C-terminal domain of rodent intestinal mucin Muc3 is proteolytically cleaved in the endoplasmic reticulum to generate extracellular and membrane components.
Although human MUC3 and rodent Muc3 are both membrane-associated intestinal mucins, the present study has explored the possibility that rodent Muc3 might exist in soluble as well as membrane forms. No evidence was obtained for the existence of soluble splice variants; however, experiments with heterologous cells transfected with cDNA encoding the 381-residue C-terminal domain of rodent Muc3 showed that a definitive proteolytic cleavage occurs during processing in the endoplasmic reticulum. The products consisted of a V5-tagged 30 kDa extracellular glycopeptide and a Myc-tagged 49 kDa membrane-associated glycopeptide. Throughout their cellular transport to the plasma membrane, the two fragments remained associated by non-covalent SDS-sensitive interactions. Site-specific mutagenesis pinpointed the need for glycine and serine residues in the cleavage sequence Leu-Ser-Lys-Gly-Ser-Ile-Val-Val, which is localized between the two epidermal-growth-factor-like motifs of the mucin. A similar cleavage sequence (Phe-Arg-Pro-Gly downward arrow Ser-Val-Val-Val, where downward arrow signifies the cleavage site) has been reported in human MUC1 and analogous sites are present in human MUC3, MUC12 and MUC17. Thus early proteolytic cleavage may be a conserved characteristic of many membrane-associated mucins, possibly as a prelude to later release of their large extracellular domains at cell surfaces.
Wang
Rongquan
R
Research Institute, Department of Structural Biology and Biochemistry, The Hospital for Sick Children, 555 University Avenue, Toronto, ON, Canada.
Khatri
Ismat A
IA
Forstner
Janet F
JF
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D002522:Chlorocebus aethiops; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D014644:Genetic Variation; D006801:Humans; D006868:Hydrolysis; D007413:Intestinal Mucosa / Q000378:metabolism*; D051379:Mice; D008871:Microvilli / Q000378:metabolism; D055263:Mucin-3; D009077:Mucins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012377:Rodentia; D014162:Transfection
C524641:Muc3 protein, mouse; D055263:Mucin-3; D009077:Mucins; D011994:Recombinant Proteins