Defining the molecular action of retromer in retrograde trafficking pathways
The endo-lysosomal network is a highly dynamic and orchestrated system which serves a vital function in coordinating the communication and the exchange of cargo molecules between intracellular membrane-bound compartments. The endo-lysosomal system is critical for the maintenance of intracellular homeostasis, and defects on it are often associated with neurological disorders. Retromer is a peripheral membrane protein complex that coordinates multiple vesicular trafficking events. However, the
... ts. However, the molecular actions of retromer in the endo-lysosomal system remain unclear and controversial. This thesis is focused on the characterization of retromer's molecular action in the retrograde trafficking of endosome transport carriers (ETCs) and dissects the functional association between retromer and its accessory proteins. Firstly, this study demonstrates a selective function of retromer in the retrograde sorting via ETCs. The incorporation of CI-M6PR, via a retromer-dependent method, into ETCs, is restricted to those captured by the trans-Golgi protein GCC88, but not other trans-golgins including golgin-97 or golgin-245. This retromerdependent retrograde trafficking pathway requires SNX3, but not other retromer-associated cargo binding proteins, such as SNX-BAR proteins and SNX27. Besides, this study shows the requirement of retromer in the maintenance of lysosomal morphology and function. The absence of retromer alters the lysosomal ultrastructure, impairs the autophagic process and the lysosomal proteolysis. Secondly, this study demonstrates that GCC88, the tethering factor in retromer-dependent trafficking pathway is required for the endosome-to-TGN retrieval of CI-M6PR and the maintenance of lysosomal proteolytic activity. However, GCC88 deficiency has no impact on the autophagy-lysosomal pathway. Lastly, the final chapter of this thesis demonstrates the Parkinson's disease-linked retromer variant -Vps35 D620N confers a partial loss of function. The presence of the Vps35 D620N variant rescues the lysosomal defects and the endosomal dissociation of TBC1D5 and the WASH complex subunit FAM21, which are caused by the absence of retromer. However, the Vps35 D620N variant fails to rescue the trafficking defects of retromer-dependent GCC88-captured ETCs. Publications during candidature Cui, Y., Yang, Z., Teasdale, R. 2017. The functional roles of retromer in Parkinson's disease.