A Metaproteomic Approach to Study Human-Microbial Ecosystems at the Mucosal Luminal Interface

Abstract

Aberrant interactions between the host and the intestinal bacteria are thought to contribute to the pathogenesis of many digestive diseases. However, studying the complex ecosystem at the human mucosal-luminal interface (MLI) is challenging and requires an integrative systems biology approach. Therefore, we developed a novel method integrating lavage sampling of the human mucosal surface, high-throughput proteomics, and a unique suite of bioinformatic and statistical analyses. Shotgun proteomic analysis of secreted proteins recovered from the MLI confirmed the presence of both human and bacterial components. To profile the MLI metaproteome, we collected 205 mucosal lavage samples from 38 healthy subjects, and subjected them to high-throughput proteomics. The spectral data were subjected to a rigorous data processing pipeline to optimize suitability for quantitation and analysis, and then were evaluated using a set of biostatistical tools. Compared to the mucosal transcriptome, the MLI metaproteome was enriched for extracellular proteins involved in response to stimulus and immune system processes. Analysis of the metaproteome revealed significant individual-related as well as anatomic region-related (biogeographic) features. Quantitative shotgun proteomics established the identity and confirmed the biogeographic association of 49 proteins (including 3 functional protein networks) demarcating the proximal and distal colon. This robust and integrated proteomic approach is thus effective for identifying functional features of the human mucosal ecosystem, and a fresh understanding of the basic biology and disease processes at the MLI.

Colorectal cancer: a tale of two sides or a continuum?: Figure 1

“…Luminal contents, including gut microbial communities and their metabolites, might trigger initiating molecular events or, alternatively, influence the tumour microenvironment and promote neoplastic progression. Microbial and biochemical components of the luminal ecosystem and interactions occurring within the mucosal–luminal interface are likely to vary gradually along the proximal–distal axis of the colorectum 49 50. Indeed, this underlying continuum was postulated by LaPointe and colleagues to be responsible for the apparent continuity observed in their gene expression data 43…”

Microbiomics: An Approach to Community Microbiology

“…The most exciting information from WMS sequencing has come from studies of the human gut microbiome and its role in obesity, energy utilization, and physiological disorders (Turnbaugh and Gordon 2008 ). Similar to metagenomes of the human gut, WMS sequencing, human stool metatranscriptomics, and medium-throughput human gut metaproteomics has provided important information on the relationship between the human body and its microbiota (Poretsky et al 2009 ;Shi et al 2009 ;Li et al 2011 ;Booijink et al 2010 ;Giannoukos et al 2012 ). This technique is good for any heterogeneous population, and the low frequency in population and biasness of the 16S rRNAsequencing methods has no infl uence (Sogin et al 2006 ).…”

“…'Metatranscriptomics'-a fi eld very similar to metagenomics-uses sequences of reversetranscribed RNAs; extending this up to the quantifi cation of protein level is 'metaproteomics', while 'metabolomics' deals with the investigation of small metabolites (Turnbaugh and Gordon 2008 ;Verberkmoes et al 2009 ;Li et al 2011 ). These fi elds are in the very early stages of development and use technologies that are almost similar to those used by their cultured counterparts.…”

Insights from quantitative metaproteomics and protein-stable isotope probing into microbial ecology

“…Feces analyses revealed only limited insight to the interface between host and microbiota. Thus, two studies focussed on the mucosa-associated microbiota in human and rats (Li et al, 2011;Haange et al, 2012). Representative, Haange et al (2012) showed phylogenetic and functional differences along the intestinal tract (mucosa and gut content) of rats.…”