IL-1 beta is an important immunoregulatory molecule which is released from human monocytes upon stimulation with LPS. Because this protein lacks a leader sequence, it is not """"""""packaged"""""""" within the endoplasmic reticulum and is therefore not released from the cells via the same mechanism as other commonly secreted proteins. The exact mechanism of release of IL-1 is not known, although one theory set forth suggests that IL-1 is released from monocytes following apoptotic cell death. The purpose of this study is to determine what effect M-CSF, a cytokine able to promote monocyte survival, has on the release of IL-1 beta from human monocytes. Using a variety of techniques, including radiolabeling of monocyte proteins and immunoprecipitation, we have thus far determined that pretreatment of monocytes with MCSF results in a decrease in the amount of IL-1 beta released from human monocytes. This decrease appears to be due specifically to a suppression in the release of IL-1 from the cells and not to changes in transcription, translation or enzymatic conversion of the IL-1 beta molecule. Another theory of IL-1 release is that IL-1 is transported directly across the plasma membrane. Heat shock proteins are responsible for the transport of proteins across the endoplasmic reticulum and mitochondrial membranes, however to date there is no evidence that proteins are directly transported across the plasma membrane. One approach we've used to address this theory is to add a membrane insoluble cross linker to the cells 3-4 hours post LPS stimulation to determine whether the IL-1 can be cross linked to a surface molecule (possibly a heat shock protein) responsible for 'pulling' it through the membrane. We've found that the addition of the cross linker results in an increase in the 33 kD internal pro form of IL-1 and a marked suppression of the processed forms. One possible explanation is that IL-1 is being cross linked to a surface protein and this in turn is preventing the release of the IL-1. Studies are currently underway to determine if this is the case.
