UBE2E ubiquitin-conjugating enzymes and ubiquitin isopeptidase Y regulate TDP-43 protein ubiquitination.
Trans-activation element DNA-binding protein of 43 kDa (TDP-43) characterizes insoluble protein aggregates in distinct subtypes of frontotemporal lobar degeneration and amyotrophic lateral sclerosis. TDP-43 mediates many RNA processing steps within distinct protein complexes. Here we identify novel TDP-43 protein interactors found in a yeast two-hybrid screen using an adult human brain cDNA library. We confirmed the TDP-43 interaction of seven hits by co-immunoprecipitation and assessed their co-localization in HEK293E cells. As pathological TDP-43 is ubiquitinated, we focused on the ubiquitin-conjugating enzyme UBE2E3 and the ubiquitin isopeptidase Y (UBPY). When cells were treated with proteasome inhibitor, ubiquitinated and insoluble TDP-43 species accumulated. All three UBE2E family members could enhance the ubiquitination of TDP-43, whereas catalytically inactive UBE2E3(C145S) was much less efficient. Conversely, silencing of UBE2E3 reduced TDP-43 ubiquitination. We examined 15 of the 48 known disease-associated TDP-43 mutants and found that one was excessively ubiquitinated. This strong TDP-43(K263E) ubiquitination was further enhanced by proteasomal inhibition as well as UBE2E3 expression. Conversely, UBE2E3 silencing and expression of UBPY reduced TDP-43(K263E) ubiquitination. Moreover, wild-type but not active site mutant UBPY reduced ubiquitination of TDP-43 C-terminal fragments and of a nuclear import-impaired mutant. In Drosophila melanogaster, UBPY silencing enhanced neurodegenerative TDP-43 phenotypes and the accumulation of insoluble high molecular weight TDP-43 and ubiquitin species. Thus, UBE2E3 and UBPY participate in the regulation of TDP-43 ubiquitination, solubility, and neurodegeneration.
From the Graduate School of Cellular and Molecular Neuroscience, University of Tübingen, Tübingen 72076, Germany, Laboratory of Functional Neurogenetics, Department of Neurodegeneration, German Center for Neurodegenerative Diseases (DZNE), Tübingen 72076, Germany, Laboratory of Functional Neurogenetics, Department of Neurodegeneration and.
Friederike
F
Hans
Laboratory of Functional Neurogenetics, Department of Neurodegeneration and.
Fabienne C
FC
Fiesel
Laboratory of Functional Neurogenetics, Department of Neurodegeneration and.
Jennifer C
JC
Strong
Laboratory of Functional Neurogenetics, Department of Neurodegeneration and.
Sandra
S
Jäckel
Synaptic Plasticity Group, Hertie Institute for Clinical Brain Research, Tübingen 72076, Germany.
Tobias M
TM
Rasse
Laboratory of Functional Neurogenetics, Department of Neurodegeneration and.
Sven
S
Geisler
Laboratory of Functional Neurogenetics, Department of Neurodegeneration, German Center for Neurodegenerative Diseases (DZNE), Tübingen 72076, Germany, Laboratory of Functional Neurogenetics, Department of Neurodegeneration and.
Wolfdieter
W
Springer
Department of Neurology, University Medical Center, Aachen 52074, Germany, and Jülich Aachen Research Alliance (JARA)-Translational Brain Medicine, Aachen 52074, Germany.
Jörg B
JB
Schulz
Department of Neurology, University Medical Center, Aachen 52074, Germany, and.
Aaron
A
Voigt
From the Graduate School of Cellular and Molecular Neuroscience, University of Tübingen, Tübingen 72076, Germany, Laboratory of Functional Neurogenetics, Department of Neurodegeneration, German Center for Neurodegenerative Diseases (DZNE), Tübingen 72076, Germany, Laboratory of Functional Neurogenetics, Department of Neurodegeneration and philipp.kahle@uni-tuebingen.de.
Philipp J
PJ
Kahle
D000328:Adult; D000818:Animals; D001921:Brain / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000378:metabolism; D010450:Endopeptidases / Q000172:deficiency / Q000378:metabolism*; D056827:Endosomal Sorting Complexes Required for Transport / Q000172:deficiency / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D009498:Neurotoxins / Q000378:metabolism; D021381:Protein Transport; D020798:Two-Hybrid System Techniques; D043222:Ubiquitin Thiolesterase / Q000172:deficiency / Q000378:metabolism*; D044763:Ubiquitin-Conjugating Enzymes / Q000378:metabolism*; D054875:Ubiquitination
D004268:DNA-Binding Proteins; D056827:Endosomal Sorting Complexes Required for Transport; D009498:Neurotoxins; D044763:Ubiquitin-Conjugating Enzymes; D010450:Endopeptidases; C541534:USP8 protein, human; D043222:Ubiquitin Thiolesterase
Amyotrophic Lateral Sclerosis (ALS) (Lou Gehrig Disease); Drosophila; Frontotemporal Dementia; Proteasome; Protein Aggregation; Protein-Protein Interaction; TDP-43; Ubiquitin-conjugating Enzyme; Ubiquitin-specific Protease; Ubiquitination