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Published on: May 31, 2018
Reduced transcript stabilization restricts TNF-alpha expression in RAW264.7 macrophages infected with pathogenic
Tina Basler1, Helmut Holtmann, Jens Abel
1Institut fur Mikrobiologie, Stiftung Tierarztliche Hochschule Hannover, Hannover, Germany.
Abstract:
Despite the critical role that TNF-alpha plays in the containment of mycobacterial infection, the mechanisms involved in regulation of its expression by mycobacteria are poorly defined. We addressed this question by studying MAP, which causes a chronic enteritis in ruminants and is linked to human Crohn's disease. We found that in MAP infected macrophages, TNF-alpha gene expression was substantially lower than in macrophages infected with nonpathogenic MS or stimulated with LPS. TNF-alpha transcriptional one could not fully explain the differential TNF-alpha mRNA expression, suggesting that there must be a substantial contribution by post-transcriptional mechanisms.Accordingly, we found reduced TNF-alpha mRNA stability in MAP-infected macrophages. Further comparison of MAP- and MS-infected macrophages revealed that lower TNF-alpha mRNA stability combined with lower mRNA and protein expression in MAP-infected macrophages correlated with lower p38 MAPK phosphorylation. These findings were independent of viability of MAP and MS. We demonstrate that the major mycobacterial cell-wall lipoglycan LM of MAP and MS induced TNF-alpha mRNA transcription,but only the MS-LM induced p38 MAPK-dependent transcript stabilization. Overall, our data suggest that pathogenic mycobacteria cause weak p38 and TNF-alpha mRNA stabilization as a result of their structural cell-wall components such as LM and thereby, restrict TNF-alpha expression in macrophages.
Insights
Pathogenic mycobacteria, like MAP, suppress tumor necrosis factor-alpha (TNF-alpha) expression in macrophages. This occurs through reduced mRNA stability and p38 MAPK signaling, influenced by cell-wall components like lipoglycan (LM).
Area of Science:
- Immunology
- Microbiology
- Molecular Biology
Background:
- Tumor necrosis factor-alpha (TNF-alpha) is crucial for controlling mycobacterial infections.
- Mechanisms regulating TNF-alpha expression by mycobacteria, particularly pathogenic strains, remain unclear.
- Mycobacterium avium subspecies paratuberculosis (MAP) causes chronic enteritis and is linked to Crohn's disease.
Purpose of the Study:
- To investigate the mechanisms by which MAP downregulates TNF-alpha expression in macrophages.
- To compare MAP's effect on TNF-alpha regulation with a nonpathogenic mycobacterium (MS).
- To elucidate the role of post-transcriptional regulation and specific mycobacterial components in this process.
Main Methods:
- Macrophages were infected with MAP or MS, or stimulated with LPS.
- TNF-alpha mRNA and protein expression levels were quantified.
- mRNA stability was assessed.
- p38 MAPK phosphorylation was measured.
- The effect of lipoglycan (LM) from MAP and MS on TNF-alpha transcription and mRNA stability was evaluated.
Main Results:
- MAP-infected macrophages exhibited significantly lower TNF-alpha expression compared to MS-infected or LPS-stimulated cells.
- Post-transcriptional mechanisms, specifically reduced TNF-alpha mRNA stability, contributed to lower expression in MAP infection.
- Lower TNF-alpha mRNA stability and expression in MAP-infected cells correlated with reduced p38 MAPK phosphorylation.
- While both MAP- and MS-LM induced TNF-alpha transcription, only MS-LM promoted p38 MAPK-dependent transcript stabilization.
Conclusions:
- Pathogenic mycobacteria, such as MAP, employ strategies to limit TNF-alpha expression in macrophages.
- Structural cell-wall components, like lipoglycan (LM), play a role in modulating TNF-alpha regulation.
- MAP's lipoglycan may lead to weak p38 MAPK activation and reduced TNF-alpha mRNA stabilization, contributing to immune evasion.
