The objective is to understand the molecular mechanisms involved in transposition of mobile genetic elements. Specifically, we are studying transposition of bacteriophage Mu DNA. The mechanisms used in Mu transposition are, in essence, highly analogous to those used by other mobile elements ranging from prokaryotic insertion sequences and transposons, which mediate movement of antibiotic resistance determinants, to the proviral forms of eukaryotic retroviruses. The unique properties of Mu as a lysogenic virus and a transposon allow the use of biochemical studies of transposition in the entirety of a population, studies which are difficult or impossible in systems with very low frequencies of transposition. We have developed improved techniques for in vivo studies, including the synchronization of a single round of Mu replicative transposition, and we are initiating in vitro studies using purified proteins. We will focus on two broad, major avenues of research: I. Studies on the molecular mechanism of transposition, including studies on the isolation and characterization of transposition intermediates, the products of a single round of transposition, selection of target sites for transposition, the influence of transposon length, and the isolation of Mu and host mutants defective in transposition. II. Studies on the role of Mu encoded """"""""accessory functions"""""""" which stimulate Mu DNA replication and transposition, including mapping and sequencing of the accessory genes kil and arm from Mu and the closely related heteroimmune phage D108, purification of thge kil and arm proteins, and studies on the mechanism of action of the stimulatory activities of kil and arm in vivo and in vitro.
