Chlamydia trachomatis is an important human pathogen, recognized as an agent of ocular and sexually transmitted diseases, and implicated as cofactor in the transmission of HIV. In part, control of chlamydial infections is hampered by our poor understanding of the unique and complex life cycle of this intracellular parasite. Recently, eukaryotic histone-like proteins have been identified in Chlamydia and are thought to play key roles in regulating their developmental cycle. These histone homologs, of which two so far have been identified, are expressed in later stages of the Chlamydia life cycle. Their expression in recombinant E. coli produces chromatin condensation similar to the nucleoid condensation observed late in the parasites own life cycle. The goal of this project is to elucidate the nature and mechanisms of chlamydial histone-DNA interactions.
The specific aims are to: (1) elucidate the functional role of C. trachomatis histone H1 like protein (Hc1) structural domains. The functions of the carboxyl and amino termini will be examined using nucleic acid and protein synthesis, DNA stability to nuclease digestion and atomic force microscopy. (2) identify Hc1 specific binding sequences on chromosomal DNA using cross linking immune precipitation assays, in vitro footprinting and gel mobility shift experiments. (3) examine the association of Hc1 with actively transcribing genes compared to inactive genes using thermoregulation of chlamydial genes as a model. (4) investigate the role of Hc1-specific protease in DNA decondensation by characterizing the newly identified protease and examining its activity during the chlamydial life cycle. The experiments described here are designed to elucidate the key features of the chlamydial developmental cycle, namely initial decondensation to metabolically active form, followed by recondensation later in the cycle. Understanding the regulation of these features may contribute to novel methods for arresting or manipulating the chlamydial life cycle.
