Unearthing the molecular mechanisms that govern L-selectin-dependent adhesion and migration

Student thesis: Doctoral ThesisDoctor of Philosophy


L-selectin has been well characterised as a cell adhesion molecule, which plays a role in the recruitment of leukocytes to sites of inflammation and is responsible for the recirculation of lymphocytes to secondary lymphoid organs. Recent evidence has shown that L-selectin also acts as a signalling molecule to activate pathways and regulate the inflammatory response. The cytosolic tail of L-selectin plays a crucial role in regulating its activity through its interaction with binding partners, such as calmodulin (CaM) and the ERM protein family. However, little is known about how the interaction between L-selectin and its binding partners is regulated. The aim of this multidisciplinary PhD project is to use biophysical and cell biological methods to address the role of the interaction between L-selectin and its binding partners during leukocyte recruitment. To this end, the interaction between CaM and the L-selectin cytosolic tail was assessed using isothermal titration calorimetry (ITC) and nuclear magnetic resonance (NMR) spectroscopy. Analysis revealed that phosphorylation of serine residues within the cytosolic tail of L-selectin did not affect CaM binding. To enable the observation of the interaction between L-selectin and CaM whilst leukocytes are undergoing transendothelial migration (TEM), the THP-1 monocytic cell line was engineered to stably express L-selectin-GFP and CaM-RFP so their interaction could be monitored at different stages of TEM. The data showed that phosphorylation of serine 364 in the L-selectin tail is important for regulating CaM interaction. Discrepancies were identified between the biophysical and cell biological results, implying the leukocyte plasma membrane may play a vital role in regulating the interaction between L-selectin and CaM. This highlights the importance of studying transmembrane proteins in the correct context.
Date of Award2015
Original languageEnglish
Awarding Institution
  • King's College London
SupervisorAlex Ivetic (Supervisor) & Maria Conte (Supervisor)

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