Mycobacterium tuberculosis is the causative agent of tuberculosis, a disease that together with human immunodeficiency virus (HIV) and malaria, is one of the main causes of mortality due to an infectious agent (31). According to the WHO, onethird of the world's population is infected asymptomatically with M. tuberculosis, representing a large reservoir of infection (11). To block further transmission and reactivation in the already-infected population, it is necessary to develop improved intervention strategies that require a better understanding of the host-pathogen interaction.Infection of a mammalian host by M. tuberculosis usually occurs by the aerosol route, and the lung is typically the principal organ affected. The bacteria initially reside in alveolar macrophages (18), where they are usually able to replicate. In order to identify M. tuberculosis components that may be responsible for successful bacterial intracellular survival, many individual genes whose expression levels are up regulated by the microorganism inside the phagosome have been analyzed (16). DNA microarray technology has made it possible to analyze the M. tuberculosis global transcriptional response to different stimuli. Experiments have been carried out in broth culture, using conditions that may mimic the macrophage environment (i.e., low pH, cell wall stress, starvation, hypoxia, heat shock, etc.) in resting or activated mouse macrophages, and in vivo, using the mouse lung model of infection. The results of these studies have recently been reviewed (2, 29). The complete gene expression profile of M. tuberculosis growing in mouse macrophages was defined by Schnappinger et al. tuberculosis has to face a DNA-and cell envelope-damaging environment that is rich in fatty acid and deficient in iron. The transcriptional profile of M. tuberculosis infecting human lungs indicates that the bacteria regulate genes involved in the evasion of the immune system (45). A similar analysis of M. tuberculosis in human monocyte-derived macrophages after 7 days of infection suggested the relevance of bacterial genes involved in transcriptional regulation (8). In addition, M. tuberculosis genes that are essential for the survival of bacteria in mouse macrophages and in mouse lungs have been identified by using the transposon site hybridization technique (46) and designer arrays for defined mutant analysis (30).In this work, we analyzed the gene expression profile of M. tuberculosis strain H37Rv infecting human macrophage-like THP-1 cells. These cells treated with phorbol-myristate acetate (PMA) differentiate into mature macrophages, providing a good model for analyzing the interaction of M. tuberculosis with primary macrophages in terms of receptor expression, bacterial uptake, survival, and replication (59). It has been demonstrated that after infection with M. tuberculosis, THP-1 cells, in a manner similar to that of monocyte-derived macrophages, produce low levels of oxygen radicals and do not produce nitric oxide (55). Moreover, THP-1 cells are a good mod...

Two-component systems are important constituents of bacterial regulatory networks. Results of this investigation into the role of the MprAB two-component system of Mycobacterium tuberculosis indicate that it is associated with the regulation of several stress-responsive regulons. Using a deletion mutant lacking portions of the response regulator, MprA, and the histidine kinase, MprB, it was demonstrated by real-time PCR, primer extension analyses and DNA microarrays that MprAB activates sigma factor genes sigE and sigB, under SDS stress and during exponential growth. SDS-inducible, MprA-dependent transcriptional start points were identified for mprA, sigE and sigB, and variations in distance between these points and MprA-binding sites suggest that MprA is involved in different mechanisms of promoter activation. Although most of the SigE regulon was downregulated in the deletion mutant, the cluster of genes Rv1129c, Rv1130 and Rv1131, which is associated with growth in monoctyes, was upregulated in the deletion mutant under SDS stress, and this upregulation was dependent upon atmospheric growth conditions. Multiple stress-associated genes of the DosR, SigD and IdeR regulons were also upregulated in the deletion mutant, during exponential growth and/or in the presence of SDS. Surprisingly, the deletion mutant had increased resistance to SDS compared to the parental strain, and enhanced growth in human peripheral blood monocytes, characteristics which may result from a loss of repression of stress-associated genes. INTRODUCTIONAs evidenced by its historical and current impact on human populations (Corbett & Raviglione, 2005;Zink et al., 2005), Mycobacterium tuberculosis is a highly successful pathogen. To withstand the challenges of aerosol transmission, growth within host macrophages, and prolonged encapsulation within lung granuloma, these organisms require the ability to respond to different types of stress. M. tuberculosis has 13 sigma factors (Cole et al., 1998), and DNA microarray analyses have shown the importance of several of these in regulating the changes in gene expression patterns associated with various stresses (Geiman et al., 2004;Manganelli et al., 2001Manganelli et al., , 2002. In addition to sigma factors, the response regulators of some two-component systems (TCSs) (Rison et al., 2005), such as DosR (Kendall et al., 2004;Park et al., 2003) and PhoP Perez et al., 2001;Walters et al., 2006), and other transcriptional regulators, such as IdeR (Dussurget et al., 1999;Manabe et al., 2005) and EmbR (Sharma et al., 2006), have been shown to modulate mycobacterial gene expression in response to particular stresses. However, in general, the mechanisms by which M. tuberculosis senses a particular stress, and activates the appropriate regulatory factor(s) while inhibiting others, are largely unknown. The GEO accession numbers for the array data associated with this paper are GSM155424-155447 (series record no. GSE6750).Four supplementary tables are available with the online version of this paper.Abbreviati...

This study was conducted to investigate the role of iron deprivation in the persistence of Mycobacterium tuberculosis. We present evidence of iron restriction in human necrotic granulomas and demonstrate that under iron starvation M. tuberculosis persists, refractive to antibiotics and capable of restarting replication when iron is made available. Transcriptomics and metabolomic analyses indicated that the persistence of M. tuberculosis under iron starvation is dependent on strict control of endogenous Fe utilization and is associated with upregulation of pathogenicity and intrinsic antibiotic resistance determinants. M. tuberculosis mutants compromised in their ability to survive Fe starvation were identified. The findings of this study advance the understanding of the physiological settings that may underpin the chronicity of human tuberculosis (TB) and are relevant to the design of effective antitubercular therapies.

Mycobacterium tuberculosis survives in macrophages and usually subverts the bactericidal mechanisms of these phagocytes. The understanding of this host-pathogen interaction is relevant for the development of new treatments for tuberculosis. The adaptation of M. tuberculosis to intracellular life depends on its ability to regulate the expression of its genes. Sigma factors are important bacterial transcription activators that bind to the RNA polymerase and give it promoter specificity. Sigma factor E (SigE) controls the expression of genes that are essential for virulence. We have identified the SigE regulon during infection of macrophages, and we analyzed the impact of this regulon on the transcriptional response of phagocytes. Our results indicate that SigE regulates the expression of genes involved in the maintenance of M. tuberculosis cell envelope integrity and function during macrophage infection. Analysis of the phagocytes' transcriptional response indicates that the SigE regulon is involved in the modulation of the inflammatory response.

Glioblastoma (GBM) is the most common and aggressive human brain tumor. Human cytomegalovirus (HCMV) immediate early (IE) proteins that are endogenously expressed in GBM cells are strong viral transactivators with onconcogenic properties. Here, we show how HCMV IE are preferentially expressed in glioma stem-like cells (GSC), where they co-localize with the other GBM stemness markers, CD133, Nestin, and Sox2. In patient-derived GSC that are endogenously infected with HCMV, attenuating IE expression by an RNA-i-based strategy, was sufficient to inhibit tumorsphere formation, Sox2 expression, cell cycle progression, and cell survival. Conversely, HCMV infection of HMCV-negative GSC elicited robust self-renewal and proliferation of cells that could be partially reversed by IE attenuation. In HCMV-positive GSC, IE attenuation induced a molecular program characterized by enhanced expression of mesenchymal markers and pro-inflammatory cytokines, resembling the therapeutically-resistant GBM phenotype. Mechanistically, HCMV/IE regulation of Sox2 occurred via inhibition of miRNA-145, a negative regulator of Sox2 protein expression. In a spontaneous mouse model of glioma, ectopic expression of the IE1 gene (UL123) specifically increased Sox2 and Nestin levels in the IE1-positive tumors, upregulating stemness and proliferation markers in vivo. Similarly, human GSC infected with the HCMV strain Towne but not the IE1-deficient strain CR208 showed enhanced growth as tumorspheres and intracranial tumor xenografts, compared to mock-infected human GSC. Overall, our findings offer new mechanistic insights into how HCMV/IE control stemness properties in glioblastoma cells.