A metaproteomic analysis of microbial communities of Ace Lake, Antarctica

摘要

The Vestfold Hills is an ice free oasis in East Antarctica with possibly the highestdistribution of stratified lakes and marine basins in the world. Ace Lake is ameromictic lake located on the Long Peninsula in the Vestfold Hills region, and isone of the most intensively studied stratified lakes in Antarctica. Since itsisolation from the Southern Ocean over 10,000 years ago, the marine derivedcommunity of Ace Lake have evolved with changes in the physical and chemicalstructure of the lake. Metaproteomics (environmental proteomics) was used toinvestigate the interactions and functional activity of microbial populations withinthe layers of Ace Lake. To achieve this, a protein extraction procedure wasdeveloped using test samples of filtered biomass from coastal marine waters(Botany Bay). The performance of two mass spectrometry analysis softwares,Mascot (spectral searching) and Peaks (de novo sequencing) were also evaluatedusing the metaproteomic datasets generated from Botany Bay. Although it wasconcluded that Mascot was a more rapid and reliable method for processing largemetaproteomic datasets, the de novo sequencing capability of Peaks hold muchpromise for identification of proteins with novel sequences. The simulated mixedcommunity study of Sphingopyxis alaskensis and Escherichia coli indicated thatwell represented organisms in terms of cell abundance resulted in higher levels ofprotein detection by mass spectrometry. The simulated community study providedmetaproteomic datasets, which enabled the performance of Mascot databasesearching and protein identification validation techniques to be tested.Cross-species matching (NR database) of proteomic data derived from microbialpopulations at different depths of Ace Lake provided insights to th e microbialcomposition and functional activity of members within the lake. A SAR11community in the surface waters of Ace Lake likely adapts to oligotrophicconditions of the lake by expressing transport proteins associated with nutrientuptake. The detection of Phycodnaviridae capsid proteins in the aerobic zonesuggests that these algal viruses may be active in the Antarctic summer duringperiods of algal bloom. Proteins involved in processes such as methanogenesis,sulfur and carbon cycling were also identified.Through metagenomics, an almost complete genome sequence of a single,dominant ass (C-Ace) was assembled from shotgun sequencing data taken fromthe oxycline (12.7 m) of the lake. Approximately 34 Mb of D A sequence datawas assembled into nine scaffolds totalling 1.79 Mb, representing a 19-foldcoverage for the C-Ace composite genome. Using a metaproteogenomic approach,metaproteomic data obtained from biomass at the chemocline was mapped back tothe refe rence C-Ace genome sequence data, which resulted in high levels (~ 31 %)of metaproteomic coverage. The findings of this study revealed the physiologicaltraits that promote the ass to compete very effectively to gain dominance undercold, nutrient-, oxygen-limited and extremely varied annual light cycles.Metaproteomics is a relatively new and rapidly emerging area of study in the fieldof environmental microbiology. An increasing number of communityproteogenomic analyses over the past few years have successfully unravelled thestructure, function, and microbial diversity of a range of different ecosystems (e.g.aquatic, soil biospheres, biofilms). Ace Lake is one of the most studiedmeromictic lakes in Antarctic, however few studies have attempted to provide acomprehensive analysis of the identity and functional activity of the microbialpopulations within the stratified layers of the lake. The metaproteomic analysispresented in this thesis has provided a broad survey of the microbial communitystructure and a glimpse into the biochemical processes occurring at the variousdepths throughout the water column. The role of the dominant ass (C-Ace) as amajor player in nutrient cycling and primary productivity was established througha detailed analysis of the metaproteogenomic data, which also provided insightsinto microbial interact ions and adaptive responses for survival in the Antarcticenvironment. Studying microbial communities ill situ is challenging hence thesuccessful application of methods developed in this metaproteomic study ofmicrobial communities from Ace Lake is valuable as it demonstrates the potentialof applying the same approach to other Antarctic aquatic environments.