The molecular basis of bacteria adhesion to gastrointestinal mucus

The human body is colonized by a vast number of microbes, most of them are present in our gut where they are collectively referred to as the human gut microbiota. The microbiota contains approximately 10-100 trillion bacteria belonging to 15,000~36,000 species with the greatest density populating the colon where they reach 1.5 kg. In fact, the microbes that we carry around outnumber our own cells by about 10-fold and collectively they have about 100-fold more genes than we do. The gut microbiota is required for the development and maintenance of human health. These bacteria help digest our food, produce nutrients, detoxify dangerous substances, protect us from harmful bacteria (pathogens) and help with the development of our immune system. However, the microbiota is not innocuous, and under conditions that compromise our ability to limit the microbiota's entry from the intestine, bacteria species can invade the body to cause disease. Furthermore, shifts in the composition of the microbiota, referred to as dysbiosis, have been linked to inflammatory bowel diseases and are also increasingly associated to a number of diseases outside the gut. There is currently no deep understanding of what triggers these changes in the microbiota. However we are starting to unravel the mechanisms that allow the majority of the bacteria to live in peaceful coexistence within our gut.
Researchers recently showed that the protective mucus layer covering cells lining the gut plays a crucial role in the maintenance of the microbiota. Mucus is produced in large amounts in the colon where most of our gut bacteria are present. Its organisation is crucial to its protective function; it is divided into a dense layer which prevents the bacteria to penetrate into our body (thus protects us against a possible invasion) and a loose layer above it which provides a home for our gut bacteria (so that we can still benefit from their protective activities without the associated risk of an invasion). This system is based on the arrangement of large proteins called mucins which contain a very complex array of sugars. Mucus also harbours a large proportion of antibodies which reinforce the confinement of our gut bacteria into the gut. It is thought that the sugars present in mucins provide an attachment site for the bacteria that help maintain normal gut function. However these hypotheses remain to be tested.
Our Group recently showed that some of the bacteria that live in the gut have mucus-binding proteins (MUB) on their surfaces which help them bind to the mucus layer. However we do not know what exactly they recognise in the mucus and how this may influence health. An important aspect of this work will be to identify the structures MUB bind to and how. Sugars are complex to analyze therefore their precise role and importance in biological systems has eluded us for many years. Recent technological advances will help us identify which mucin sugars are involved in the interaction. Complementary biochemical analyses will provide further insights into the specificity and strength of binding presented by the multiple protein units constituting MUB. Using crystallography and mutagenesis we will also determine the precise amino acids involved in the interaction with mucins and antibodies, this will help understand differences in the way harmful or protective bacteria interact with the gut. We will expand this in vitro work to intestinal cell models to study the interaction of MUB purified from the bacteria and of bacteria harbouring MUB in a biologically relevant system. We will determine the consequences of the association with antibodies to the adhesion of bacteria to mucus and how this may change the way the intestinal cells respond to bacteria. Results from this work will help us understand how to keep a beneficial relationship with our gut bacteria and may lead to the development of novel strategies to readjust microbial community or prevent dysbiosis.

The mammalian gastrointestinal (GI) tract is home to complex bacterial communities. Most of the bacteria establish beneficial symbiotic relationships with their host, making important contributions to metabolism and digestive efficiency. The vast numbers of bacteria in the GI tract and their proximity to host tissues raise the question of how homeostatic host-bacterial relationships are established and maintained without eliciting potentially harmful immune responses. The mucus barrier, antimicrobial proteins and IgA operate together to confine symbiotic bacteria towards the outer mucus layer and protect the epithelial surface by limiting bacterial penetration. This homeostatic interaction between mucus and gut commensals is critical to maintain gut health. Intestinal pathogens have evolved strategies for subverting and evading these barriers to access host tissues. By contrast, there is currently very limited knowledge of the mechanisms by which resident members of the intestinal microbiota interact with components of the mucus barrier. Mucus-binding proteins (MUBs) are a class of cell-surface adhesins found in Gram-positive gut symbionts, shown to mediate interaction of bacteria to mucus. Building on our extensive preliminary data on the structural and biochemical properties of MUB, the purpose of this study is to determine the molecular and cellular basis of MUB interactions with receptors of the mucus layer, mucins glycans and mucosal antibodies, and assess the potential of the impact of MUB and IgA interaction on bacteria adhesion to mucus and cell response (focusing on changes in mucin, cytokine, polymeric Ig receptor expression), providing mechanistic insights into the regulation of gut microbiota homeostasis. Understanding the molecular basis of these processes will help understand how some bacteria strains exert intestinal health protection and devise intervention strategies to reinforce gut health.

The research fits with the BBSRC's Bioscience Underpinning Health priority and its Grand Challenge 3 - Fundamental bioscience enhancing lives and improving wellbeing (BBSRC Delivery Plan 2011-15). The research has the potential to impact on the nation's health and welfare through reducing the onset and progression of gut-associated diseases. Inappropriate host-microbial interactions initiate the dysregulation and disruption of the intestinal immune balance, which result in inflammatory and metabolic disorders including IBD (Crohn's disease and ulcerative colitis) and obesity. IBD in particular involves a break of tolerance to the commensal microbiota. The mucus layer is very important for maintaining homeostasis and mucus defects can allow bacteria to reach the epithelium and trigger inflammation. This protective effect involves homeostatic interactions of the bacteria with the mucosal IgA system and mucins. A better insight into the biochemical nature of these interactions is fundamental for understanding how this protective function is achieved and can be maintained throughout life. The benefits for industry are not immediate or direct. However, in the age of extensive microbiota sequencing where an ever-increasing number of bacteria genomes and metagenomes are being unravelled, the prospect of accessing our individual microbiota profile as part of a routine health check is becoming closer and the need to further our understanding of the fundamental mechanisms involved in the interaction of the microbiome with the host is thus urgently required in order to capitalise on these technical advances and be able to translate findings into biomarkers of health and disease.

IBD affects 240,000 people in the UK. Education, employment, social and family life are all disrupted by the unpredictable occurrence of the disease. The symptoms of IBD can severely affect social functioning, particularly among young and newly-diagnosed individuals. Patients with severe IBD diseases may develop potentially life-threatening complications and have an increased risk of colorectal cancer. The cost of IBD to the NHS is about £720 million/year. Expensive drugs to suppress the immune system account for about a quarter of total health service cost for IBD and half of the healthcare costs relates to the inpatient management of a minority of patients who need intensive medical or surgical intervention. There is a significant morbidity and mortality in young adults, which has a major adverse impact on the potential economic contribution of this demographic. It is anticipated that within 10-15 years, the detailed understanding of gut bacteria-host molecular interactions such as those described in this proposal will be used by pharmaceutical/food companies to address the causes of dysbiosis and develop routes to restore microbial balance. Such preventive treatments will significantly reduce NHS costs, and improve the health of the nation. In the shorter term, the information generated in this proposal will facilitate the rational selection or engineering of probiotic strains. Furthermore, the innate binding specificity of the glycan binding protein under study may offer a versatile approach for mapping mucin glycosylation in human health as well as a potential diagnostic marker.

The work will be published in internationally leading peer-reviewed journals. Results will be presented by the PI and PDRA at national and international conferences, public lectures and meetings with academic collaborators. This will be extended to the popular press when appropriate. Local schools and interested public will benefit from the proposed outreach activities. The PDRA will benefit from training from PI and collaborators who have expertise and interests in glycobiology, microbiology and immunology. The PDRA will also acquire transferable skills both from working on the project and through attending courses e.g. project management, public engagement and business.