Related Experiment Videos
IFN-gamma increases cathepsin H mRNA levels in mouse macrophages
W P Lafuse1, D Brown, L Castle
1Department of Medical Microbiology, Ohio State University, Columbus 43210, USA.
Abstract:
Expression of major histocompatibility complex (MHC) class II molecules and ability to present antigen to T lymphocytes is acquired upon activation of the macrophage by interferon-gamma (IFN-gamma). Little information is available concerning immune regulation of protease gene expression in mouse macrophages. We have isolated a cDNA clone for cathepsin H, a lysosomal cysteine proteinase from a cDNA subtraction library of mouse macrophage genes induced by IFN-gamma, and have characterized its expression. The level of cathepsin H mRNA increased in mouse peritoneal macrophages following addition of IFN-gamma. Cathepsin H mRNA levels began to increase 8 h after the addition of IFN-gamma and was maximal at 24-48 h. This increase was concordant in time with appearance of MHC class II E beta mRNA and Ia invariant chain mRNA. The increase in cathepsin H mRNA levels by IFN-gamma was dose dependent. Cycloheximide treatment of peritoneal macrophages inhibited the increase in cathepsin H mRNA levels induced by IFN-gamma, suggesting that the increase in cathepsin mRNA levels requires de novo protein synthesis. Lipopolysaccharide and cytokines interleukin-2 (IL-2), IL-4, IL-10, and tumor necrosis factor alpha were found to have no effect on cathepsin H mRNA levels in mouse peritoneal macrophages.
Insights
Interferon-gamma (IFN-gamma) upregulates cathepsin H mRNA in mouse macrophages, a process requiring new protein synthesis. This immune regulation is linked to major histocompatibility complex (MHC) class II expression.
Area of Science:
- Immunology
- Molecular Biology
- Cell Biology
Background:
- Macrophages play a crucial role in immune responses, including antigen presentation via MHC class II molecules.
- Interferon-gamma (IFN-gamma) is a key cytokine that activates macrophages and enhances their immune functions.
- Regulation of protease gene expression in macrophages, particularly in response to immune stimuli, is not well understood.
Purpose of the Study:
- To investigate the immune regulation of cathepsin H, a lysosomal cysteine proteinase, gene expression in mouse macrophages.
- To characterize the effect of IFN-gamma on cathepsin H mRNA levels and its temporal dynamics.
- To explore the requirement of de novo protein synthesis for IFN-gamma-induced cathepsin H expression.
Main Methods:
- Isolation of a cathepsin H cDNA clone from a subtraction library of IFN-gamma-induced mouse macrophage genes.
- Quantitative analysis of cathepsin H mRNA levels in mouse peritoneal macrophages using Northern blotting or similar techniques.
- Treatment of macrophages with IFN-gamma, cycloheximide, lipopolysaccharide, and various cytokines to assess their effects on cathepsin H mRNA expression.
Main Results:
- IFN-gamma significantly increased cathepsin H mRNA levels in mouse peritoneal macrophages in a dose-dependent manner.
- The upregulation of cathepsin H mRNA by IFN-gamma occurred between 8 and 48 hours, coinciding with MHC class II E beta and Ia invariant chain mRNA expression.
- Inhibition of de novo protein synthesis by cycloheximide blocked the IFN-gamma-induced increase in cathepsin H mRNA, indicating a requirement for new protein synthesis.
- Other tested stimuli, including lipopolysaccharide and cytokines IL-2, IL-4, IL-10, and TNF-alpha, did not affect cathepsin H mRNA levels.
Conclusions:
- IFN-gamma is a potent inducer of cathepsin H gene expression in mouse macrophages.
- The induction of cathepsin H by IFN-gamma is a regulated process dependent on de novo protein synthesis and occurs concurrently with the upregulation of MHC class II-related genes.
- These findings shed light on the molecular mechanisms underlying macrophage activation and immune function by IFN-gamma.