Abstract Details

Title ALS/FTD-linked UBQLN2-P497H Increases K63-linked Polyubiquitinated Protein Levels

Topic Neuromuscular and Clinical Neurophysiology (EMG)

Presentation(s) P1 - Poster Session 1 (5:30 PM-6:30 PM)

Poster/Presentation
Number 12-030

Objective To identify specific ubiquitin linkage types present in the ubiquitinated aggregates of the UBQLN2^P497H transgenic mouse brain.

Background A UBQLN2^P497H mutation has been identified in patients with ALS/FTD. Ubiquilin2, ubiquitin, and P62 are components of pathological protein inclusions in the brains and spinal cords of UBQLN2^P497H patients as well as many other ALS/FTD patients without a UBQLN2 mutation. Ubiquitin is a ubiquitously expressed regulatory protein that is covalently linked to other proteins to alter their cellular location, activity, and interactions or mark them for degradation by the proteasome. These signals are mediated by different ubiquitin linkage types. We have shown that UBQLN2^P497H disrupts the Ubiquitin-Proteasome System (UPS), which degrades K29- and K48-linked substrates. However, it is unknown which specific ubiquitin linkages are present in the pathological protein inclusions found in ALS/FTD patients.

Design/Methods Whole brain lysates were prepared from non-transgenic, UBQLN2^WT transgenic mice, UBQLN2^P497H transgenic mice, and Ubqln2^KO mice. Immunoprecipitation assays using Tandem Ubiquitin Binding Entities (TUBEs) and western blotting were carried out. Immunohistochemistry was performed to determine if specific ubiquitin linkages localize to pathological protein inclusions in UBQLN2^P497H or other locations in transgenic mice.

Results We detect increased levels of K48 and K63-linked polyubiquitinated protein in UBQLN2^P497H transgenic mouse brain, but not in non-transgenic, UBQLN2^WT transgenic mice or Ubqln2^KO mice. K48-linked proteins are detected in the pathological protein inclusions in UBQLN2^P497H transgenic mice. Experiments to determine if K63-linked proteins are also present are ongoing.

Conclusions An increase of K48-linked proteins is expected due to the known UPS impairment caused by UBQLN2^P497H. Given that K63-linked proteins are not known to be a UPS substrate, it was not expected to be increased. Our results suggest an additional pathological mechanism giving rise to the K63-linked protein increase. Additionally, the absence of these alterations in Ubqln2^KO mice suggests a gain-of-function mechanism for the pathogenesis caused by UBQLN2^P497H.

Authors/Disclosures
PRESENTER	No disclosure on file
Thomas J. Lukas, PhD (Northwestern University)	No disclosure on file
Yong Shi, PhD (Northwestern University)	No disclosure on file
	No disclosure on file
Han-Xiang Deng, MD	Dr. Deng has nothing to disclose.
Teepu Siddique, MD, FAAN	No disclosure on file

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