Elucidating the regulation of lipid flippases required for vesicle biogenesis

Different MSc projects about flippases are suggested.

 

Elucidating the regulation of lipid flippases required for vesicle biogenesis

A fundamental feature of eukaryotic cells is their compartmentalization into distinct organelles by lipid membranes. This allows different functions to occur efficiently and simultaneously in different parts of the cell. Among these functions are nutrient and ion transport, oxidative and photosynthetic phosphorylation, signal transduction, and electrical excitability. To fulfil these various functions, cells tightly regulate vesicular traffic between cellular compartments. Specific membrane pumps, termed lipid flippases, play an essential role in this process but how do they work and how are they regulated? The objective of this project is to analyse the regulation of two plant lipid flippases, localized to different compartments, by the action of different kinases and phosphatases.

 

Applied techniques: split-ubiquitin analysis, site-directed mutagenesis, phosphorylation/dephosphorylation assays, membrane protein purification and co-immunoprecipitation. The work will be carried out at the PUMPKIN Centre of Excellence in the Faculty of Life Sciences. 
 

Tracking down interacting partners of plant lipid flippases

Lipid flippases are involved in vesicle formation in the secretory pathway. There role is fundamental for cell survival as the secretory pathway is involved in processes such as host-pathogen interactions, nutrient and gravity sensing, and protein sorting. As active components of the vesiculation machinery, these flippases, also known as P4-ATPases,  are expected to interact with several proteins involved in this process. The objective of this project is to identify interacting  partners for different P4-ATPases using a modification of the yeast-2-hybrid adapted to membrane proteins (the Split-ubiquitin system). In order to achieve this goal, a cDNA library will be generated and used for high-throughput screening. The most promising candidates will be subjected for further characterization.

 

Applied techniques: RNA isolation, cDNA library design and titration, yeast transformation, immunoprecipitation, Immobilized Metal Affinity Chromatography (IMAC), transient expression in tobacco leaf epidermal cells, bioimaging. The work will be carried out at the PUMPKIN Centre of Excellence in the Faculty of Life Sciences.