Integrated MicroRNA-mRNA-Analysis of Human Monocyte Derived Macrophages upon Mycobacterium avium subsp. hominissuis Infection

Abstract

BackgroundMany efforts have been made to understand basal mechanisms of mycobacterial infections. Macrophages are the first line of host immune defence to encounter and eradicate mycobacteria. Pathogenic species have evolved different mechanisms to evade host response, e.g. by influencing macrophage apoptotic pathways. However, the underlying molecular regulation is not fully understood. A new layer of eukaryotic regulation of gene expression is constituted by microRNAs. Therefore, we present a comprehensive study for identification of these key regulators and their targets in the context of host macrophage response to mycobacterial infections.Methodology/Principal FindingsWe performed microRNA as well as mRNA expression analysis of human monocyte derived macrophages infected with several Mycobacterium avium hominissuis strains by means of microarrays as well as quantitative reverse transcription PCR (qRT-PCR). The data revealed the ability of all strains to inhibit apoptosis by transcriptional regulation of BCL2 family members. Accordingly, at 48 h after infection macrophages infected with all M. avium strains showed significantly decreased caspase 3 and 7 activities compared to the controls. Expression of let-7e, miR-29a and miR-886-5p were increased in response to mycobacterial infection at 48 h. The integrated analysis of microRNA and mRNA expression as well as target prediction pointed out regulative networks identifying caspase 3 and 7 as potential targets of let-7e and miR-29a, respectively. Consecutive reporter assays verified the regulation of caspase 3 and 7 by these microRNAs.Conclusions/SignificanceWe show for the first time that mycobacterial infection of human macrophages causes a specific microRNA response. We furthermore outlined a regulatory network of potential interactions between microRNAs and mRNAs. This study provides a theoretical concept for unveiling how distinct mycobacteria could manipulate host cell response. In addition, functional relevance was confirmed by uncovering the control of major caspases 3 and 7 by let-7e and miR-29a, respectively.

The mammalian microRNA response to bacterial infections

“…75 The integrated analysis of microRNA and mRNA expression as well as target prediction and reporter assays identified caspases 3 and 7, major effector caspases essential in the triggering of apoptosis, as targets of let-7e and miR-29a, respectively, thus showing that inhibition of apoptosis after mycobacteria infection is controlled by miRNAs. Along the same line, miR-29a was upregulated in both serum and sputum of patients with active pulmonary tuberculosis compared with the levels in healthy controls.…”

Circulating MicroRNAs in Patients with Active Pulmonary Tuberculosis

“…(12,22). A recent study showed that miR-29a was specifically upregulated after the mycobacterial infection of human macrophages (24). The infection of mice with Listeria monocytogenes or Mycobacterium bovis bacillus Calmette-Guérin (BCG) downregulated miR-29 expression in IFN-␥-producing natural killer cells, CD4 ϩ T cells, and CD8 ϩ T cells.…”

“…China has the world's second largest tuberculosis epidemic, which has resulted in increased health care costs and other socioeconomic burdens. Recent studies showed that the mycobacterial infection of human macrophages causes a specific miRNA response (24), and the infection of mice with Mycobacterium bovis bacillus Calmette-Guérin (BCG) downregulates miR-29 expression in gamma interferon (IFN-␥)-producing natural killer cells, CD4 ϩ T cells, and CD8 ϩ T cells (16). These findings should open a new and interesting field in elucidating the relationship between miRNAs and TB infection.…”

Feeding of Enterococcus faecium NCIMB 10415 Leads to Intestinal miRNA-423-5p-Induced Regulation of Immune-Relevant Genes

“…Endotoxin-free plasmids were created for transfection according to the manufacturer's protocols for the NucleoBond Xtra Midi kit (Macherey-Nagel GmbH & Co. KG). Cotransfection of the reporter plasmid and miRNA mimics as well as measurement of luciferase activity were described previously (29).…”