Galectin-4 and sulfatides in apical membrane trafficking in enterocyte-like cells

We have previously reported that 1-benzyl-2-acetamido-2-deoxy-α-d-galactopyranoside (GalNAcα-O-bn), an inhibitor of glycosylation, perturbed apical biosynthetic trafficking in polarized HT-29 cells suggesting an involvement of a lectin-based mechanism. Here, we have identified galectin-4 as one of the major components of detergent-resistant membranes (DRMs) isolated from HT-29 5M12 cells. Galectin-4 was also found in post-Golgi carrier vesicles. The functional role of galectin-4 in polarized trafficking in HT-29 5M12 cells was studied by using a retrovirus-mediated RNA interference. In galectin-4–depleted HT-29 5M12 cells apical membrane markers accumulated intracellularly. In contrast, basolateral membrane markers were not affected. Moreover, galectin-4 depletion altered the DRM association characteristics of apical proteins. Sulfatides with long chain-hydroxylated fatty acids, which were also enriched in DRMs, were identified as high-affinity ligands for galectin-4. Together, our data propose that interaction between galectin-4 and sulfatides plays a functional role in the clustering of lipid rafts for apical delivery

Systematic Gene Overexpression in Candida albicans identifies a Regulator of Early Adaptation to the Mammalian Gut

We are grateful to members of the genomics core facility (PF2, Génopole) for the availability of the microarray scanner and the Alain Jacquier’s lab for making the GenePix software available. We are grateful to Drs. Suzanne Noble and Aaron Mitchell for providing C. albicans mutant collections. We thank all members of the Fungal Biology & Pathogenicity Unit, particularly Drs. Anne Neville and Adeline Feri for their numerous insights during the course of this project. This work has been supported by grants from the Agence Nationale de la Recherche (KANJI, ANR-08-MIE-033-01 to C.d’E. and F.D.; ERA-Net Infect-ERA, FUNCOMPATH, ANR-14-IFEC-0004; and CANDIHUB, ANR-14-CE-0018 to C.d’E.), the French Government’s Investissement d’Avenir program (Laboratoire d’Excellence Integrative Biology of Emerging Infectious Diseases, ANR-10-LABX-62-IBEID to C.d’E.; Institut de Recherche Technologique BIOASTER, ANR-10-AIRT-03 to C.d’E., F.D. and T.J.), the European Commission (FinSysB PITN-GA-2008-214004 to C.d’E.) and the Wellcome Trust (The Candida albicans ORFeome project, WT088858MA to C.d’E. and C.M.). C.M. acknowledges support from the Medical Research Council, UK (New Investigator Award, G0400284), the MRC Centre for Medical Mycology (MR/N006364/1) and the University of Aberdeen. S.Z. is an Institut Pasteur International Network Affiliate Program Fellow. S.Z., L.v.W. and A.H.C. were the recipients of post-doctoral fellowships from the European Commission (FINSysB, PITN-GA-2008-214004 to S.Z.), the Agence Nationale de la Recherche (KANJI, ANR-08-MIE-033-01 to S.Z.; ERA-Net Infect-ERA, FUNCOMPATH, ANR-14-IFEC-0004 to A.H.C.; CANDIHUB, ANR-14-CE-0018 to L.v.W) and the French Government’s Investissement d’Avenir program (Institut de Recherche Technologique BIOASTER, ANR-10-AIRT-03 to S.Z. and A.H.C.). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Peer reviewedPublisher PD

Biological modeling of mucus to modulate mucus barriers.

International audienceA recent study using a transgenic mouse, whose intestinal mucus contains a molecule made of 12 copies of a domain found in many gelling mucins, demonstrates that it is possible to strengthen mucus properties in situ, leading to promising new treatment strategies in diseases in which the mucosal barrier is impaired

The extraordinarily complex but highly structured organization of intestinal mucus-gel unveiled in multicolor images.

The mucus that coats the gastrointestinal tract of all mammals is a dynamic and sticky gel layer and represents the first protective barrier between the host and the hostile environment. There is, however, a lack of detailed knowledge about the mucus gel organization because of the high water content and the complexity of MUC2, the main gel-forming molecule in the intestine. Histological staining and a multilabel immunofluorescence method were used to examine mucus blankets and Muc2 in mouse colon and ileum samples fixed in Carnoy's solution, unveiling an extraordinarily complex but highly structured mucus gel organization. The inner firmly adherent mucus blanket consists of alternating layers. The thicker outer loosely adherent mucus blanket in the colon is made of alternating laminated layers and loose curl-like structures. The layers consist of Muc2 molecules with different fucosylation states and glycoforms remain unmixed in the mucus. Importantly, distinct goblet cell subpopulations throughout the ileum along the crypt-to-villus axis with an alternation of goblet cells secreting fucosylated and non-fucosylated Muc2 are observed. A better understanding of the mucus structure should contribute to improve the efficiency of DNA and drug delivery and will allow for a better understanding and treatment of inflammatory and infectious intestinal diseases

Preclinical mouse model to monitor live Muc5b-producing conjunctival goblet cell density under pharmacological treatments.

PURPOSE:Modification of mucous cell density and gel-forming mucin production are established hallmarks of mucosal diseases. Our aim was to develop and validate a mouse model to study live goblet cell density in pathological situations and under pharmacological treatments. METHODS:We created a reporter mouse for the gel-forming mucin gene Muc5b. Muc5b-positive goblet cells were studied in the eye conjunctiva by immunohistochemistry and probe-based confocal laser endomicroscopy (pCLE) in living mice. Dry eye syndrome (DES) model was induced by topical application of benzalkonium chloride (BAK) and recombinant interleukine (rIL) 13 was administered to reverse the goblet cell loss in the DES model. RESULTS:Almost 50% of the total of conjunctival goblet cells are Muc5b+ in unchallenged mice. The decrease density of Muc5b+ conjunctival goblet cell population in the DES model reflects the whole conjunctival goblet cell loss. Ten days of BAK in one eye followed by 4 days without any treatment induced a -18.3% decrease in conjunctival goblet cell density. A four days of rIL13 application in the DES model restored the normal goblet cell density. CONCLUSION:Muc5b is a biological marker of DES mouse models. We bring the proof of concept that our model is unique and allows a better understanding of the mechanisms that regulate gel-forming mucin production/secretion and mucous cell differentiation in the conjunctiva of living mice and can be used to test treatment compounds in mucosal disease models

MUC5B leads to aggressive behavior of breast cancer MCF7 cells.

The mucin MUC5B has a critical protective function in the normal lung, salivary glands, esophagus, and gallbladder, and has been reported to be aberrantly expressed in breast cancer, the second leading cause of cancer-related deaths among women worldwide. To understand better the implication of MUC5B in cancer pathogenesis, the luminal human breast cancer cell line MCF7 was transfected with a vector encoding a recombinant mini-mucin MUC5B and was then infected with a virus to deliver a short hairpin RNA to knock down the mini-mucin. The proliferative and invasive properties in Matrigel of MCF7 subclones and subpopulations were evaluated in vitro. A xenograft model was established by subcutaneous inoculation of MCF7 clones and subpopulations in SCID mice. Tumor growth was measured, and the tumors and metastases were assessed by histological and immunological analysis. The mini-mucin MUC5B promoted MCF7 cell proliferation and invasion in vitro. The xenograft experiments demonstrated that the mini-mucin promoted tumor growth and MCF7 cell dissemination. In conclusion, MUC5B expression is associated with aggressive behavior of MCF7 breast cancer cells. This study suggests that MUC5B may represent a good target for slowing tumor growth and metastasis

Différenciation des cellules à mucus et régulation de la mucine gélifiante Muc5b : un nouvel outil pour des études ex vivo et précliniques in vivo

International audienc

The Cervicovaginal Mucus Barrier

Preterm births are a global health priority that affects 15 million babies every year worldwide. There are no effective prognostic and therapeutic strategies relating to preterm delivery, but uterine infections appear to be a major cause. The vaginal epithelium is covered by the cervicovaginal mucus, which is essential to health because of its direct involvement in reproduction and functions as a selective barrier by sheltering the beneficial lactobacilli while helping to clear pathogens. During pregnancy, the cervical canal is sealed with a cervical mucus plug that prevents the vaginal flora from ascending toward the uterine compartment, which protects the fetus from pathogens. Abnormalities of the cervical mucus plug and bacterial vaginosis are associated with a higher risk of preterm delivery. This review addresses the current understanding of the cervicovaginal mucus and the cervical mucus plug and their interactions with the microbial communities in both the physiological state and bacterial vaginosis, with a focus on gel-forming mucins. We also review the current state of knowledge of gel-forming mucins contained in mouse cervicovaginal mucus and the mouse models used to study bacterial vaginosis

Ocular mucins in dry eye disease.

International audienceDry eye disease is a common and multifactorial disease with a high prevalence worldwide. Water loss, reduced expression of glycocalyx mucins, and loss of goblet cells secreting gel-forming mucins are hallmarks of dry eye disease. Mucins are large and complex heavily glycosylated proteins. Their organization in the tear film remains unclear, but they play a key role to protect and maintain integrity of the ocular surface. Mice have been extremely valuable mammalian models with which to study ocular physiology and disease, and to evaluate eye therapies. Genetically modified mice and spontaneously occurring mutants with eye defects have proven to be powerful tools for the pharmaceutical industry, clinicians, and basic researchers investigating dry eye disease. However, ocular mucins remain relatively under-studied and inadequately characterized. This review aims to summarize current knowledge about mucin production at the ocular surface in healthy individuals and in dry eye disease, and to compile an overview of mouse models available for the study of mucins in dry eye disease