Activation of Tumor Necrosis Factor-α-converting Enzyme-mediated Ectodomain Shedding by Nitric Oxide

Zili Zhang,Jay K Kolls,Peter Oliver,David Good,Paul O Schwarzenberger,Mahesh S Joshi,Julie L Ponthier,Jack R Lancaster

doi:10.1074/jbc.m000604200

Zili Zhang, Jay K Kolls + Show 6 more

Open Access

https://doi.org/10.1074/jbc.m000604200

Copy DOI

Abstract

Ectodomain shedding of cell surface proteins is an important process in a wide variety of physiological and developmental events. Recently, tumor necrosis factor-alpha-converting enzyme (TACE) has been found to play an essential role in the shedding of several critical surface proteins, which is evidenced by multiple developmental defects exhibited by TACE knockout mice. However, little is known about the physiological activation of TACE. Here, we show that nitric oxide (NO) activates TACE-mediated ectodomain shedding. Using an in vitro model of TACE activation, we show that NO activates TACE by nitrosation of the inhibitory motif of the TACE prodomain. Thus, NO production activates the release of cytokines, cytokine receptors, and adhesion molecules, and NO may be involved in other ectodomain shedding processes.

Highlights

Ectodomain shedding of cell surface proteins is an important process in a wide variety of physiological and developmental events
Tumor necrosis factor␣-converting enzyme (TACE) has been found to play an essential role in the shedding of several critical surface proteins, which is evidenced by multiple developmental defects exhibited by tumor necrosis factor␣-converting enzyme (TACE) knockout mice
Enhancement of TNF␣ Shedding by nitric oxide (NO)—To test the possible involvement of NO in activation of TACE, we added an exogenous source of NO, PAPA/NO, to LPS/PMA-stimulated Mono Mac 6 cells, a human monocytic cell line that expresses TACE [8]

Summary

EXPERIMENTAL PROCEDURES

Chemicals—(Z)-1-[N-(3-ammoniopropyl)-N-(n-propyl)-amino]-diazen1-ium-1,2-diolate (PAPA/NO), NG-monomethyl-L-arginine (NMA), and 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one were obtained from Alexis Biochemicals (San Diego, CA). IEC-6 cells (ATCC, Manassas, VA) were cultured in Dulbecco’s modified Eagle’s medium with 5% fetal bovine serum. Cells were Ͼ95% viable macrophages (determined by trypan blue staining) and were cultured in Dulbecco’s modified Eagle’s medium with 10% fetal bovine serum. TNF␣ production was induced in Mono Mac 6 cells by stimulation with a combination of 20 ng/ml LPS and 60 ng/ml PMA for 3 h in the presence of varying concentrations of PAPA/NO. Co-culture of Mono Mac 6 and IEC-6 Cells—IEC-6 cells, a rat intestinal epithelial cell line, were stimulated with both murine interferon-␥ (10 units/ml) and LPS (20 ng/ml) for 18 h to induce NO production. Mono Mac 6 cells were co-cultured with this cell line in the presence of continuous murine interferon-␥ and LPS stimulation for 3 h. Experiments were performed in the presence of 100 ␮M NMA or 5 ␮M HbO2 to antagonize NO production or activity, respectively

Activation of TACE by NO

RESULTS

DISCUSSION