In vivo Transcriptional Analysis of Latent Tuberculosis

描述（由申请人提供）：我们在分子水平上对结核病慢性期的理解是不完整的，肯定需要改进。对结核病早期和晚期的基因表达谱进行比较分析，将丰富我们对结核病不同阶段的认识。该项目的主要目的是确定负责在小鼠中建立慢性结核病的基因调控网络。基于一组强有力的初步结果，我们假设结核杆菌积极转录特定的基因集来调节其在宿主内的代谢活动，尽管在疾病的慢性阶段保持恒定的水平。为了确定慢性感染过程中分枝杆菌基因调控网络的关键组成部分，我们将采用体内微阵列分析：

DESCRIPTION (provided by applicant): Our understanding of the chronic stages of tuberculosis on a molecular level is incomplete and definitely needs improvement. Comparative analysis of gene expression profiled during early vs. late stages of tuberculosis will enrich our understanding of various stages of tuberculosis. The main objective of this project is to identify gene regulatory networks responsible for establishing chronic tuberculosis in mice. Based on a strong set of preliminary results, we hypothesize that the tuberculous bacilli are actively transcribing specific sets of genes to regulate their metabolic activity inside the host despite remaining at a constant level during the chronic stase of the disease. To identify key components of the mycobacterial gene regulatory network during chronic infection, we will employ the in vivo microarray analysis to: 1. Identify M.tb. genes responsible for entering into the chronic stage of tuberculosis. We will employ the mouse model of chronic tuberculosis combined with microarray analysis to identify differentially expressed genes in mice during the first 220 days of infection as well as during in vitro cultures. A reactivation model of mouse tuberculosis also will be assessed to mimic human tuberculosis during immune suppression. 2. Characterize the role of putative persistence genes in establishing chronic tuberculosis. We will follow organ colonization of M.tb. strains with defective persistence genes (generated by allelic exchange) after aerosol infection of BALB/c mice in comparison to the wild type strain of M.tb. H37Rv. Also, the transcriptional profile of mutants growing under defined in vitro stress conditions will be assessed using DNA microarrays. We believe that the approaches exploited in this proposal will generate a wealth of information related to the metabolic states of M.tb. during chronic stages of infection. The proposed studies will further improve our understanding of the nature of the host-pathogen interactions that can provide potential targets for drug development. Outcomes from this project will provide the necessary foundation for more detailed and focused studies on specific sets of genes utilizing human tissues. Transcriptional regulators that could contribute to both chronic and reactivation stages of tuberculosis will be the target for future analysis.