Serine-65 phosphorylated ubiquitin associates with soluble tau and seeding activity in Alzheimer’s disease

Tau propagation is a key driver of Alzheimer’s disease (AD), yet selective markers of seeding-competent tau remain elusive. Ubiquitin phosphorylated at Serine-65 (pS65-Ub) is a PINK1-derived cellular stress signal that accumulates in AD with elevated tau pathology, but its relationship to tau seeding is unknown. Here, using postmortem brain tissue from female and male donors, we show that a subset of tau is covalently modified by pS65-Ub in AD brain.

The pS65-Ub-associated tau population is elevated in AD and is enriched in soluble, high-molecular-weight, oligomeric, and hyperphosphorylated tau species. Across AD cases, soluble pS65-Ub-associated tau levels strongly correlate with tau seeding activity. Tau species isolated with pS65-Ub antibodies exhibit seeding activity, whereas removal of pS65-Ub-associated material markedly reduces seeding from AD brain extracts.

Together, these findings identify pS65-Ub as a disease-enriched molecular feature of a seed-competent tau population and link cellular stress signaling with pathological tau propagation in AD. We thank all patients and family members who made this study possible. We thank Monica Castanedes Casey, Bailey Rawlinson, and Michael DeTure from the Neuropathology Laboratory at Mayo Clinic Jacksonville for processing human post-mortem brain tissue.

We thank Cody Hoffmann and Benjamin J. Madden from the Mayo Clinic Proteomics Core Facility for technical support and data generation, supported in part by the Mayo Clinic Comprehensive Cancer Center Grant (P30CA15083). We thank Casey Cook for help with the original MSD assay.

We thank Marc Diamond’s lab for kindly sharing tau-FRET biosensor cells. We are grateful to the late Peter Davies for providing the pS202-tau (CP13) and pS396/404-tau (PHF1) antibodies. Figures were created with BioRender.com, with permission.

This work was supported by the Florida Department of Health – Ed and Ethel Moore Alzheimer’s Disease Research Program 25A11 (to T.Y.), 22A07 and 26A09 (to F.C.F.), and 9AZ10 (to W.S.); Alzheimer’s Association Research Fellowships AARF-22-974258 (to T.Y.) and AARF-22-973152 (to X.H.); Alzheimer’s Association and National Alzheimer’s Coordinating Center New Investigator Awards NIAP25-1446587 (to T.Y.) and NIAP25-1445722 (to X.H.); the Mayo Clinic Alzheimer Disease Research Center Developmental Project Award P30AG062677 (to X.H.); the Michael J. Fox Foundation for Parkinson’s Research (to F.C.F. and W.S.); the Mayo Clinic Center for Biomedical Discovery and Office of Belonging Research Catalyst Award (to F.C.F.); National Institute on Aging R56AG062556 (to W.S.); National Institute of Neurological Disorders and Stroke RF1NS085070 and U54NS110435 (to W.S.); The Ted Nash Long Life Foundation (to W.S.); and the Mayo Clinic Robert and Arlene Kogod Center on Aging (to W.S.). Department of Neuroscience, Mayo Clinic, Jacksonville, FL, USA Tingxiang Yan, Xu Hou, Jens O.