Biosynthesis of Gastric Mucus Glycoprotein of the Rat

Dekker, Jan; Beurden-Lamers, W M O Van; Strous, Ger J.

doi:10.1016/s0021-9258(18)81639-8

Cited by 50 publications

(6 citation statements)

References 34 publications

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“…In some rats, the RCM precursor is visible as a double band, most likely representing two independent gene products, as the bands were equally intense after pulse-labelling and remained equally intense during a 4 h chase-incubation. This probably reflects expression of two different alleles, as has previously been shown for rat gastric mucin [24]. Heterogeneity between individual rats is most likely of genetic origin, and may reflect differences in number of tandem repeats, as described for the human MUC1 gene [40].…”

Section: Discussionmentioning

confidence: 63%

“…The mucosae were scraped. Total mucin was isolated from scrapings as was previously described for rat gastric mucin and HCM [23,24]. Briefly, scrapings were homogenized for 20 h at 4°C in homogenization buffer, containing 6 M guanidinium-HCl (Sigma) and 100 mM dithiothreitol (DTT; Sigma) for reduction in order to enhance solubility.…”

Section: Isolation Of Rcmmentioning

confidence: 99%

“…For comparison with HCM, human colonic biopsies were obtained at endoscopy from patients without colonic disease and the same protocol for metabolic labelling was followed. Biosynthesis was studied as previously described [23,24]. Briefly, to study the biosynthesis of RCM polypeptide, the tissue was first depleted of methionine and cysteine.…”

Section: Metabolic Labelling Studiesmentioning

confidence: 99%

“…Immunoprecipitated mucins were spun down and washed several times. To be able to distinguish between precursor proteins and mature fully glycosylated RCM, some immunoprecipitated samples were treated with 750 U endo-fl-N-acetylglucosaminidase H (endo H; New England Biolabs, Beverly, MA, U.S.A.) as described earlier[23,24,35]. Samples were run on SDS/PAGE, gels were stained with PAS and radiolabelled mucins were analysed by towards peptide epitopes was tested by digesting the peptide fluorography RESULTS Isolation of RCM RCMs were isolated using triple CsCl density gradients.…”

mentioning

confidence: 99%

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Biosynthesis of rat MUC2 in colon and its analogy with human MUC2

Tytgat

Bovelander

Opdam

et al. 1995

Biochemical Journal

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In order to identify the mucins synthesized and secreted in the rat colon, we studied their biochemical characteristics and biosynthesis and evaluated their analogy to human colonic mucins. Purified mucin from both species appeared similar with respect to composition, buoyant density and mobility on SDS/PAGE. Isolated rat colonic mucin (RCM) was used to elicit a polyclonal antiserum, which was used in metabolic labelling studies to identify mucins and mucin precursors. RCM is synthesized as a 600 kDa precursor protein, which oligomerizes before O-glycosylation. The mature, high-molecular mass mucin is secreted and displays an anomalous molecular mass on SDS/PAGE of approximately 650 kDa. Polymorphism in precursor size was found among different rats, suggesting genetic heterogeneity. Molecular mass, biosynthesis and secretion of RCM appeared similar to human MUC2. Moreover, RCM precursor could be immunoprecipitated using specific anti-(human MUC2) antisera, indicating that the RCM can be designated rat MUC2. This study describes the biosynthesis of two homologous mucins in two different species. The high degree of similarity suggests functional analogy.

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Section: Discussionmentioning

confidence: 63%

Section: Isolation Of Rcmmentioning

confidence: 99%

Section: Metabolic Labelling Studiesmentioning

confidence: 99%

mentioning

confidence: 99%

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Biosynthesis of rat MUC2 in colon and its analogy with human MUC2

Tytgat

Bovelander

Opdam

et al. 1995

Biochemical Journal

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“…As discussed at length previously (31), the mobilities of mature mucins are generally anomalous and are dependent on the intrinsic negative charge of the mucins, which is imposed by the high sialic acid content. Also, the presence of sulfate esters has been shown to have dramatic effects on the mobility of some mucins on gel (6,34); therefore, potential differences in sulfation may also contribute to the observed differences in mobility between the small intestinal and colonic MUC2. Thus the Muc2 in the small and large intestine of the mouse may differ in their glycan structure and composition.…”

Section: G122mentioning

confidence: 99%

Gastrointestinal expression and partial cDNA cloning of murine Muc2

Klinken¹,

Einerhand

Duits³

et al. 1999

American Journal of Physiology-Gastrointestinal and Liver Physi

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To help us investigate the role of mucin in the protection of the colonic epithelium in the mouse, we aimed to identify the murine colonic mucin (MCM) and its encoding gene. We isolated MCM, raised an anti-MCM antiserum, and studied the biosynthesis of MCM in the gastrointestinal tract. Isolated MCM resembled other mucins in physicochemical properties. Anti-MCM recognized MCM as well as rat and human MUC2 on Western blots, interacting primarily with peptide epitopes, indicating that MCM was identical to murine Muc2. Using anti-MCM and previously characterized anti-human and anti-rat MUC2 antibodies, we identified a murine Muc2 precursor in the colon of ∼600 kDa, which appeared similar in size to rat and human MUC2 precursors. Western blotting, immunoprecipitation of metabolically labeled mucins, and immunohistochemistry showed that murine Muc2 was expressed in the colon and the small intestine but was absent in the stomach. To independently identify murine Muc2, we cloned a cDNA fragment from murine colonic mRNA, encoding the 302 NH2-terminal amino acids of murine Muc2. The NH2 terminus of murine Muc2 showed 86 and 75% identity to the corresponding rat and human MUC2 peptide sequences, respectively. Northern blotting with a murine Muc2 cDNA probe showed hybridization to a very large mRNA, which was expressed highly in the colon and to some extend in the small intestine but was absent in the stomach. In situ hybridization showed that the murine Muc2 mRNA was confined to intestinal goblet cells. In conclusion, by two independent sets of experiments we identified murine Muc2, which appears homologous to rat and human MUC2. Because Muc2 is prominently expressed in the colon, it is most likely to be the predominant mucin in the colonic mucus layer.

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Review Article. Mucus Glycoproteins and Their Role in Colorectal Disease

Corfield

Warren

1996

J. Pathol.

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Biosynthesis of Gastric Mucus Glycoprotein of the Rat

Cited by 50 publications

References 34 publications

Biosynthesis of rat MUC2 in colon and its analogy with human MUC2

Biosynthesis of rat MUC2 in colon and its analogy with human MUC2

Gastrointestinal expression and partial cDNA cloning of murine Muc2

Review Article. Mucus Glycoproteins and Their Role in Colorectal Disease

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