The compartmentation and distribution of metabolites between mitochondria and the rest of the cell is a key parameter of cell signalling and pathology. Here, we have developed a rapid fractionation procedure that enables us to take mouse heart and liver from in vivo and within ~ 30 s stabilise the distribution of metabolites between mitochondria an...
Mitochondrial transplantation is emerging as a novel cellular biotherapy to alleviate mitochondrial damage and dysfunction. Mitochondria play a crucial role in establishing cellular homeostasis and providing cell with the energy necessary to accomplish its function. Owing to its endosymbiotic origin, mitochondria share many features with their bact...
Cyanobacteria are complex prokaryotes, incorporating a Gram-negative cell wall and internal thylakoid membranes (TMs). However, localization of proteins within cyanobacterial cells is poorly understood. Using subcellular fractionation and quantitative proteomics, we produced an extensive subcellular proteome map of an entire cyanobacterial cell, id...
The study of protein localisation has greatly benefited from high-throughput methods utilising cellular fractionation and proteomic profiling. Hyperplexed Localisation of Organelle Proteins by Isotope Tagging (hyperLOPIT) is a well-established method in this area. It achieves high-resolution separation of organelles and subcellular compartments but...
Ultracentrifugation (UC) is recognized as a robust approach for the isolation of extracellular vesicles (EVs). However, recent studies have highlighted limitations of UC including low recovery efficiencies and aggregation of EVs that could impact downstream functional analyses. We tested the benefit of using a liquid cushion of iodixanol during UC ...
Published in Cardiovascular research
Extracellular vesicles (EVs)-particularly exosomes and microvesicles (MVs)-are attracting considerable interest in the cardiovascular field as the wide range of their functions is recognized. These capabilities include transporting regulatory molecules including different RNA species, lipids, and proteins through the extracellular space including b...
We previously developed a mass spectrometry-based method, Dynamic Organellar Maps, for the determination of protein subcellular localisation, and the identification of translocation events in comparative experiments. The use of metabolic labelling for quantification (SILAC) renders the method best suited to cells grown in culture. Here we have adap...
Chromatin structure plays a pivotal role in facilitating proper control of gene expression. Transcription factor (TF) binding of cis-elements is often associated with accessible chromatin regions. Therefore, the ability to identify these accessible regions throughout plant genomes will advance understanding of the relationship between TF binding, c...
Stable isotope labelling by amino acids in cell culture (SILAC) is a technique that allows proteomic profiling of cells. In this chapter we describe a protocol for the identification and quantification of newly synthesised proteins. The methodology can be applied to any cultured cell system with relevance to schizophrenia, affective disorders and a...
Stromules are stroma-containing tubules that have been observed to emanate from the main plastidic body in vivo. These structures have been shown to require cytoskeletal components for movement. Though numerous studies have shown a close association with the endoplasmic reticulum, nucleus, mitochondria, and other plastids, the mechanism of formatio...
