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.
Highlights
Mononuclear phagocytes represent gateways of the host immune system for encountering and eliminating pathogens
Conclusions/Significance: We show for the first time that mycobacterial infection of human macrophages causes a specific microRNA response
We extended our investigations by quantitative reverse transcription PCR (qRT-PCR) analysis and by using an additional M. avium subsp. hominissuis (MAH) strain isolated from water
Summary
Mononuclear phagocytes represent gateways of the host immune system for encountering and eliminating pathogens. The genus Mycobacterium comprises highly pathogenic species and opportunists such as M. avium, which are able to withstand the hostile phagosomal environment and cause disseminated infections. Mycobacterial infections of humans and animals give cause for serious concern. Less pathogenic mycobacteria e.g. members of the M. avium complex (MAC) are able to cause disseminated infections in immuno-compromised persons such as AIDS patients [1]. MAC comprises the species M. intracellulare and M. avium. The latter is divided into four subspecies: M. avium subsp. Human mycobacteriosis is predominantly caused by MAH, while MAA and MAS affect primarily birds [4] and MAP is the causative agent of Johne’s disease. Pathogenic species have evolved different mechanisms to evade host response, e.g. by influencing macrophage apoptotic pathways. We present a comprehensive study for identification of these key regulators and their targets in the context of host macrophage response to mycobacterial infections
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