Regional
USP25-Mediated Talin-1 stabilization in platelets: A novel mechanism of hyperreactivity and thrombosis risk during aging
Xuemei Jia
Shanghai First People's Hospital, Shanghai, China
Rong Xia
Huashan Hospital, Fudan University, China.
Aging is a well-recognized risk factor for platelet hyperreactivity and thrombotic events, yet the molecular underpinnings remain incompletely defined. In this study, Jia and colleagues investigated whether age-associated remodeling of protein homeostasis—specifically ubiquitination and deubiquitination—contributes to enhanced platelet reactivity in the elderly. Using parallel murine and human datasets, the authors established a consistent phenotype of aging-associated platelet hyperresponsiveness and then linked this phenotype to a specific deubiquitinating enzyme (DUB)–substrate axis centered on USP25 and talin-1.
The authors first validated the aging phenotype across species. Compared with young mice, platelets from aged mice displayed heightened collagen-induced aggregation, increased ATP release, augmented integrin αIIbβ3 activation, and enhanced spreading on fibrinogen. A similar pattern was observed in healthy human donors, with older individuals (>65 years) exhibiting greater platelet aggregation and spreading than young controls (18–23 years). These findings support the concept that platelet hyperreactivity is an evolutionarily conserved feature of aging.
To explore the mechanistic basis, the investigators performed ubiquitin-remnant (di-Gly) enrichment proteomics (“ubiquitin-modified proteome”) and immunoblotting. They observed a global reduction in ubiquitination in resting platelets from aged mice and older human donors. Importantly, this decline was not explained by reduced transcription of ubiquitin-encoding genes or diminished free ubiquitin protein levels, pointing instead to altered ubiquitination dynamics. Given that ubiquitination is reversible and tightly regulated by DUBs, the authors tested whether DUB activity contributed to the age-associated ubiquitination shift. Pharmacologic DUB inhibition (PR-619) suppressed platelet aggregation and secretion, and notably restored ubiquitin conjugates in aged platelets. Moreover, platelets from aged mice were more sensitive to low-dose PR-619, consistent with increased DUB activity as a functional driver of the observed ubiquitination phenotype.
The study then focused on identifying which DUB(s) might mediate this effect. By integrating publicly available platelet RNA-seq datasets with protein validation, USP25 emerged as a highly expressed DUB that is significantly upregulated in platelets from aged mice and older human donors, with partial localization near the platelet membrane. Genetic loss-of-function experiments in USP25 knockout mice demonstrated that USP25 is required for full platelet functional capacity: USP25 deficiency impaired aggregation induced by multiple agonists (including ADP, collagen, and thrombin), reduced dense granule secretion, weakened integrin “outside-in”–dependent processes (spreading and clot retraction), and decreased integrin αIIbβ3 activation. In vivo, USP25 deficiency delayed FeCl3-induced arterial thrombosis and reduced thrombus size, while prolonging tail bleeding time. Platelet depletion–reconstitution experiments confirmed that these effects were platelet-intrinsic.
Mechanistically, the authors combined two orthogonal discovery approaches: (i) ubiquitin-modified proteomics comparing wild-type and USP25-deficient platelets and (ii) immunoprecipitation–mass spectrometry to identify USP25-interacting partners in human platelets. Intersection analysis highlighted talin-1 (TLN1) as a prime candidate substrate. Subsequent co-immunoprecipitation and confocal microscopy confirmed endogenous USP25–talin-1 interaction in resting and activated platelets, with increased co-localization in older donors. Domain mapping suggested that USP25’s catalytic and coiled-coil regions contribute to binding, and a defined talin-1 region mediates association. Functionally, USP25 stabilized talin-1 protein through its deubiquitinase activity: talin-1 levels were reduced in USP25-deficient platelets without changes in talin-1 mRNA, and proteasome inhibition (MG132) abrogated USP25-dependent talin-1 stabilization, indicating a proteasome-dependent degradation mechanism. The catalytically inactive USP25 mutant failed to stabilize talin-1, supporting an enzymatic requirement. Importantly, USP25 preferentially stabilized the talin-1 rod domain, a region implicated in integrin signaling competence, and USP25 deficiency eliminated the age-associated rise in talin-1 levels observed in wild-type platelets. The study further characterized ubiquitin linkage preferences and identified key talin-1 lysine residues (including K869 and K2049) relevant to USP25-mediated deubiquitination.
Finally, the authors explored translational feasibility using AZ1, a dual USP25/USP28 inhibitor. In platelets from older human donors and aged mice, AZ1 reduced talin-1 protein abundance (time-dependent) and suppressed platelet aggregation, secretion, integrin activation, spreading, and clot retraction. In vivo, AZ1 attenuated age-dependent thrombotic susceptibility. However, residual effects of AZ1—particularly on collagen/GPVI-related responses even in USP25-deficient platelets—suggest potential off-target or USP25-independent actions, underscoring the need for improved selectivity and careful assessment of the bleeding–thrombosis therapeutic window. Collectively, the work defines an aging-associated remodeling of platelet ubiquitination and establishes USP25-mediated talin-1 stabilization as a mechanistic axis that promotes platelet hyperreactivity and thrombotic susceptibility in older individuals.
Brief perspective: recent progress and how this work advances the field
Aging-related platelet hyperreactivity has been linked to systemic inflammation and mitochondrial/redox dysregulation. The current study complements those frameworks by introducing protein homeostasis (ubiquitination dynamics) as an additional mechanistic layer that converges on a terminal effector of platelet function—integrin αIIbβ3 activation—via talin-1 stabilization.
Ubiquitin–proteasome and deubiquitination pathways are increasingly recognized as active regulators in anucleate platelets. This study provides a particularly strong “mechanism-to-phenotype” chain by pinpointing a specific DUB (USP25), validating platelet-intrinsic function through knockout and reconstitution experiments, and identifying a biologically coherent substrate (talin-1) that directly explains altered integrin signaling and thrombosis phenotypes.
References
1. Jia X, Jiang S, Cheng H, Wang Z, Chen Z, Dong W, et al. USP25-Mediated Talin-1 Stabilization in Platelets: A Novel Mechanism of Hyperreactivity and Thrombosis Risk During Aging. Blood. 2025:blood.2023023352. doi: 10.1182/blood.2023023352.
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