Neurofibromatosis type 2, an autosomal dominant genetic disorder, causes predisposed individuals to develop various benign central and peripheral nervous system tumors. The characteristic tumors of this disease are schwannomas, which are tumors of the Schwann cells, typically on the vestibular nerve. These and the other associated tumors slowly compress nervous system structures causing deafness and loss of balance, resulting in an average life-span of less than 40 years. The product of the Nf2 gene is the protein named merlin or schwannomin. In individuals diagnosed with NF2, merlin is either absent or mutated to the point of inactivation. As such, merlin functions as a negative growth regulator in that it suppresses tumor growth. Being that NF2 is predominately a disease of the Schwann cells, merlin's functional role within the signal transduction pathways of Schwann cell growth and differentiation are being investigated. This thesis explores the molecular relationships between merlin and its various interactors within Schwann cells, and illuminates one step in elucidating merlin's functional mechanism of action. Merlin has been shown to associate with paxillin in a density-dependant manner and to bind directly to paxillin through two specific paxillin binding domains. Individual paxillin LD domain fusion proteins were produced, as well as recombinant merlin lacking the paxillin binding domains. Direct binding assays were performed in order to determine which specific paxillin domains merlin might interact with directly. The results indicate that, in vitro, merlin binds, through its PBD1 domain, to the paxillin LD3 motif. Supporting this data, the results also demonstrate that when the merlin PBD1 domain is deleted, merlin binding to paxillin LD3 is abrogated. The direct binding shown here between paxillin and merlin, coupled with research demonstrating that merlin is present in β1 integrin immuno-precipitations, leads to the question of whether merlin binds directly to β1 integrin or associates with β1 integrin through paxillin. Using direct binding assays, this research shows that the merlin C-terminus binds directly to the cytoplasmic domain of β1 integrin, in vitro. Lastly, since merlin is an ERM family protein and has been shown to dimerize with ezrin (another ERM family member), and because merlin has been shown to bind directly to paxillin, the question asked is whether paxillin can interact directly with ezrin. The results indicate that paxillin can bind directly to the N-terminus of ezrin, in vitro. The findings presented here, when examined together, provide a framework for the proposal of a model in which paxillin LD3 mediates the localization of merlin to the plasma membrane, where it associates with the β1 integrin cytoplasmic domain and ezrin. These results and the proposed model offer additional insight into the mechanism of action of merlin's negative growth regulating function in Schwann cells.
Master of Science (M.S.)
Burnett College of Biomedical Sciences
Molecular Biology and Microbiology
Length of Campus-only Access
Masters Thesis (Open Access)
Geden, Sandra E. Beattie, "A Novel Binding Interaction For The Paxillin Ld3 Motif: Paxillin Ld3 Mediates Merlin-Paxillin Binding At Paxillin Binding Domain 1" (2005). Electronic Theses and Dissertations. 6099.