Elucidating the functions of the E3 ubiquitin ligase FBXO7 - with a particular focus on myelinating cells
Molecular Biology and Physiology of Neurons and Glial Cells
Final Report Abstract
Our research results on the atypical parkinsonism-associated E3 ubiquitin ligase FBXO7 in a mouse genetics study (i) and in a molecular/biochemical approach (ii) are: (i) Owing to the prominent expression of FBXO7 in white matter and the overlap of symptoms that characterize parkinsonism and the white matter disease multiple system atrophy, we took a mouse genetic approach to examine the role of FBXO7 in myelinating cells. Myelination of axons facilitates the rapid propagation of electrical signals and the long-term integrity of axons. The ubiquitin proteasome system is essential for proper protein homeostasis, which is particularly crucial for interactions of postmitotic cells. In our study, we examined how the FBXO7 affects the axon-myelin unit. Deletion of Fbxo7 in oligodendrocytes and Schwann cells in mice led to motor impairment due to hindlimb paresis. Here, it triggered axonal degeneration in the CNS and resulted in the severe degeneration of axons in the PNS, inducing a full-blown neuropathy. Both the CNS and PNS displayed inflammation, while the PNS was also characterized by fibrosis, massive infiltration of macrophages, and edema. Tamoxifen-induced deletion of Fbxo7 after myelination, led to a small number of degenerated axons and hence a very mild peripheral neuropathy. Importantly, cultured FBXO7 knockout Schwann cells showed reduced proteasome activity compared to wild type Schwann cells. Taken together, our results demonstrate an essential role for FBXO7 in myelinating cells to support associated axons, which is fundamental to the proper developmental establishment and the long-term integrity of the axon-myelin unit. (ii) In the second part, we followed up an unbiased quantitative mass spectrometry approach to uncover the FBXO7 interactome. Here, we report the association of the E3 ubiquitin ligase FBXO7- SCF (SKP1, cullin-1, F-box protein) with the BAG6 complex, a previously characterized chaperone complex. We identified the subunit GET4 as a direct interactor of FBXO7 and we showed that the subunits GET4 and UBL4A were required for proper proteasome activity. Our findings demonstrated reduced binding of a FBXO7 variants to GET4 and that FBXO7 variants harbored reduced proteasome activity. In addition, we found that GET4 was a non-proteolytic substrate of FBXO7, that binding of GET4 to BAG6 was enhanced in the presence of active FBXO7-SCF and that the cytoplasmic localization of the BAG6 complex was dependent on the ligase activity. Taken together, our data show that the parkinsonism-associated FBXO7 cooperates with the BAG6 complex in proteasome function and determines the subcellular localization of this chaperone complex. Finally, we present preliminary research to characterize the association of FBXO7 and ataxin-3, a de-ubiquitinase implicated in spinocerebellar ataxia type 3. We show that FBXO7 associates with ataxin-3 and that gain- and loss-of-function experiments demonstrate a role for ataxin-3 in proteasome activity regulation.
Publications
- 2017, Mechanistic contributions of FBXO7 to Parkinson’s disease, Journal of Neurochemistry
Joseph S., Schulz, J.B. Stegmüller J.
(See online at https://doi.org/10.1111/jnc.14253) - 2019, Gallyas silver staining of myelination nerve fibers, Bio-Protoc 9(22)
Joseph S., Werner H.B., Stegmüller J.
(See online at https://dx.doi.org/10.21769%2FBioProtoc.3436) - 2019, Myelinating glia-specific deletion of Fbxo7 in mice triggers axonal degeneration in the central nervous system together with peripheral neuropathy, JNeurosci, 39(28)
Joseph S., Vingill S., Jahn O., Fledrich R., Werner H.B., Katona I., Möbius W., Mitkovski M, Huang Y., Weis J., Sereda M.W., Schulz J.B., Nave K.A., Stegmüller J.
(See online at https://doi.org/10.1523/JNEUROSCI.3094-18.2019) - 2021, The parkinsonism-associated protein FBXO7 cooperates with the BAG6 complex in proteasome function and controls subcellular localization of the complex, Biochemical J, 487(12)
Wang Q., Crnkovic V., Preisinger C., Stegmüller J.
(See online at https://doi.org/10.1042/BCJ20201000) - 2021, The role of E3 ubiquitin ligases in synapse function in the healthy and diseased brain, Molecular and Cellular Neuroscience
Kawabe H., Stegmüller J.
(See online at https://doi.org/10.1016/j.mcn.2021.103602)
