Regulation of cellular trafficking and pathogenesis by mucins

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The aims of this proposal was to elucidate the biological roles of mucins in a variety of pathogenic processes focusing on their involvement in cellular recognition and trafficking. Epithelial mucins were investigated for their distribution, their biosynthetic regulation, and the mechanism of recognition by carbohydrate binding molecules (lectins). The importance of such recognition processes in cancer invasion and metastasis, allergic dermatitis, and other diseases was investigated. Important findings are listed herein : (1) A unique carbohydrate epitope in human colonic sulfomucin was identified as sulfo-LeィイD1aィエD1(HSOィイD23ィエD2-Galβ1-3[Fucα1-4]GlcNAc-). Mucins with this epitope, presumably MUC5B was found associated with gallstone in humans. Positional localization of mucins with this epitope was very similar between humans and mice as far as in the intestinal tracts. (2) The initial step of glycosylation of MUC2 mucin was investigated focusing on the maximum number and the order of … More GalNAc incorporation into multiple threonine residues. When a microsomal fraction of LS174T human colon carcinoma cells was used as a source of the enzyme, GalNAc incorporation was 100% to consecutive threonine residues whereas 〜50% to alternating threonine residues. Three recombinant UDP-GalNAc : peptide/N-acetylgalactosaminyltransferases (ppGalNAc-T1, T2, and T3) were tested for their capacity to transfer GalNAc into three consecutive threonine residues, the order and the maximum number was unique to each enzyme. (3) Recombinant human macrophage galactose/N-acetylgalactosamine-specific calcium-type lectin (hMGL) was tested for its interaction with MUC2 mucin peptides with various arrangement of attached GalNAc residues. hMGL, forming a trimeric configuration, was shown to preferentially bind MUC2 peptides with GalNAc on consecutive threonines. (4) A mucin-like high-molecular-weight and heavily glycosylated molecule was identified to be associated with mouse ovarian tumor OV2944 HM-1 cells. This molecule was recognized by a mouse macrophage C-type lectin (mMGL). OV2944 HM-1 variant cells selected for low mMGL binding, deficient in this cell surface mucin, were more metastatic to lymph nodes when injected subcutaneously into syngeneic mice. (5) Migration of mMGL positive dermal macrophages from the skin to lymph nodes were observed during the sensitization phase of contact dermatitis. Although mucins were already identified as one of natural ligands of mMGL, whether such a molecule in lymph nodes is involved in macrophage trafficking remains to be elucidated. Less