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Ecophysiology of membrane lipid remodelling in marine bacteria

Periodic Reporting for period 3 - EcoLipid (Ecophysiology of membrane lipid remodelling in marine bacteria)

Reporting period: 2020-04-01 to 2021-09-30

Membrane lipids form the structural basis of all cells. In bacteria Escherichia coli uses predominantly phosphorus-containing lipids (phospholipids) in its cell envelope, including phosphatidylethanolamine and phosphatidylglycerol. However, beyond E. coli a range of lipids are found in bacterial membranes, including phospholipids as well as phosphorus (P)-free lipids such as betaine lipids, ornithine lipids, sulfolipids and glycolipids. In the marine environment, it is well established that P availability significantly affects lipid composition in the phytoplankton, whereby non-P sulfur-containing lipids are used to substitute phospholipids in response to P stress. This remodeling offers a significant competitive advantage for these organisms, allowing them to adapt to oligotrophic environments low in P. Until very recently, abundant marine heterotrophic bacteria were thought to lack the capacity for lipid remodelling in response to P deficiency. However, recent work by myself and others has now demonstrated that lipid remodelling occurs in many ecologically important marine heterotrophs, such as the SAR11 and Roseobacter clades, which are not only numerically abundant in marine waters but also crucial players in the biogeochemical cycling of key elements. However, the ecological and physiological consequences of lipid remodeling, in response to nutrient limitation, remain unknown. This project aims to provide a mechanistic understanding of the physiological response of lipid remodelling to nutrient limitation using model marine bacteria. It seems to bridge the gap in our understanding of lipid remodeling and its potential biological and ecological consequences. The project aims to use a synthesis of molecular biology, microbial physiology, and "omics" approaches to reveal the fitness trade-offs of lipid remodelling in cosmopolitan marine heterotrophic bacteria, providing novel insights into the ecophysiology of lipid remodelling and its consequences for marine nutrient cycling.
The project is further divided into three main work packages. Progress to date includes
1) comparative lipidomics experiments using Ruegerial pomeroyi as the model and the identification of the novel glutamine-containing lipids in this bacterium and its role in the adaptation to phosphorus stress. This work has recently been accepted in publication in the ISME Journal (Smith et al., 2019).
2) Mechanistic insight into lipid remodelling. We have successfully generated several mutants, including plcP, olsAB, glsB which will be used in this work package. We have recently obtained a novel mutant involved in the production of a new homotaurin containing lipids in marine roseobacteris. This new lipid has not been reported previously in any live organisms. This manuscript for reporting the identification is currently under preparation to be submitted to ISME Journal. We have also successfully tested the methodology of isotope labelling using radio-active compounds. Furthermore, we have also purified and biochemically characterised the key enzyme PlcP which is essential in lipid remodelling. this work has recently been published in Applied and Environmental Microbiology (Wei et al., 2018).
3) We have successfully isolated several bacterial phages which infect our model bacteria. This will be used to understand the role of remodelling in phage interaction. We have tested infection dynamics of phage against our model bacterium DSS3 and analysed lipidomics changes and proteomics changes between wild-type and mutants. Two of the phages infecting Roseobacter pomeryo DSS-3 have unexpected DNA substitution from thymidine by deoxyuridine (dU) and do not resemble anything else in the current bacteriophage genome database. We have established that they form a novel family of bacterial virus. This student is currently under review for publication (Branko et al.)
4) We have successfully obtained several eukaryotes and managed to maintain and grow them successfully in the laboratory. We are currently investigating the interaction between wild type Roseobacter strains and the lipid remodelling mutants.
The project aims to bridge a knowledge gap in our understanding of lipid remodelling and its potential biological and ecological consequences. We have made significant progress in all work packages and established methods and protocols during this reporting period in order to test our hypotheses. The project is therefore on track to transform our understanding of the ecophysiology of adaptation to nutrient limitation in model marine bacteria and open up a whole new horizon for future investigation of the role in nature systems.