Project Name: Regulation of Cell Polarity and Tumor Suppression by Dynamic Palmitoylation
Most cancers arise when epithelial cells, which make up the tissues that cover and line the body, undergo a malignant transformation. This transformation, called "epithelial-mesenchymal transition" or EMT, causes cells to grow in excess, detach from their normal environments, acquire invasive, migratory capabilities, and eventually colonize or metastasize to other regions of the body. Epithelial cells normally have a defined structure that is lost in EMT. My work aims to understand how certain proteins in the body maintain epithelial cell structure and therefore protect against EMT.
More specifically, I am interested the function of these proteins, called tumor suppressors, is influenced by chemical modifications. We recently used mass spectrometry and proteomics to compile a list of proteins that undergo modifications by fatty acids in an aggressive lymphoma cell line. Mass spectrometry is an analytical technique that determines the molecular composition of a sample, including all of the proteins present in the sample. The compilation of proteins is referred to as the proteome, and the study of these sets of proteins is called proteomics. We can use these techniques to compare the compositions of proteins in different samples in order to identify changes. One of the interesting proteins that appeared to be modified by fatty acids in the screen was the tumor suppressor Scribble (Scrib), which serves as a gatekeeper of cell structure.
Scrib is a large protein that serves as a "scaffold;" much like the foundation of a home, Scrib brings together many other proteins, connecting them together so they can serve a specific structural role. Scrib is a critical architectural element for epithelial cells. Interestingly, in order to serve its role, Scrib must be present at the cell membrane. When epithelial cells undergo EMT, Scrib appears to become increasingly diffused within the intracellular space, rather than remaining on the cell membrane, and can no longer fulfill its role in connecting proteins key to normal cell structure. This tells us that Scrib can only act as a tumor suppressor when it is on the membrane. We hypothesize that Scrib's presence on the cell membrane is due in large part to the fatty acid modifications that we previously identified through our screening, and that its diffusion to the intracellular space in EMT is due in part to hyperactive enzymes known as thioesterases, which we believe remove fatty acids from Scrib.
I am currently working on three independent projects. The goals of each project are:
1) Screen for proteins undergoing fatty acid modifications in cellular models of EMT. I will use proteomics to identify proteins that have changed levels in EMT, as well as thioesterase enzymes that display either heightened or diminished activity in EMT. This will help us understand the mechanism that forces Scrib from the membrane and will guide our future work to develop drugs to target these enzymes.
2) Identify the molecular mechanism of Scrib's presence at the membrane by directly assessing the fatty acid dynamics of Scrib. The biggest mystery in the field is why Scrib attaches and detaches from the cell membrane according to cellular malignancy. To test my hypothesis that the diffusion of Scrib to the intracellular space is due to a loss of the chemical modifications by fatty acids, I will manipulate the Scrib protein where I believe the modifications take place. This will prevent the fatty acids from attaching, and allow me to assess whether Scribâ€™s localization changes at all, potentially identifying the mechanism which promotes Scrib presence at the membrane.
3) Evaluate drugs that inhibit thioesterase enzymes as a therapeutic approach for restoring Scrib localization and tumor suppression activity. Thioesterases remove fatty acids from proteins. We hypothesize that these enzymes are hyperactive in EMT, therefore removing fatty acids from Scrib and promoting diffusion away from the membrane and toward the intracellular space. We previously identified small molecules that specifically target two thioesterase enzymes, and found that treating EMT cells with these drugs partially restores Scrib to the cell membrane. I will continue to evaluate these drugs in other EMT lines and assess the status of fatty acid binding to Scrib, localization of Scrib after treatment with drugs, and determine whether the drugs can diminish the aggressive nature of EMT cells.
Summer 2014 Update
In May 2014, MCRF Trustees had a very engaging tour of the Department of Chemistry lab where Jeannie and her colleagues conduct their research. During this tour, Trustees and guests had the opportunity to observe cells undergoing manipulation, learn about specialized tools and equipment essential to the research process, and even had some hands-on participation in lab experiments.
Summer 2016 Update
Jeannie and her colleagues at the University of Michigan Department of Chemistry were pleased to publish their findings in an article in the Royal Society of Chemistry’s journal Molecular BioSystems in May 2016.