Alzheimer disease-specific conformation of hyperphosphorylated paired helical filament-Tau is polyubiquitinated through Lys-48, Lys-11, and Lys-6 ubiquitin conjugation.
One of the key pathological hallmarks of Alzheimer disease (AD) is the accumulation of paired helical filaments (PHFs) of hyperphosphorylated microtubule-associated protein Tau. Tandem mass spectrometry was employed to examine PHF-Tau post-translational modifications, in particular protein phosphorylation and ubiquitination, to shed light on their role in the early stages of Alzheimer disease. PHF-Tau from Alzheimer disease brain was affinity-purified by MC1 monoclonal antibody to isolate a soluble fraction of PHF-Tau in a conformation unique to human AD brain. A large number of phosphorylation sites were identified by employing a data-dependent neutral loss algorithm to trigger MS3 scans of phosphopeptides. It was found that soluble PHF-Tau is ubiquitinated at its microtubule-binding domain at residues Lys-254, Lys-311, and Lys-353, suggesting that ubiquitination of PHF-Tau may be an earlier pathological event than previously thought and that ubiquitination could play a regulatory role in modulating the integrity of microtubules during the course of AD. Tandem mass spectrometry data for ubiquitin itself indicate that PHF-Tau is modified by three polyubiquitin linkages, at Lys-6, Lys-11, and Lys-48. Relative quantitative analysis indicates that Lys-48-linked polyubiquitination is the primary form of polyubiquitination with a minor portion of ubiquitin linked at Lys-6 and Lys-11. Because modification by Lys-48-linked polyubiquitin chains is known to serve as the essential means of targeting proteins for degradation by the ubiquitin-proteasome system, and it has been reported that modification at Lys-6 inhibits ubiquitin-dependent protein degradation, a failure of the ubiquitin-proteasome system could play a role in initiating the formation of degradation-resistant PHF tangles.
Cripps
Diane
D
Department of Pharmaceutical Sciences, University of Southern California, Los Angeles, California 90089, USA.
Thomas
Stefani N
SN
Jeng
Young
Y
Yang
Frank
F
Davies
Peter
P
Yang
Austin J
AJ
D000465:Algorithms; D000544:Alzheimer Disease / Q000378:metabolism*; D000595:Amino Acid Sequence; D002853:Chromatography, Liquid; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D008239:Lysine / Q000737:chemistry*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009410:Nerve Degeneration / Q000378:metabolism; D016874:Neurofibrillary Tangles / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D010766:Phosphorylation; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000737:chemistry; D012997:Solvents / Q000737:chemistry; D013912:Threonine / Q000737:chemistry; D014357:Trypsin / Q000737:chemistry; D025801:Ubiquitin / Q000737:chemistry*; D016875:tau Proteins / Q000737:chemistry*
D010455:Peptides; D012997:Solvents; D025801:Ubiquitin; D016875:tau Proteins; D013912:Threonine; D012694:Serine; D014357:Trypsin; D046988:Proteasome Endopeptidase Complex; D008239:Lysine