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TNF-alpha-converting enzyme cleaves the macrophage colony-stimulating factor receptor in macrophages undergoing
E Rovida1, A Paccagnini, M Del Rosso
1Dipartimento di Patologia e Oncologia Sperimentali, Università di Firenze, Florence, Italy. Immunex, Seattle, WA 98101, USA.
Abstract:
We previously reported that macrophage activators such as LPS, IL-2, and IL-4 down-modulate the M-CSFR via a mechanism involving protein kinase C and phospholipase C. In this study, we showed that M-CSFR is shed from macrophage surface and identified the protease responsible for M-CSFR cleavage and down-modulation. The shedding of M-CSFR elicited by phorbol esters (tetradecanoylphorbol myristate acetate (TPA)) or LPS in murine BAC.1-2F5 macrophages was prevented by cation chelators, as well as hydroxamate-based competitive inhibitors of metalloproteases. We found that the protease cleaving M-CSFR is a transmembrane enzyme and that its expression is controlled by furin-like serine endoproteases, which selectively process transmembrane metalloproteases. M-CSFR down-modulation was inhibited by treating cells in vivo, before TPA stimulation, with an Ab raised against the extracellular, catalytic domain of proTNF-converting enzyme (TACE). TACE expression was confirmed in BAC.1-2F5 cells and found inhibited after blocking furin-dependent processing. Using TACE-negative murine Dexter-ras-myc cell monocytes, we found that in these cells TPA is unable to down-modulate M-CSFR expression. These data indicated that TACE is required for the TPA-induced M-CSFR cleavage. The possibility that the cleavage is indirectly driven by TACE via the release of TNF was excluded by treating cells in vivo with anti-TNF Ab. Thus, we concluded that TACE is the protease responsible for M-CSFR shedding and down-modulation in mononuclear phagocytes undergoing activation. The possible physiological relevance of this mechanism is discussed.
Insights
Macrophage activators cause shedding of the M-CSFR protein by activating the TACE enzyme. This shedding, crucial for macrophage activation, is mediated by TACE, a metalloprotease involved in M-CSFR down-modulation.
Area of Science:
- Immunology
- Cell Biology
- Molecular Biology
Background:
- Macrophage colony-stimulating factor receptor (M-CSFR) is crucial for mononuclear phagocyte function.
- Macrophage activators like LPS, IL-2, and IL-4 down-modulate M-CSFR through protein kinase C and phospholipase C pathways.
- The precise mechanism of M-CSFR shedding and down-modulation during macrophage activation remained unclear.
Purpose of the Study:
- To identify the protease responsible for M-CSFR shedding and down-modulation.
- To elucidate the mechanism by which macrophage activators induce M-CSFR cleavage.
- To investigate the role of TACE (TNF-converting enzyme) in M-CSFR regulation.
Main Methods:
- Utilized murine macrophages (BAC.1-2F5 and Dexter-ras-myc cell monocytes).
- Employed phorbol esters (TPA) and LPS as macrophage activators.
- Used cation chelators, metalloprotease inhibitors, and antibodies against TACE and TNF.
- Investigated the role of furin-like serine endoproteases in regulating the protease.
Main Results:
- M-CSFR shedding induced by TPA or LPS was inhibited by metalloprotease inhibitors.
- A specific transmembrane metalloprotease, TACE, was identified as the key protease responsible for M-CSFR cleavage.
- TACE expression and activity were found to be regulated by furin-like serine endoproteases.
- TACE-negative cells showed no M-CSFR down-modulation upon TPA stimulation, confirming TACE's essential role.
- Evidence excluded indirect TACE-mediated cleavage via TNF release.
Conclusions:
- TACE is the primary protease responsible for M-CSFR shedding and down-modulation in activated mononuclear phagocytes.
- This TACE-mediated M-CSFR cleavage is a critical mechanism in macrophage activation.
- The findings provide insights into the regulation of M-CSFR and its role in immune cell function.