Receptor-mediated internalization of bradykinin: DDT1 MF-2 smooth muscle cells process internalized bradykinin via multiple degradative pathways
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This study was undertaken to evaluate the role of internalization in the action of the peptide autacoid bradykinin (BK). At 4°C [3H]BK binds to an apparently single class of B2 kinin receptors on DDT1 MF-2 smooth muscle cells (C. M. Munoz, S. Cotecchia, and L. M. F. Leeb-Lundberg, manuscript submitted). At this temperature the [3H]BK binding was confined exclusively to the cell surface. On the other hand, at 37°C the B2 receptor-specific cell surface [3H]BK binding was rapidly followed by a receptor-specific internalization of [3H]BK (t1/2 ∼ 9 min). The internalization reached a steady-state level after 30-40 min that was 80-100% of the level of specifically bound [3H]BK on the cell surface at 4°C, and this level was maintained for ≥2 h. Internalized [3H]BK was routed via at least two intracellular degradative pathways which were distinguished primarily based on subcellular localization but also on a small but significant difference in the rate of [3H]BK degradation. One pathway was localized in a plasma membrane-enriched fraction and had a relatively high degradative capacity. Another pathway was localized in a microsomal fraction and had a relatively low degradative capacity. The internalized [3H]BK activity was rapidly released into the media (t1/2 ∼ 24 min). Following a single round of internalization, the released activity consisted almost exclusively of small [3H]BK fragments (<[3H]BK(1-5)). In contrast, at steady-state [3H]BK represented 30-40% of the released activity. While chloroquine (100 μM) did not alter the rate of [3H]BK internalization or release or the intracellular distribution of [3H] BK, this agent significantly decreased the rate of [3H] BK degradation in both pathways. In all, these results show that B2 kinin receptor-mediated internalization of BK is a process integral to the interaction of BK with DDT1 MF-2 smooth muscle cells and may be a mechanism for terminating BK actions by rapidly removing extracellular free and receptor-bound BK and accessing various intracellular BK degradative pathways.
|Research areas and keywords||
Subject classification (UKÄ) – MANDATORY
|Journal||Journal of Biological Chemistry|
|Publication status||Published - 1992 Dec 1|