Date of Award

2021

Document Type

Dissertation - MUSC Only

Degree Name

Doctor of Philosophy (PhD)

Department

Molecular and Cellular Biology and Pathobiology

College

College of Graduate Studies

First Advisor

Steven Carroll

Second Advisor

Carl Atkinson

Third Advisor

Amanda LaRue

Fourth Advisor

Robin C. Muise-Helmericks

Fifth Advisor

John O'Bryan

Abstract

In patients with the genetic disorder Neurofibromatosis type 1 (NF1), the loss of the Ras GTPase-activating tumor suppressor protein neurofibromin (also NF1) promotes the development of multiple nervous system tumors and other clinical manifestations. Neurofibromin loss potentially results in hyperactivation of both classic Ras (H-Ras, N-Ras, K-Ras) and non-classical Ras (R-Ras, R- Ras2/TC21, R-Ras3/M-Ras) subfamilies of proteins, thus promoting transformation and malignancy. Malignant Peripheral Nerve Sheath Tumors (MPNSTs) are often associated with NF1 loss and we have previously shown that the resulting hyperactivation of classic Ras proteins contribute to MPNST proliferation and survival. However, the roles of R-Ras subfamily proteins in MPNSTs have not previously been elucidated. We found that R-Ras and R-Ras2 are expressed and functional in neurofibromin-null MPNST cells. We discovered that R-Ras mediated signaling promotes MPNST mitogenesis, migration, and an invasive growth phenotype. Using a dominant negative (DN) R-Ras expression vector and mass spectrometry-based phosphoproteomics, we identified thirteen R-Ras specific regulatory networks. The most prominently affected networks involved cellular movement. We validated a key migration and cytoskeletal organization regulator, ROCK1, as an essential effector molecule downstream of activated R-Ras in MPNSTs that could be therapeutically targeted. Due to the drug resistant nature of MPNSTs, we sought to identify a combinatorial regimen by identifying a novel target upstream of R-Ras. Using a global approach, we identified the LPA receptors as upstream mediators of R- Ras signaling. We discovered that LPA functions in a receptor dependent manner, with LPAR1 working through R-Ras and ROCK1 to specifically promote migration while LPAR3 specifically promotes proliferation of MPNST cells. We conclude that R-Ras activated proteins promote tumorigenesis via signaling pathways distinct from the more-studied classic Ras proteins in MPNSTs. In this dissertation, we show that LPA and R-Ras orchestrate mechanisms regulating proliferation, invasion, and migration in MPNSTs thus providing potential novel therapeutic targets for patients with MPNSTs.

Rights

All rights reserved. Copyright is held by the author.

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