Design of a Specific Colonic Mucus Marker Using a Human Commensal Bacterium Cell Surface Domain

• Published in: The Journal of biological chemistry 2012-05, Vol.287 (19), p.15916-15922
• Main Authors: Coïc, Yves-Marie, Baleux, Francoise, Poyraz, Ömer, Thibeaux, Roman, Labruyere, Elisabeth, Chretien, Fabrice, Sobhani, Iradj, Lazure, Thierry, Wyplosz, Benjamin, Schneider, Gunter, Mulard, Laurence, Sansonetti, Philippe J., Marteyn, Benoit S.
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Bacterial Adhesion; Bacterial Proteins - chemistry; Bacterial Proteins - metabolism; Bacterial Proteins - pharmacology; Bacteriology; Bioengineering; Biomarkers; Cell Biology; Cell Survival; Chemical Sciences; Colon - metabolism; Colon - microbiology; Colonic Mucinous Carcinoma Diagnostic; Colonic Mucus; Electrophoresis, Polyacrylamide Gel; Fluorescence; Fluorescent Dyes - chemistry; Glycosylation; Guinea Pigs; HeLa Cells; Host-Pathogen Interactions; HT29 Cells; Humans; Imaging; Immunohistochemistry; Lactobacillus reuteri - genetics; Lactobacillus reuteri - metabolism; Lactobacillus reuteri - physiology; Life Sciences; Live Imaging; Medicin och hälsovetenskap; Mice; Microbiology and Parasitology; Microscopy, Fluorescence; Molecular Sequence Data; Muc2; Mucin-2 - metabolism; Mucins; Mucus - metabolism; Mucus - microbiology; Peptide Chemical Synthesis; Protein Binding; Protein-Protein Interactions; Rabbits; Sequence Homology, Amino Acid; Species Specificity; Tumor Marker
• ISSN: 0021-9258; 1083-351X; 1083-351X
• DOI: 10.1074/jbc.M111.310003

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 (MUB70) expressed on the surface of Lactobacillus strains. The chemical synthesis of MUB70 was achieved using the human commensal bacterium Lactobacillus reuteri AF120104 protein as a template. The synthesized Cy5-conjugated MUB70 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 MUB70, we demonstrated that this peptide binds specifically to Muc2, the most abundant secreted mucin, through its glycosylated moieties. Hence, Cy5-MUB70 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. Background: Imaging colonic mucus on living cells, tissues, and organs is required for live microscopy. Results: We have identified, synthesized, and validated a new human colonic mucus bacterial marker (MUB70). Conclusion: This non-toxic marker is used to image the secreted colonic mucus. Significance: Beyond imaging applications, Cy5-MUB70 might be used for diagnostic and prognosis applications on colonic mucinous carcinoma.