The objective of this research is to define the mechanisms involved in regulation of transcription of bacteriophage Mu. Mu early transcription is carried out by host function; middle transcription requires Mu functions and replication; late transcription requires the middle operon product C. This project will involve (1) isolation and analysis of phage containing middle operon mutations, (2) definition of the role of a transactivator protein Tam in activation of the middle promoter Pm by cloning of the tam gene, overproduction and purification of Tam protein, and characterization of its mechanism of action in an in vitro transcription system and (3) determination of the role of DNA replication, overlapping transcription termination, and specific DNA binding proteins (Mu repressors, transposase, and host IHF) on Pm activation by S1 nuclease and Pm-lacZ fusion analysis under differing levels of replication, termination and binding proteins. The results should provide a detailed understanding of the mechanisms of transcriptional control exhibited by this novel bacteriophage, and thus should enhance our knowledge regarding this important target for cellular regulation.
