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

Xiaoxiao Li,Vanathi Gopalakrishnan,Jonathan Braun,David Elashoff,James Borneman,Bennett Roth,Sharon S Chen,Lee Goodglick,Laura L Presley,Jeff Allard,William L Bigbee,Thomas G Graeber,James Leblanc,Jonathan L Lustgarten,Andrew Ippoliti,Allison Truong,Ravi Vuthoori,Jörg Hermann Fritz

doi:10.1371/journal.pone.0026542

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.

Highlights

The intestinal mucosal surface plays diverse and critical roles in nutrient uptake, host defense, and local and systemic endocrinology [1,2,3,4]
The results showed that this new integrated sampling and analytical approach is capable of analyzing molecular compositions at different locations along the gastrointestinal tract, and provides a new dimension to the characterization of host-microbial interaction at the human mucosal-luminal interface (MLI)
To study the mucosal luminal interface, we established a novel protocol to directly examine this environment using samples obtained by endoscopic lavage

Summary

Introduction

The intestinal mucosal surface plays diverse and critical roles in nutrient uptake, host defense, and local and systemic endocrinology [1,2,3,4]. Functional assessment of the microbiome has recently emphasized metagenomic and biochemical analysis, uncovering commonalities in metabolic and other traits responsive to diet, and reciprocal interactions with host physiology [10,11,12,13]. Far, such studies have almost exclusively focused on the biology of the fecal compartment, and there have been only limited assessments of the mucosal surface itself. There are few strategies available to directly analyze the function of the MLI in humans as an integrated hostcommensal biologic unit

Methods

Results

Conclusion