Abstract Details

Title A New Mutation in Ubiquilin Gene Family and Its Effect on Protein Degradation

Topic Anterior Horn

Presentation(s) P02 - (-)

Poster/Presentation
Number 168

Objective

Background BACKGROUND: We reported mutations of Ubiquilin2 gene causes familial amyotrophic lateral sclerosis (FALS) and ALS with dementia. Spinal cord and brain with or without UBQLN2 mutations showed ubiquilin2 positive protein aggregates. The ubiquilin gene families has four members, Ubiquilin1,2,3 and 4, all has a N-terminal ubiquitin like (UBL) domain and a C-terminal ubiquitin associated (UBA) domain.

Design/Methods DESIGN/METHODS: Genomic DNA from 357 Familial ALS and 411 SALS were screened for Ubiquilin 4 variations via SSCP and Sanger sequencing. Variations unidentified in SNP database were further sequenced in 332 normal controls. Recombinant Ubiquilin4 wild type and mutant tagged to mCherry were used to transiently transfect Neuro2A cells (N2A). Confocal microscopy was performed to evaluate protein expression and colocalisation. N2A were co-transfected LC3-GFP and Rab7-GFP and analyzed using a BD LSRFortessa flow cytometer .

Results RESULTS: We identify a mutation in the exon 3 of Ubiquilin4 gene in a FALS patient that was absent from 332 controls. N2A cells transiently transfected with Ubiquilin4 tagged to mCherry showed punctuate structures in both wild type and mutant. Flow cytometry reveled a significant accumulation of LC3-GFP which corresponds to an inhibition of autophagic activity in mutant Ubiquilin4 transfected cells. We also observed that both wild and mutant Ubiquilin4 co-localize with LC3, p62 and subsequently Ubiquilin2. However we found a significant decrease in the Rab7-GFP accumulation in the mutant Ubiquilin4 transfected cells which may indicate impairment of autophagosome late endosome formation.

Conclusions CONCLUSIONS: These data suggest that Ubiquilin4 is mutated in ALS and Ubiquilin4 mutation impairs the UPS and autophagy protein degradation pathway. Such mechanisms could lead to protein aggregation in ALS, and serve as targets for designing rational therapeutics in ALS.

Authors/Disclosures
Jianhua Yan, PhD (Northwestern University) PRESENTER	No disclosure on file
	No disclosure on file
Faisal Fecto, MD, PhD (Advocate Medical Group)	The institution of Dr. Fecto has received research support from NINDS.
Yong Shi, PhD (Northwestern University)	No disclosure on file
Dileep R. Yavagal, MD, FAAN (University of Miami Miller School of Medicine)	Dr. Yavagal has received personal compensation in the range of $10,000-$49,999 for serving as a Consultant for Vascular Dynamics. Dr. Yavagal has received personal compensation in the range of $5,000-$9,999 for serving as a Consultant for Guidepoint. The institution of Dr. Yavagal has received personal compensation in the range of $500-$4,999 for serving as a Consultant for GLG. Dr. Yavagal has received personal compensation in the range of $0-$499 for serving as a Consultant for Medtronic. Dr. Yavagal has received personal compensation in the range of $0-$499 for serving as a Consultant for Cerenovus/Johnson & Johnson. Dr. Yavagal has received personal compensation in the range of $0-$499 for serving as a Consultant for Stryker. Dr. Yavagal has received personal compensation in the range of $0-$499 for serving as a Consultant for Poseydon. Dr. Yavagal has received personal compensation in the range of $10,000-$49,999 for serving as a Consultant for Athersys. Dr. Yavagal has received personal compensation in the range of $500-$4,999 for serving as a Consultant for Gravity Medical Technology. Dr. Yavagal has received personal compensation in the range of $0-$499 for serving on a Scientific Advisory or Data Safety Monitoring board for Poseydon. Dr. Yavagal has stock in Poseydon. Dr. Yavagal has stock in Galaxy Therapeutics. Dr. Yavagal has stock in Stentrode. Dr. Yavagal has stock in Gravity Medical. Dr. Yavagal has stock in Gravity Medical Technology. Dr. Yavagal has stock in Athersys. Dr. Yavagal has stock in Rapid Medical . Dr. Yavagal has received intellectual property interests from a discovery or technology relating to health care. Dr. Yavagal has received personal compensation in the range of $5,000-$9,999 for serving as a Medicolegal Expert with Royal Carribean Cruise Line. Dr. Yavagal has received personal compensation in the range of $5,000-$9,999 for serving as a Medicolegal Consultant with Carnival Cruise Line.
	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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