טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentLiani-Gura Esti
SubjectUbiquitylation of Synphilin-1 and its Role in Parkinson's
Disease
DepartmentDepartment of Medicine
Supervisor Professor Simone Engelender
Full Thesis textFull thesis text - English Version


Abstract

Parkinson’s disease (PD) is the second most common neurodegenerative disorders in humans, characterized by loss of dopaminergic neurons in the substantia nigra and the presence of Lewy bodies in the surviving neurons. Although the etiology of PD is not well understood, the discovery of genes responsible for familial forms of the disease provided insights into its pathogenesis.   

Mutations in the a-synuclein gene and its presence in Lewy bodies of PD patients indicate that a-synuclein plays an important role in the disease. We have previously found that a-synuclein interacts with a protein called synphilin-1. Synphilin-1 interacts with a-synuclein in vivo and is present in Lewy bodies of sporadic PD patients. Co-transfection of synphilin-1 with a-synuclein into mammalian cells leads to the formation of eosinophilic inclusions that resemble Lewy bodies, suggesting that synphilin-1 could modulate a-synuclein aggregation.

            Although synphilin-1 is a neuronal protein that is localized to presynaptic nerve terminals and associates in vivo with synaptic vesicles, its function remains largely unknown. In an attempt to dissect synphilin-1 function, we sought to identify additional proteins that interact with synphilin-1. Utilizing the Yeast Two-Hybrid technique, we found that synphilin-1 interacts with the E3 ubiquitin-ligases SIAH-1 and SIAH-2. SIAH proteins promote the ubiquitylation and degradation of synphilin-1 through the ubiquitin-proteasome system, and the formation of a large amount of inclusion bodies in the presence of proteasome inhibitors. Ubiquitylation is required for inclusion formation, since a catalytically inactive mutant of SIAH-1, which still binds to synphilin-1, fails to promote inclusions. Synphilin-1/SIAH inclusions are ubiquitin-positive and can recruit a-synuclein, suggesting that they may be relevant to the PD pathology. We also found that, like synphilin-1, a-synuclein associates with SIAH in cells, but the interaction with SIAH-2 was stronger than with SIAH-1. In vitro experiments show that SIAH-2 monoubiquitylates a-synuclein. Further evidence that SIAH may play a role in inclusion formation comes from the demonstration of SIAH immunoreactivity in Lewy bodies of PD patients. We hypothesize that dysfunction of ubiquitin-proteasome pathway and accumulation of ubiquitylated synphilin-1 and a-synuclein could be an early event in Lewy body formation and in the pathogenesis of PD.