Year: 2022
Type: dissertation
Abstract: The internalisation of different cell surface receptors, their ligands, macromolecules, and pathogens from the extracellular space is mediated by a highly regulated process called endocytosis. The fate of internalised cargoes is either to be trafficked to different subcellular locations (such as the trans-Golgi network or recycled back to the plasma membrane) or the cargo is directed to be degraded in the lysosome. Issues in endocytosis have been implicated in the development of neurodegenerative diseases, and cardiovascular and inflammatory diseases (several types of cancer). The cargo sorting occurs in the endosomal network. The early endosome is the first step in the sorting process. The early endosome recruits several sorting complexes namely retromer, commander and WASH complex to facilitate the proper trafficking of cargo. The sorting complexes such as WASH and retromer complex are known to interact with each other and influence each other’s endosomal localisation. The sorting complexes are also known to interact with other membrane organisers such as Rab GTPases like RAB21. RAB21 is previously shown to influence the cargo sorting process of specific cargoes based on recent work from Pr Jean laboratory. The primary purpose of this project is to understand the interrelationship between the different endosomal sorting regulators such as retromer, WASH complex and RAB21 in the cargo sorting process. The KOs of specific sorting regulator proteins such as FAM21(WASH) or VPS29(Retromer) along with RAB21 depletions were used in combination with immunofluorescence to assess the loss of sorting regulators. An investigation into the influence of RAB21 activation on WASH-mediated endosomal F-actin polymerisation crucial for cargo sorting was also performed. My work showed that the VPS29-depleted cells show a complete loss of endosomal FAM21. The observations of VPS29-depleted cells showed similar results obtained from the KOs of FAM21 and FAM21+RAB21. We used RAB21-depleted cells and rescued them with wildtype RAB21 (eGFP:RAB21-WT). These experiments suggested an increase in endosomal F-actin levels. My studies furthered the understanding of the intricate relationship between the different endosomal sorting regulators and could provide additional lines of investigation to understand the endosomal sorting of specific cargoes.
Author: Maran, Suveda Vignesh
Language: English
Subfield: Cell Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W7201924290