title: Design of a specific colonic mucus marker using a human commensal bacterium cell surface domain.
abstract: Imaging living cells and organs requires innovative, specific, efficient, and well tolerated fluorescent markers targeting cellular components. Such tools will allow proceeding to the dynamic analysis of cells and the adaptation of tissues to environmental cues. In this study, we have identified and synthesized a novel non-toxic fluorescent marker allowing a specific fluorescent staining of the human colonic mucus. Our strategy to identify a molecule able to specifically bind to the human colonic mucus was on the basis of the mucus adhesion properties of commensal bacteria. We identified and characterized the mucus-binding property of a 70-amino acid domain (MUB(70)) expressed on the surface of Lactobacillus strains. The chemical synthesis of MUB(70) was achieved using the human commensal bacterium Lactobacillus reuteri AF120104 protein as a template. The synthesized Cy5-conjugated MUB(70) marker specifically stained the colonic mucus on fixed human, rabbit, and guinea pig tissues. Interestingly, murine tissue was not stained, suggesting significant differences in the composition of the murine colonic mucus. In addition, this marker stained the mucus of living cultured human colonic cells (HT29-MTX) and human colonic tissue explants. Using a biotinylated derivative of MUB(70), we demonstrated that this peptide binds specifically to Muc2, the most abundant secreted mucin, through its glycosylated moieties. Hence, Cy5-MUB(70) is a novel and specific fluorescent marker for mammalian colonic mucus. It may be used for live imaging analysis but also, as demonstrated in this study, as a marker for the diagnosis and the prognosis of colonic mucinous carcinomas.
authors.affiliation: Unité de Chimie des Biomolécules, Institut Pasteur, 28 rue du Dr. Roux, 75724 Paris Cedex 15, France.
authors.forename: Yves-Marie
authors.initials: YM
authors.lastname: Coïc
authors.forename: Francoise
authors.initials: F
authors.lastname: Baleux
authors.forename: Ömer
authors.initials: Ö
authors.lastname: Poyraz
authors.forename: Roman
authors.initials: R
authors.lastname: Thibeaux
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Labruyere
authors.forename: Fabrice
authors.initials: F
authors.lastname: Chretien
authors.forename: Iradj
authors.initials: I
authors.lastname: Sobhani
authors.forename: Thierry
authors.initials: T
authors.lastname: Lazure
authors.forename: Benjamin
authors.initials: B
authors.lastname: Wyplosz
authors.forename: Gunter
authors.initials: G
authors.lastname: Schneider
authors.forename: Laurence
authors.initials: L
authors.lastname: Mulard
authors.forename: Philippe J
authors.initials: PJ
authors.lastname: Sansonetti
authors.forename: Benoit S
authors.initials: BS
authors.lastname: Marteyn
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D002470:Cell Survival; D003106:Colon / Q000378:metabolism* / Q000382:microbiology; D004591:Electrophoresis, Polyacrylamide Gel; D005456:Fluorescent Dyes / Q000737:chemistry; D006031:Glycosylation; D006168:Guinea Pigs; D019073:HT29 Cells; D006367:HeLa Cells; D054884:Host-Pathogen Interactions; D006801:Humans; D007150:Immunohistochemistry; D052200:Limosilactobacillus reuteri / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D051379:Mice; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D055262:Mucin-2 / Q000378:metabolism*; D009093:Mucus / Q000378:metabolism* / Q000382:microbiology; D011485:Protein Binding; D011817:Rabbits; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity
chemical_list: D001426:Bacterial Proteins; D005456:Fluorescent Dyes; D055262:Mucin-2