The fate of pro-TNF-alpha following inhibition of metalloprotease-dependent processing to soluble TNF-alpha in human monocytes
| Autor: | K A Solomon, M B Covington, C P DeCicco, R C Newton |
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| Rok vydání: | 1997 |
| Předmět: | |
| Zdroj: | The Journal of Immunology. 159:4524-4531 |
| ISSN: | 1550-6606 0022-1767 |
| Popis: | Human monocytes rapidly produce TNF-alpha following activation by bacterial LPS. The message for TNF-alpha encodes a 26-kDa protein that is proteolytically processed to the secreted 17-kDa form. Sequencing of the N terminus of the protein secreted by monocytes shows processing of the 26-kDa pro-TNF-alpha to a mature form at the projected metalloprotease cleavage site to generate 17-kDa TNF-alpha with the N terminus VRSSSR-. The addition of hydroxamic acid-based metalloprotease inhibitors to the cell culture is capable of blocking >95% of the production of soluble TNF-alpha and leads to a transient, but reproducible, increase in cell surface TNF-alpha as measured by FACS analysis. The cell surface TNF-alpha was demonstrated to increase the cell's ability to kill L929 tumor targets and induce PG production from human gingival fibroblasts. The buildup of cell surface TNF-alpha is unable to account for the TNF-alpha that is not secreted when inhibitor is present. Pulse-chase analysis of the cells demonstrates rapid degradation of the pro-TNF-alpha that remains unprocessed in the monocytes. Through inhibition of processing and secretion by brefeldin A, processing was shown to occur at a postendoplasmic reticulum site and is closely associated with movement to the cell surface. |
| Databáze: | OpenAIRE |
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