title: Endotoxin-stimulated monocytes release multiple forms of IL-1 beta, including a proIL-1 beta form whose detection is affected by export.
abstract: The processing and release of 31-kDa proIL-1 beta to the mature 17-kDa form of IL-1 beta are still poorly understood. To help elucidate the mechanisms involved in IL-1 beta processing and release, we measured IL-1 beta forms released from endotoxin-stimulated monocytes by immunoprecipitation of [35S]methionine-labeled protein, by Western blots, and by our recently developed ELISA specific for proIL-1 beta. Our studies demonstrate that in addition to the 17-kDa mature IL-1 beta, IL-1 beta is also released as 31-, 28-, and 3-kDa molecules. The 31-kDa-released form of proIL-1 beta represented 20-40% of the total released IL-1 beta, as measured by SDS-PAGE with densitometry. This released proIL-1 beta was susceptible to ICE processing; however, this proIL-1 beta was not detectable by either a mature or proIL-1 beta-specific ELISA, suggesting that release induces a conformational change. The ELISA inability to detect proIL-1 beta was not due to inadequate sensitivity or subsequent degradation in the ELISA. Furthermore, while immunoaffinity-purified cytosolic proIL-1 beta could complex the type II IL-1R, released proIL-1 beta did not. Finally, the absence of a band shift in nondenaturing gel electrophoresis excluded proIL-1 beta binding to another protein. These findings imply that IL-1 beta is exported from monocytes as 3-, 17-, 28-, and 31-kDa forms and that the released 31-kDa form differs from cytosolic proIL-1 beta.
authors: Department of Pulmonary and Critical Care Medicine, Ohio State University, Columbus 43210, USA. wewers.2@osu.edu
authors: M D
authors: MD
authors: Wewers
authors: A V
authors: AV
authors: Winnard
authors: H A
authors: HA
authors: Dare
mesh_terms: D020170:Caspase 1 / Q000378:metabolism; D002474:Cell-Free System; D002478:Cells, Cultured; D003600:Cytosol / Q000276:immunology / Q000378:metabolism; D004731:Endotoxins / Q000494:pharmacology*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D015151:Immunoblotting; D007375:Interleukin-1 / Q000302:isolation & purification / Q000378:metabolism*; D008970:Molecular Weight; D009000:Monocytes / Q000201:enzymology / Q000276:immunology / Q000378:metabolism*; D011233:Precipitin Tests; D011485:Protein Binding / Q000276:immunology; D020033:Protein Isoforms / Q000302:isolation & purification / Q000378:metabolism; D011498:Protein Precursors / Q000302:isolation & purification / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000276:immunology*; D017472:Receptors, Interleukin-1 / Q000378:metabolism; D053574:Receptors, Interleukin-1 Type II
chemical_list: D004731:Endotoxins; D007375:Interleukin-1; D020033:Protein Isoforms; D011498:Protein Precursors; D017472:Receptors, Interleukin-1; D053574:Receptors, Interleukin-1 Type II; C043830:interleukin 1 precursor; D020170:Caspase 1