Characterization of MUC1 complexes in human oral epithelial cells
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Mucins are a famiIy of heavy glycosylated high molecular weight glycoproteins which are invoIved in the protection and lubrication of luminal epithelial surfaces. At least 19 human mucin genes have been distinguished by cDNA cloning—MUC1, MUC2, MUC3, MUC4, MUC5AC, MUC5B, MUC6-MUC9, MUC11-MUC13, and MUC15-MUC20. They have been classified into three distinct families: gel forming (MUC2, MUC5AC, MUC5B, MUC6 and MUC19), soluble (MUC7), and membrane-associated (MUC1, MUC3, MUC4 and MUC12, MUC13, MUC15, MUC16, MUC17, MUC18 and MUC20). MUC8, MUC9 and MUC11 remain unclassified. MUC1 is a large, heavily O-glycosylated, transmembrane protein expressed on the apical membrane of many epithelial tissues, including oral epithelium. It is expressed as a heterodimer after translation of a single polypeptide and cleavage into two subunits in the endoplasmic reticulum. The extracellular domain of MUC1 can interact with other molecules which can alter the adhesive/anti-adhesive properties of cells. The MUC1-CT (intracellular domain) interacts with a variety of kinases and other proteins and is involved in signal transduction pathways. Therefore, our hypothesis is that MUC1 might serve as an outside-to-inside signal that alters the proliferation, differentiation or cell-adhesion status of the epithelial cell. In the first part of this project, we concentrated our study on the interaction of MUC1-CT with other proteins in human oral epithelial cells using Tandem Affinity purification and Mass Spectrometry. We identified GRP-78, the78-kDa glucose-regulated protein as a protein which interacts with the cytoplasmic tail of MUC1. GRP78 is thought to be crucial in protein folding, proper glycosylation, protein-protein interactions in oligomeric structures, and elimination of misfolded polipeptides. Recently, MUC1 has been reported to be able to translocate to mitochontria and nucleus, but the mechanism remains unclear. Considering our results and the function of GRP78, we suggest that GRP78 (glucose-regulated protein 78) may be involved in either the posttranslational cleavage of MUC1 or may regulate intracellular trafficking of the MUC1-CT. In the second part of this project, in order to study the function and interaction properties of intact MUC1, we cloned a full length MUC1 containing the Tandem Affinity tags and expressed it in oral epithelial cells. MU1 was found to interact with IQGAP1 (IQ motif containing GTPase activating protein 1). IQGAP1 is considered as a scaffold which participates in multiple fundamental cellular activities, including transcription, cell-cell attachment, and regulation of the cytoskeleton, and it has the ability to turn off activated Ras, which consequently interferes with the activation of several kinases in the Ras/Ref/MEK/ERK signaling pathway. However, MUC1 has been reported to activate the Grb2-Sos-Ras-MEK-ERK pathway. The interaction between these two proteins may contribute to a very important signaling event, by balancing activation and inactivation of this pathway. Further studies are likely to yield additional targets and functions for MUC1 and its interacting proteins.
PLEASE NOTE: This work is protected by copyright. Downloading is restricted to the BU community: please click Download and log in with a valid BU account to access. If you are the author of this work and would like to make it publicly available, please contact email@example.com.Thesis (D.Sc.)--Boston University, Henry M. Goldman School of Dental Medicine, 2007 (Dept. of Periodontology and Oral Biology).Includes bibliography: leaves 148-183.
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