Biology subjects

Ngu, H.

Publications and source records attributed to Ngu, H..

6 recordsLinked to original sources

bioRxiv2026

Phosphorylated ubiquitin is a secondary messenger and an epigenetic mark mediating mitochondria to nucleus signaling

AbstractParkinsons disease (PD) is commonly associated with dysfunctional mitochondrial homeostasis. PINK1, a S/T kinase mutated in early-onset PD, generates phosphoserine 65 ubiquitin (pS65Ub) on damaged mitochondria facilitating their removal. Here, we show that pS65Ub translocates into the nucleus after generation at damaged mitochondria and is directly attached to substrates by resident E3 ligases. Histone H2A is a major substrate and is modified at lysine 119 (H2AK119) by the polycomb silencer, E3 ligase RING1B. At nucleosomes, pS65Ub simultaneously suppresses RING1B and potentiates H2A deubiquitinases USP16 and USP21. Epigenetic profiling and RNA sequencing reveal that pS65Ub is enriched at the promoters of poorly expressed yet dynamically regulated genes and is associated with H2AK119ub depletion. Functionally, we show that pS65Ub enrichment drives polycomb target gene expression, which accelerates the maturation of dopaminergic neurons. Importantly, post-mortem PD brains exhibit elevated nuclear pS65Ub, potentially linking nuclear pS65Ub accumulation with disease pathogenesis. Together, these data indicate that pS65Ub generated at damaged mitochondria regulates fundamental cellular processes at distant sites.

cell biology↗

bioRxiv2026

Practical quantification of immunohistochemistry antigen concentrations and reaction-diffusion parameters

The immunohistochemistry (IHC) methods widely used in diagnostic medicine and biomedical research are kinetically complex reaction-diffusion processes that, ideally, produce stain intensities correlated with the local antigen concentration. Yet after 75 years of use, practical theoretical tools to rigorously plan and interpret IHC experiments are still lacking. Because modeling the reactions requires time-consuming computer simulation, impractical for regular use, most protocols are optimized empirically, without detailed knowledge of the reaction rates and antigen-antibody equilibria. The resulting stain intensities can be calibrated against standards with known antigen abundance, but they are typically not interpretable in terms of chemical antigen concentrations. To address these limitations, we developed a fast interpolation method to model reaction-diffusion behavior, and experimental methods to characterize IHC kinetic parameters in formalin-fixed paraffin-embedded (FFPE) samples. Used together, these allow experimental measurement of both the chemical concentration of antigen in the sample and the reaction-diffusion parameters consistent with the assay results. Results show 1) direct immunofluorescent detection has low nanomolar sensitivity with >1000-fold dynamic range, and 2) antibody diffusion rates in FFPE samples can be >1000-fold slower than in aqueous solutions, producing diffusion-limited conditions in which the IHC reaction time course may depend on the sample antigen concentration. Awareness of these details is necessary to avoid potential underestimation of both the absolute and relative antigen concentrations in different samples that may occur if staining is stopped before reaching equilibrium. Software tools are provided to allow users to rapidly model IHC reaction time courses and to fit experimental time course data with candidate reaction parameters. The principles described here apply equally to other tissue-based "spatial omics" analyses and should be considered when designing and interpreting experiments requiring any macromolecule to diffuse into and react in a tissue section. SIGNIFICANCEThe theoretical and experimental framework described here advances IHC staining from a qualitative or semi-quantitative method towards a more rigorously quantitative assay. The practical ability to predict IHC reaction kinetics and fit reaction parameters to experimental data has the potential to advance IHC applications in diagnostic medicine and biomedical research in three ways: 1) interpretation of experimental and diagnostic samples stained under different conditions can be more objective, facilitating comparison of results from different protocols and different laboratories; 2) IHC staining can be interpreted as molar chemical antigen-antibody concentrations calculated from the reaction parameters measured in the studied sample; 3) the correlation between antigen concentration and biological behavior can be examined more reliably. Practical software tools are provided.

pathology↗

bioRxiv2025

Intranigral injection of Alpha-Synuclein pre-formed fibrils leads to BBB compromise and Bilateral Dopaminergic Neurodegeneration in A53T Alpha-Synuclein transgenic mice.

Parkinsons disease (PD) is a progressive neurodegenerative disorder characterized by alpha-()-Synuclein neuronal aggregation and loss of dopaminergic (DA) neurons. Developing animal models that replicate PDs neuropathological phenotypes is critical for understanding its pathophysiology and evaluating potential therapeutic targets. In this study, we show that direct unilateral injection of human -Synuclein PFFs into the Substantia Nigra (SN) of mutant A53T -synuclein overexpressing mice induce bilateral phosphorylated -Synuclein (pS129) pathology in the SN. This pathology spreads to the striatum, cerebral cortex, and midbrain within 60 days and is accompanied by neuroinflammation in the midbrain and cerebral cortex. Additionally, we observed synuclein-dependent neurodegeneration, with a 50% reduction in Tyrosine Hydroxylase (TH) intensity in the SN and a 40% reduction in Striatum, both bilaterally. The model also revealed a compromised blood-brain barrier (BBB) and T-cell infiltration in the PFF injected animals, correlating with pS129 pathology and neuroinflammation. Taken together, we developed a mouse model that recapitulates multiple PD phenotypes, providing a valuable platform for testing therapeutic strategies targeting human -Synuclein pathology and for exploring CNS-peripheral immune interactions in PD. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=135 SRC="FIGDIR/small/690780v2_ufig1.gif" ALT="Figure 1"> View larger version (20K): org.highwire.dtl.DTLVardef@19c6234org.highwire.dtl.DTLVardef@1f36bbaorg.highwire.dtl.DTLVardef@25ac6borg.highwire.dtl.DTLVardef@15ad61d_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2025

Complement C3aR deletion does not attenuate neurodegeneration in a tauopathy model or alter acute inflammation-induced gene expression changes in the brain

Aberrant activation of the classical complement pathway in the brain is implicated in contributing to synapse loss and neurodegeneration in various neurodegenerative conditions. Given that C3aR is a druggable target in the complement pathway, we evaluated the potential of C3aR KO to rescue neurodegeneration in a tauopathy model and neuroinflammatory responses in an acute endotoxemia model. We found that C3aR KO did not rescue tau pathology, microglia activation markers, neurodegeneration, or behavioral abnormalities in tauopathy model mice. While we found that endotoxemia resulted in numerous transcriptional changes including distinct alterations in subpopulations of microglia, astrocytes and oligodendrocytes, C3aR KO did not impact these alterations. Together, our results suggest that the beneficial effects of blocking the complement classical pathway in neurodegeneration models is likely independent of C3aR activation and raise questions about the rationale for therapeutically targeting C3aR for neurodegenerative disease.

neuroscience↗

bioRxiv2022

TPL2 kinase activity regulates microglial inflammatory responses and promotes neurodegeneration in tauopathy mice

TPL2 (MAP3K8) is a central signaling node in the inflammatory response of peripheral immune cells. We find that TPL2 kinase activity modulates microglial cytokine release and is required for microglia-mediated neuron death in vitro. In acute in vivo neuroinflammation settings, TPL2 kinase activity regulates microglia activation states and brain cytokine levels. In a tauopathy model of chronic neurodegeneration, loss of TPL2 kinase activity reduces neuroinflammation and rescues synapse loss, brain volume loss, and behavioral deficits. Single-cell RNAseq analysis indicates protection in the tauopathy model was associated with reductions in activated microglia subpopulations as well as infiltrating peripheral immune cells. Overall, using various models, we find that TPL2 kinase activity can promote multiple harmful consequences of microglial activation in the brain including cytokine release, iNOS induction, astrocyte activation, and immune cell infiltration. Consequently, inhibiting TPL2 kinase activity could represent a potential therapeutic strategy in neurodegenerative conditions.

neuroscience↗

bioRxiv2020

Global proteomics of Ubqln2-based murine models of ALS

Familial forms of neurodegenerative diseases commonly involve mutation of aggregation-prone proteins or components of the protein degradation machinery that act on aberrant proteins. Ubqln2 encodes a member of the UBL/UBA family of proteasome shuttle factors that is thought to facilitate proteasomal degradation of substrates, and mutation of this gene results in a familial form of ALS/FTD in humans. How Ubqln2 dysfunction leads to neurodegeneration, however, remains uncertain. We undertook a comprehensive study to identify proteomic changes upon Ubqln2 perturbation in multiple murine models of Ubqln2-mediated neurodegenerative disease. By performing quantitative multiplexed proteomics on neural tissues of affected animals, we identified a small group of proteins whose abundance is tightly linked to UBQLN2 function: the ubiquitin ligase TRIM32 and two retroelement-derived proteins, PEG10 and CXX1B. Further studies using cultured cells of human origin, including induced neurons, found similar changes in protein abundance upon Ubqln2 loss, and pulse-chase studies suggested that PEG10 and TRIM32 are direct clients of UBQLN2. In conclusion, our study provides a deep understanding of the proteomic landscape of ALS-related Ubqln2 mutants and identifies candidate client proteins that are altered in vivo in disease models and whose degradation is promoted by UBQLN2.

cell biology↗