Content area
Abstract
FE65 is a brain-enriched, developmentally regulated adaptor protein which was first identified as a binding partner of amyloid precursor protein (APP), an important molecule in Alzheimer’s disease (AD). FE65 possesses three protein interaction domains, including an N-terminal WW domain and two C-terminal phosphotyrosine binding (PTB) domains, and is capable of mediating assembly of multimolecular complexes. Growing evidence suggests that FE65, in concert with its interacting partners, plays a role in various biological processes including neurodegeneration.
In order to gain a better understanding in the biological functions of FE65, I attempted to look for novel FE65-interacting proteins, and identified Huntingtin (Htt) as a binding partner of FE65. Htt is a ubiquitously expressed 348 kDa protein with little sequence homology to other proteins. It contains a polyglutamine (polyQ) tract followed by a proline-rich domain (PRD) at the N-terminus. An unstable expansion of the polyQ sequence to more than 35 residues is the culprit of Huntington’s disease (HD). In the study, I demonstrated that FE65 WW domain interacts with Htt in a polyQ-dependent manner. Intriguingly, FE65 knockdown results in reduction of mutant Htt protein level. The reduction is attenuated when the ubiquitin-proteasome system (UPS) is inhibited. Moreover, the ubiquitination level of mutant Htt is enhanced when FE65 is knocked down. Additionally, it was found that overexpression of FE65 increases mutant Htt-induced cell death. These results suggest that FE65 promotes the accumulation of mutant Htt in cells by preventing its degradation via the UPS, and thereby enhances the toxicity of mutant Htt.
In addition to recruiting other proteins, the functions of FE65 can be modulated by phosphorylation. Although a number of phosphorylated residues have been identified in FE65, the functional significance of these phosphorylation sites is largely unknown. In the second part of the study, the effect of FE65 Serine-610 (S610), a previously reported phosphorylation site of serum- and glucocorticoidinduced kinase 1 (SGK1), on APP processing, a critical event in AD pathogenesis, was investigated. I demonstrated that phosphorylation of FE65 S610 by SGK1 attenuates APP-FE65 interaction. Consistent with this finding, immunofluorescence staining of APP and FE65 S610 mutants reveals that while FE65 and FE65 S610A colocalize with APP in the perinuclear region, FE65 S610D retains in the nucleus. The effect of FE65 S610 phosphorylation on APP processing at various levels was also examined. The data suggest that when FE65 S610 is phosphorylated by SGK1, the enhancing effect of FE65 on APP processing and Aâ liberation is diminished. Importantly, the phosphorylation status of FE65 S610 was shown be a molecular switch of APP turnover via the UPS. This explains, at least in part, why APP processing is affected.
Collectively, my study broadens our understanding on the functions of FE65 in neurodegenerative processes and provides important insight in future research direction.
