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Updated: Aug 18, 2026
Mouse Naïve CD4+ T Cell Isolation and In vitro Differentiation into T Cell Subsets
Published on: April 16, 2015
Differences in internalization and intracellular processing of interleukin-1 associated with the two forms of
1Medical Products Division, E.I. Du-Pont De Nemours & Co., Glenolden Laboratory, PA 19036.
There are at least two classes of interleukin-1 (IL-1) receptor, namely p80, and 80 kDa single-chain protein found in T-cells, fibroblasts and many other cell types, and p68, a 68 kDa protein expressed in B-cells and macrophages. The to classes of IL-1 receptor show distinct differences in their substrate-binding site and molecular properties. In this study we show that the kinetics of IL-1 internalization and the consequences of ligand processing in the two subtypes of IL-1 receptor are also very different. The Raji cell line was used as a source of the p68 form of the IL-1 receptor, whereas the YT cell line was used as a source of p80 receptor. Under conditions of steady-state binding the IL-1 was equally distributed between cell-surface and intracellular sites in YT cells, compared with predominantly cell-surface binding (85%) in Raji cells. The mechanism of IL-1 processing was also different in the two cell types. In Raji cells 60% of internalized IL-1 was released from the cells in an intact form, whereas the remainder was degraded. All of the IL-1 extruded from YT cells was intact. The kinetics of IL-1 release was faster in Raji cells, with a half-time of 4.5 h compared with over 15 h in YT cells. SDS/PAGE analysis of internalized IL-1 in Raji cells revealed that the ligand was sequentially processed to trichloroacetic acid-soluble products. The YT receptor-ligand complex was resistant to dissociation at pH 5, whereas that in Raji cells rapidly dissociated at this pH. Treatment of Raji cells with the lysosomotropic agent chloroquine inhibited the degradation of IL-1 without having any effect on the amount of intact IL-1 in the intracellular compartments. From these data we conclude that the pathways of internalization, intracellular trafficking and overall processing of IL-1 are different for p68 IL-1 receptors compared with p80. This could have direct consequences for IL-1 action and IL-1 receptor regulation in the cell.
There are at least two classes of interleukin-1 (IL-1) receptor, namely p80, and 80 kDa single-chain protein found in T-cells, fibroblasts and many other cell types, and p68, a 68 kDa protein expressed in B-cells and macrophages. The to classes of IL-1 receptor show distinct differences in their substrate-binding site and molecular properties. In this study we show that the kinetics of IL-1 internalization and the consequences of ligand processing in the two subtypes of IL-1 receptor are also very different. The Raji cell line was used as a source of the p68 form of the IL-1 receptor, whereas the YT cell line was used as a source of p80 receptor. Under conditions of steady-state binding the IL-1 was equally distributed between cell-surface and intracellular sites in YT cells, compared with predominantly cell-surface binding (85%) in Raji cells. The mechanism of IL-1 processing was also different in the two cell types. In Raji cells 60% of internalized IL-1 was released from the cells in an intact form, whereas the remainder was degraded. All of the IL-1 extruded from YT cells was intact. The kinetics of IL-1 release was faster in Raji cells, with a half-time of 4.5 h compared with over 15 h in YT cells. SDS/PAGE analysis of internalized IL-1 in Raji cells revealed that the ligand was sequentially processed to trichloroacetic acid-soluble products. The YT receptor-ligand complex was resistant to dissociation at pH 5, whereas that in Raji cells rapidly dissociated at this pH. Treatment of Raji cells with the lysosomotropic agent chloroquine inhibited the degradation of IL-1 without having any effect on the amount of intact IL-1 in the intracellular compartments. From these data we conclude that the pathways of internalization, intracellular trafficking and overall processing of IL-1 are different for p68 IL-1 receptors compared with p80. This could have direct consequences for IL-1 action and IL-1 receptor regulation in the cell.
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