Transcriptional Profile of Mycobacterium tuberculosis Replicating in Type II Alveolar Epithelial Cells

Abstract

Mycobacterium tuberculosis (M. tb) infection is initiated by the few bacilli inhaled into the alveolus. Studies in lungs of aerosol-infected mice provided evidence for extensive replication of M. tb in non-migrating, non-antigen-presenting cells in the alveoli during the first 2–3 weeks post-infection. Alveoli are lined by type II and type I alveolar epithelial cells (AEC) which outnumber alveolar macrophages by several hundred-fold. M. tb DNA and viable M. tb have been demonstrated in AEC and other non-macrophage cells of the kidney, liver, and spleen in autopsied tissues from latently-infected subjects from TB-endemic regions indicating systemic bacterial dissemination during primary infection. M. tb have also been demonstrated to replicate rapidly in A549 cells (type II AEC line) and acquire increased invasiveness for endothelial cells. Together, these results suggest that AEC could provide an important niche for bacterial expansion and development of a phenotype that promotes dissemination during primary infection. In the current studies, we have compared the transcriptional profile of M. tb replicating intracellularly in A549 cells to that of M. tb replicating in laboratory broth, by microarray analysis. Genes significantly upregulated during intracellular residence were consistent with an active, replicative, metabolic, and aerobic state, as were genes for tryptophan synthesis and for increased virulence (ESAT-6, and ESAT-6-like genes, esxH, esxJ, esxK, esxP, and esxW). In contrast, significant downregulation of the DevR (DosR) regulon and several hypoxia-induced genes was observed. Stress response genes were either not differentially expressed or were downregulated with the exception of the heat shock response and those induced by low pH. The intra-type II AEC M. tb transcriptome strongly suggests that AEC could provide a safe haven in which M. tb can expand dramatically and disseminate from the lung prior to the elicitation of adaptive immune responses.

Understanding dissemination of Mycobacterium tuberculosis from the lungs during primary infection

“…In contrast, intramacrophage M. tuberculosis show upregulation of DosR (DevR) regulon genes and stress-related genes associated with transition to dormancy (Schnappinger et al, 2003;Fontán et al, 2008). While esat-6 is downregulated in M. tuberculosis in macrophages, esat-6 and five genes encoding ESAT-6-like proteins, esxH, esxW, esxJ, esxK and esxP, are upregulated in M. tuberculosis in type II AECs (Ryndak et al, 2015), as are genes encoding the laminin-binding adhesin MS, which promotes bacterial adherence to type II AECs (Kinhikar et al, 2006), Ag 85C (Rv0129c), which is a member of the fibronectinbinding Ag85 complex (Kuo et al, 2012), and Rv3922c, which has sequence similarity to haemolysins of other bacteria. Importantly, transcripts for these genes are unaffected during bacterial residence in macrophages (Schnappinger et al, 2003;Fontán et al, 2008).…”

“…This is supported by the transcriptional profile of M. tuberculosis replicating in human type II AECs (Ryndak et al, 2015). Thus, intra-AEC M. tuberculosis downregulate DosR (DevR) regulon genes and dormancyinducing stress-related genes, and upregulate genes involved in energy production, protein synthesis and cellwall synthesis (Ryndak et al, 2015).…”

“…), and toxins facilitate dissemination by increasing barrier permeability via cell lysis. The upregulation of transcripts for east-6 and hbha in M. tuberculosis replicating in AECs (Ryndak et al, 2015;Kinhikar et al, 2010;Delogu et al, 2006), and the greater attenuation of BCG Dhbha strain for migration across the in vitro alveolar barrier, is reminiscent of the synergism of adhesin(s) and toxin(s) reported for other bacteria for direct dissemination. Thus, Listeria monocytogenes adhesins (FbpA, ActA, InlA) promote attachment/invasion of small intestinal enterocytes (Dramsi et al, 2004;Suárez et al, 2001;Bierne et al, 2007) and listeriolysin O (LLO), a pore-forming toxin, impairs the integrity of the intestinal epithelial barrier (Richter et al, 2009).…”

“…This is supported by the transcriptional profile of M. tuberculosis replicating in human type II AECs (Ryndak et al, 2015). Thus, intra-AEC M. tuberculosis downregulate DosR (DevR) regulon genes and dormancyinducing stress-related genes, and upregulate genes involved in energy production, protein synthesis and cellwall synthesis (Ryndak et al, 2015). In contrast, intramacrophage M. tuberculosis show upregulation of DosR (DevR) regulon genes and stress-related genes associated with transition to dormancy (Schnappinger et al, 2003;Fontán et al, 2008).…”

“…Transcripts for ESAT-6, MS and HBHA are upregulated in M. tuberculosis in AECs (Ryndak et al, 2015;Delogu et al, 2006), while they are unaffected in bacteria replicating in macrophages (Schnappinger et al, 2003;Fontán et al, 2008). M. bovis BCG, M. tuberculosis H37Rv Desat-6 and M. bovis BCG Dhbha are attenuated for dissemination from the lungs in animal models (Lewis et al, 2003;Pethe et al, 2001).…”