Date of Award

Winter 12-18-2015

Document Type

Thesis

Degree Name

Master of Science (MS)

Programs

Cancer Research

First Advisor

Hamid Band

Abstract

Endocytic trafficking is an essential process in eukaryotic cells, specifically for the transport of nutrients, membrane components, and receptors. Cargo destined for endocytic traffic is internalized at the cell surface via clathrin-dependent and clathrin-independent pathways, and brought to the early or sorting endosomes. From there, cargo is further trafficked to lysosomes for degradation, trafficked to other compartments in the cell, or recycled back to the cell surface (either directly or via the endocytic recycling compartment).

Mammalian C-terminal Eps15 homology domain-containing proteins, or EHD proteins (EHD1 to 4), are a family of highly conserved ATPases that function as key regulators of specific steps of the endocytic recycling process in metazoans. Research has shown that EHD proteins can oligomerize and bind to negatively charged membranes, which stimulates nucleotide hydrolysis. This is thought to assist in the process of membrane tubulation or the budding of vesicles, which is important in the movement of cellular components. Additionally, EHD proteins contain EH domains, which are known to bind to asparagine-proline-phenylalanine (NPF) motif-containing peptides, suggesting hundreds of potential interacting partners, albeit only a handful are thus far experimentally validated.

Recent findings have implicated EHD proteins in endothelial cells. Endothelial cells (ECs) form a barrier between blood and tissues, and play an essential role in many vital physiological functions including delivery of nutrients, angiogenesis, and innate and adaptive immune responses. Despite sharing a common purpose, ECs are structurally and functionally heterogeneous. Immunofluorescence staining of sections from multiple wild type mouse organs was carried out and the results support a conclusion of differential expression of EHD proteins within different endothelial beds. In vitro analysis of multiple cultured endothelial cell lines showed that differential EHD protein expression is lost as all four EHD proteins were expressed. Finally, siRNA-mediated knockdown of individual EHD proteins was found to affect tubulation of endothelial cells grown in reconstituted basement (Matrigel). These findings support a role for EHD proteins in endothelial cell biology.

Figure 1 Permission.pdf (135 kB)
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Figure 3 Permission.pdf (208 kB)
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