinct arrangements of the ends. In their normal relative orientation, L and R were either plectonemically interwrapped or aligned by random collision. Addition of the enhancer to this system channeled synapsis toward the interwrapped pathway. When the ends were in the wrong relative orientation, synapsis occurred exclusively by random collision. In the second enhancer-independent condition, which retains the requirement for a specific orientation of Mu ends, synapsis of L and R was entirely by interwrapping. The two distinct kinds of synapses also were identified by gel electrophoresis. We discuss these results in the context of the "topological filter" model and consider the many contributions the enhancer makes to the biologically relevant interwrapped synapse.