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MUC1: the polymorphic appearance of a human mucin 总被引:14,自引:0,他引:14
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The complete genomic organization of the two mucin genes MUC2 and MUC6 was obtained by comparison of new and published mRNA sequences with newly available human genomic sequence. The two genes are located 38.5 kb apart in a head-to-head orientation within a gene complex on chromosome 11p15.5. The N-terminal organization of MUC6 is highly similar to that of MUC2, containing the D1, D2, D', and D3 Von Willebrand factor domains followed by the large tandem repeat domains located in exons 31 and 30, respectively. MUC6 has a much smaller C-terminal domain (101 amino acids) encoded by 2 exons containing only the CK domain, compared with MUC2, which has a C-terminal domain of 859 amino acids containing the D4, C, D, and CK domains, encoded by 19 exons. The gene structures agreed partially but not completely with predictions from gene prediction programs. 相似文献
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The N terminus of the MUC2 mucin forms trimers that are held together within a trypsin-resistant core fragment 总被引:2,自引:0,他引:2
Godl K Johansson ME Lidell ME Mörgelin M Karlsson H Olson FJ Gum JR Kim YS Hansson GC 《The Journal of biological chemistry》2002,277(49):47248-47256
The N terminus of the human MUC2 mucin (amino acids 1-1397) has been expressed as a recombinant tagged protein in Chinese hamster ovary cells. The intracellular form was found to be an endoglycosidase H-sensitive monomer, whereas the secreted form was an oligomer that gave monomers upon disulfide bond reduction. The secreted MUC2 N terminus contained a trypsin-resistant core fragment. Edman sequencing and mass spectrometry of the peptides obtained localized this core fragment to the C-terminal end of the recombinant protein. This core retained its oligomeric nature with an apparent mass of approximately 240 kDa. Upon reduction, peptides of approximately 85 kDa were found, suggesting that the N terminus forms trimers. This interpretation was also supported by gel electrophoresis and gel filtration of the intact MUC2 N terminus. Electron microscopy revealed three globular domains each linked via an extended and flexible region to a central part in a trefoil-like manner. Immunostaining with gold-labeled antibodies localized the N-terminal end to the three globular structures, and the antibodies directed against the Myc and green fluorescent protein tags attached at the C terminus localized these to the stalk side of the central trefoil. The N terminus of the MUC2 mucin is thus assembled into trimers that contain proteolytically stable parts, suggesting that MUC2 can only be partly degraded by intestinal proteases and thus is able to maintain a mucin network protecting the intestine. 相似文献
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During purification of a recombinant MUC2 C terminus expressed in CHO-K1 cells, the protein was partly cleaved when buffers with a pH of 6.0 were used. When buffers with higher pH values were used, less cleavage was found. Disulfide bonds held the two fragments generated together as these were only observed after reduction. Edman sequencing of the C-terminal 110-kDa fragment revealed that the cleavage had occurred at an Asp-Pro bond, a site described previously to generate the so-called "link peptide" after disulfide bond reduction. In vitro studies on the conditions for cleavage showed that it occurred in a time-dependent manner at a pH below 6.0. Furthermore, the reaction was not enzyme-mediated as it occurred in pure preparations of the MUC2 C terminus and was not inhibited by protease inhibitors. When expressed in the mucin producing cell line LS 174T, the C terminus was cleaved to a higher extent compared with the CHO-K1 cells. Neutralizing the secretory pathway with either NH(4)Cl or bafilomycin A1 inhibited this cleavage. Altogether, our results suggest that the cleavage is an autocatalytic reaction that occurs in the acidic environment of the late secretory pathway. Furthermore, the cleavage produced a new, reactive C terminus that has the potential to attach the mucin to itself or other molecules. Because a pH below 6 can be reached in the late secretory pathway and on mucosal surfaces, the cleavage and possible cross-linking are likely to be of biological importance. 相似文献
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A study of the intracellular and secreted forms of the MUC2 mucin from the PC/AA intestinal cell line. 总被引:2,自引:0,他引:2
In this study we present data on the entire population of MUC2 molecules secreted from and within the cell layer of an intestinal cell line. The molecular size distribution of the extracted molecules and their reactivity with two different MUC2 polypeptide antibodies indicated the presence of precursor and mature forms of the mucin. Oligomerized forms of the mucin were found in both the cell layer and medium; however, precursor forms were confined to the cell layer. Isopycnic density gradient centrifugation gave good resolution of mature and precursor forms of MUC2 as assessed by agarose gel electrophoresis. Three different populations of MUC2 were identified: one at low density (>1.3 g/ml) containing the N-glycosylated, non-O-glycosylated polypeptide; a second at intermediate density (1.3-1.35 g/ml) which may represent partially O-glycosylated intermediates; and a third at high density (1.36-1.48 g/ml) containing the mature MUC2 mucins. Rate-zonal centrifugation and agarose electrophoretic analysis of the low-density fraction indicated that the N-glycosylated MUC2 polypeptide was present as putative monomer and dimer/oligomer species. The combination of isopycnic density gradient centrifugation with agarose electrophoresis provides a new and simple approach that allows us to follow the MUC2 gene product from polypeptide through to the mature glycosylated mucin. 相似文献
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J R Gum J W Hicks D M Swallow R L Lagace J C Byrd D T Lamport B Siddiki Y S Kim 《Biochemical and biophysical research communications》1990,171(1):407-415
A human small intestinal lambda gt11 cDNA library was screened with antibodies to deglycosylated small intestinal mucin. Four partial cDNA clones were isolated that define a novel human mucin gene. These include two partial cDNA clones, SIB 124 and SIB 139, that contain 51 nucleotide tandem repeats which encode a seventeen amino acid repetitive peptide with a consensus sequence of HSTPSFTSSITTTETTS. SIB 139 hybridized to messages produced by small intestine, colon, colonic tumors and also by high mucin variant LS174T colon cancer cells. The gene from which cDNAs SIB 124 and SIB 139 are derived (proposed name MUC 3) maps to chromosome 7, distinct from other known human mucin genes. 相似文献
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Aspirin changes the secretion rate and amino acid composition of human small intestinal mucin in subjects with ileal conduits 总被引:1,自引:0,他引:1
Anthony M. Roberton Bandule Rabel Linda Stubbs Clifford Tasman-Jones Sum P. Lee 《Glycoconjugate journal》1996,13(5):781-789
The effect of aspirin on the rate of secretion and amino acid composition of human ileal mucin was studied, using subjects with ileal conduits as a model system in which mucin secreted from the ileal conduit tissue is flushed out in the urine and can be measured and analysed. Aspirin (600 mg per day, administered orally) increased the daily mucin output by 37–104% in subjects by days 3 or 4, but thereafter the mucin output declined to below the baseline level by day 10. Mucin samples, purified from the ileal conduit urine during the control period and during aspirin administration, were compared. There were no discernible changes in the degree of polymerisation or the density, but during aspirin administration the amino acid composition was significantly changed, and in particular threonine and proline were enriched. One possible explanation, consistent with the compositional analyses, is that the N- and C-terminal regions of the mucin subunits have been cleaved off and lost during aspirin administration. The observed changes in mucin secretion may have implications for the mechanism of the toxic effects of aspirin on the small intestine by altering the barrier properties of the mucus layer.Abbreviations EGF
epidermal growth factor
- NSAID
non-steroidal anti-inflammatory drug 相似文献
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Julia Pinkhasov M. Lucrecia Alvarez Latha B. Pathangey Teresa L. Tinder Hugh S. Mason Amanda M. Walmsley Sandra J. Gendler Pinku Mukherjee 《Cancer immunology, immunotherapy : CII》2010,59(12):1801-1811
Since epithelial mucin 1 (MUC1) is associated with several adenocarcinomas at the mucosal sites, it is pertinent to test the
efficacy of a mucosally targeted vaccine formulation. The B subunit of the Vibrio cholerae cholera toxin (CTB) has great potential to act as a mucosal carrier for subunit vaccines. In the present study we evaluated
whether a MUC1 tandem repeat (TR) peptide chemically linked to CTB would break self-antigen tolerance in the transgenic MUC1-tolerant
mouse model (MUC1.Tg) through oral or parenteral immunizations. We report that oral immunization with the CTB–MUC1 conjugate
along with mucosal adjuvant, unmethylated CpG oligodeoxynucleotide (ODN) and interleukin-12 (IL-12) did not break self-antigen
tolerance in MUC1.Tg mice, but induced a strong humoral response in wild-type C57BL/6 mice. However, self-antigen tolerance
in the MUC1.Tg mouse model was broken after parenteral immunizations with different doses of the CTB–MUC1 conjugate protein
and with the adjuvant CpG ODN co-delivered with CTB–MUC1. Importantly, mice immunized systemically with CpG ODN alone and
with CTB–MUC1 exhibited decreased tumor burden when challenged with a mammary gland tumor cell line that expresses human MUC1. 相似文献
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