Biology subjects

Lao, Y.

Publications and source records attributed to Lao, Y..

6 recordsLinked to original sources

bioRxiv2025

Transcriptomic and protein analysis of human cortex reveals genes and pathways linked to NPTX2 disruption in Alzheimer's disease

BackgroundCognitive resilience to Alzheimers disease (AD) pathology is associated with preserved expression of NPTX2, an activity-regulated synaptic protein involved in circuit plasticity, excitation-inhibition balance, and complement-linked synapse regulation. However, the broader molecular programs coordinated with NPTX2 in resilient individuals remain unclear. MethodsWe analyzed postmortem middle temporal gyrus tissue using targeted PRM-MS proteomics in 135 individuals and bulk RNA-seq in an expanded 575-sample cohort. NPTX2-associated molecular coordination was assessed within cognitively normal low-pathology controls (CN-Lo), cognitively normal high-pathology controls (CN-Hi), mild cognitive impairment (MCI), and AD. Correlation-based approaches were applied using NPTX2 protein and NPTX2 mRNA expression as anchors to define resilience mechanisms in CN-Hi subjects. ResultsNPTX2 protein abundance was preserved across all controls regardless of age and pathology but reduced in MCI and AD. NPTX2 mRNA expression was also invariant across pathology within controls and reduced in MCI and AD but decreased markedly with age. Targeted proteomics identified NPTX2 relationships with synaptic and inhibitory-circuit proteins that were preserved across control groups, alongside CN-Hi-specific recruitment of trafficking, lysosomal, metabolic, and proteostasis-associated proteins. Transcriptome-wide correlations with NPTX2 revealed differences in gene co-expression between groups, identifying a prominent activity-dependent program including BDNF, VGF, SCG2, SST, SERTM1, DUSP4, and EGR4, that was preserved in both CN-Lo and CN-Hi subjects, while genes recruited to the NPTX2 network specifically in CN-Hi implicated immune, neuroprotective, translation, and proteostasis-related pathways. Coupling differential gene expression analysis with co-expression, we further identified five candidate resilience genes whose expression and NPTX correlation was preserved across controls, but lost in MCI and AD: SST, MAL2, TAC1, SERTM1, and RFK. Expression of genes in distinct NPTX2 co-expression classes can be freely explored in our bulk RNA-seq data and other public AD transcriptomic datasets at NeMO Analytics. ConclusionFindings suggest that cognitive resilience in the context of AD neuropathology engages a coordinated molecular state distinct from both persevered cognition without pathology and MCI/AD, which is organized around preserved and selectively remodeled NPTX2-associations. Rather than reflecting broad transcript abundance changes, resilience was characterized by maintained synaptic and inhibitory programs, and adaptive proteostasis and trafficking pathways that distinguish resilient high-pathology individuals from low-pathology controls or symptomatic AD.

bioinformatics↗

bioRxiv2025

Antimuscarinic drugs exert β-arrestin-biased agonism at the muscarinic acetylcholine type 1 receptor

Previous studies indicate that both pirenzepine (PZ), a selective orthosteric muscarinic acetylcholine type 1 receptor (M1R) antagonist, and muscarinic toxin 7 (MT7), a negative M1R allosteric modulator (NAM), act via M1R to promote neuritogenesis in cultured adult rodent primary dorsal root ganglia (DRG) sensory neurons, in part, through {beta}-arrestin-dependent activation of extracellular signal-regulated protein kinase 1/2 (ERK1/2). Furthermore, these antagonists reverse nerve degeneration in a variety of rodent models of peripheral neuropathy through multiple complementary pathways. To understand the therapeutic effects and mechanism of M1R antagonist-induced ERK1/2 phosphorylation, we tested the hypothesis that PZ and MT7 possess {beta}-arrestin-biased agonism at M1R to drive activation of ERK and enhance neurite outgrowth. Treatment for up to 30 min with PZ and MT7 dose-dependently recruited {beta}-arrestin2 to M1R (analyzed using nano-BRET) and increased ERK phosphorylation in both HEK293 cells and DRG neurons. DRG neurons of different sub-types express M1R, and ERK activation by MT7 was only observed in M1R-positive neurons. These novel pharmacological effects occurred in the absence of activation of G protein signaling or receptor internalization. PZ phosphorylated M1R at six specific serine/threonine residues (T230, S251, T254, S321, T354, S356) of intracellular loop 3 (ICL3) and deletion mutation of these sites suppressed PZ and MT7 induction of {beta}-arrestin binding to M1R and inhibited ERK activation. With regard to PZ signaling, alanine substitution at S251 and T254 was sufficient to impede {beta}-arrestin binding and ERK activation. {beta}-arrestin-biased activity of PZ and MT7 involved the mobilization of casein kinase 2 (CK2) and this occurred in the absence of Gq or G protein receptor kinase (GRK) activity. Pharmacological or siRNA-based inhibition of CK2 blocked PZ-induction of {beta}-arrestin association, ERK activation and neurite outgrowth in DRG neurons. In conclusion, PZ/MT7 activated M1R toward the {beta}-arrestin signaling pathway in both HEK293 cells and DRG neurons to augment ERK activation and neurite outgrowth via engagement of CK2. One-sentence summaryAntimuscarinic drugs act as {beta}-arrestin-biased agonists via casein kinase 2 activation to promote ERK1/2 phosphorylation and neurite outgrowth O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/649213v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@1bb8da7org.highwire.dtl.DTLVardef@510218org.highwire.dtl.DTLVardef@608d69org.highwire.dtl.DTLVardef@e40d9e_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstract.C_FLOATNO Schematic presentation of the effect of muscarinic ligands at M1R associated signaling pathway. (A) Muscarine/carbachol acts as a balanced ligand by engaging both Gq and -arrestin signaling pathways and treatment with pirenzepine/MT7 blocks these effects. (B) Pirenzepine/MT7 acts as a -arrestin biased ligand by 1) phosphorylating of ICL3 region of M1R via CK2 (but not GRKs), 2) no activation of G protein signaling, 3) recruitment of -arrestin 2 and 4) ERK1/2 activation leading to neurite outgrowth in DRG sensory neurons. This figure was generated by BioRender under license number EK285MUOPQ. C_FIG

pharmacology and toxicology↗

bioRxiv2024

SysQuan: repurposing SILAC mice for the affordable absolute quantitation of the human proteome

Relative quantitation, used by most MS-based proteomics laboratories to determine protein fold-changes, requires samples being processed and analyzed together for best comparability through minimizing batch differences. This limits the adoption of MS-based proteomics in population-wide studies, and the detection of subtle but relevant changes in heterogeneous samples. Absolute quantitation circumvents these limitations and enables comparison of results across laboratories, studies, and longitudinally. However, high costs of the essential stable isotope labeled (SIL) standards prevents widespread access and limits the number of quantifiable proteins. Our new approach, called "SysQuan", repurposes SILAC mouse tissues/biofluids as system-wide internal standards for matched human samples to enable absolute quantitation of, theoretically, two-thirds of the human proteome using 157,086 shared tryptic peptides. We demonstrate that SysQuan enables quantification of 70% and 31% of the liver and plasma proteomes, respectively. We demonstrate for 14 metabolic proteins that abundant SIL mouse tissues enable cost-effective reverse absolute quantitation in, theoretically, 1000s of human samples. Moreover, 10,000s of light/heavy doublets in untargeted SysQuan datasets enable unique post-acquisition absolute quantitation. SysQuan empowers researchers to replace relative quantitation with affordable absolute quantitation at scale, making data comparable across laboratories, diseases and tissues, enabling completely novel study designs and increasing reusability of data in repositories. TOC Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=115 SRC="FIGDIR/small/622109v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@c7a58org.highwire.dtl.DTLVardef@1344320org.highwire.dtl.DTLVardef@2321e3org.highwire.dtl.DTLVardef@85bf2d_HPS_FORMAT_FIGEXP M_FIG C_FIG

systems biology↗

bioRxiv2024

Identification of novel myokines and putative protein targets that mediate functional adaptations in response to chronic contractile activity induced skeletal muscle-extracellular vesicle treatment

We have previously shown that skeletal muscle-derived extracellular vesicles (EVs) released post-chronic contractile activity (CCA) increased mitochondrial biogenesis in murine myoblasts, and decreased cell viability and induced apoptosis and senescence in non-small cell lung cancer cells. While the underlying mechanisms are unknown, the effects perpetuated were dependent on membrane-bound proteins. Here, we performed an extensive LC-MS/MS proteomic analysis on EVs from control and CCA myotubes. A total of 2900 proteins were identified in CON-EVs and CCA-EVs, including EV-associated proteins such as TSG101, tetraspanins (CD9, CD81, and CD63), flotillin-1, and annexins. Of these, 856 proteins are novel and not listed in EV databases (ExoCarta and Vesiclepedia), indicating that myotube-EVs harbor proteins not yet identified in EVs of different origin. Additionally, we identified 2062 unique proteins that have not yet been previously reported in myotube-EVs to date. Remarkably, of the 2900 total proteins identified, we observed 46 upregulated, and 25 downregulated differentially expressed proteins (DEPs) in CCA-EVs vs. control-EVs. Most of upregulated DEPs include EV-associated proteins. Comparing the 71 DEPs with proteins expressed in skeletal muscle indicated 61 of these as potential myokines. We identified actin cytoskeleton signaling, integrin signaling and muscle contraction as the most enriched pathways among the DEPs using different databases/software including FunRich, KEGG, STRING and Ingenuity Pathway Analysis. Using a relevance score that prioritized membrane-bound proteins with known function in mitochondrial biogenesis and inhibition of cancer growth, we identified top-scoring highly enriched DEPs of interest: IGF1R, ATP7A, PFN1, GJA1, PRKCA and ITGA6. We confirmed upregulation of these targets in EVs using immunoblotting. Among these top-scoring DEPs, PFN1, and ITGA6 are associated with EVs, with expression upregulated following acute exercise. In summary, we report the first comprehensive analysis of skeletal muscle-EV proteome following CCA, with identification of putative protein targets and signaling pathways that may execute the pro-metabolic and anti-tumorigenic effects of CCA-EVs.

cell biology↗

bioRxiv2023

Activation of Shh/Smo is sufficient to maintain oligodendrocyte precursor cells in an undifferentiated state but is not necessary for differentiation

Myelination is the terminal step in a complex and precisely timed program that orchestrates the proliferation, migration and differentiation of oligodendroglial cells. It is thought that Sonic Hedgehog (Shh) acting on Smoothened (Smo) participates in regulating this process, but that these effects are highly context dependent. Here, we investigate oligodendroglial development and remyelination from three specific transgenic lines: NG2-CreERT2 (control), Smofl/fl/NG2-CreERT2 (loss of function) and SmoM2/NG2-CreERT2 (gain of function), as well as pharmacological manipulation that enhance or inhibit the Smo pathway (SAG or cyclopamine treatment respectively). To explore the effects of Shh/Smo on differentiation and myelination in vivo, we developed a highly quantifiable model by transplanting OPCs in the retina. We find that myelination is greatly enhanced upon cyclopamine treatment and hypothesize that Shh/Smo could promote OPC proliferation to subsequently inhibit differentiation. Consistent with this hypothesis, we find that the genetic activation of Smo significantly increased numbers of OPCs and decreased oligodendrocyte differentiation when we examined the corpus callosum during development and after cuprizone demyelination and remyelination. However, upon loss of function with the conditional ablation of Smo, myelination in the same scenarios are unchanged. Taken together, our present findings suggest that the Shh pathway is sufficient to maintain OPCs in an undifferentiated state, but is not necessary for myelination and remyelination.

neuroscience↗

bioRxiv2023

Supporting central nervous system neuroprotection and remyelination by specific TLR4 antagonism

ApTOLL is an aptamer specifically designed to antagonize Toll-Like Receptor 4 (TLR4), a relevant actor for innate immunity involved in inflammatory responses in multiple sclerosis (MS) and other diseases. MS is a primary demyelinating, chronic, inmune and neurodegenerative disease of the central nervous system that normally debuts in young adults. The currently available therapeutic arsenal to treat MS is composed of immunomodulators but, to date, there are no (re)myelinating drugs available in clinics. Our present study shows cells expressing TLR4 in demyelinating lesions of MS patients (postmortem samples from cerebral cortex) and, as a derivative, we studied the effect of TLR4 inhibition with ApTOLL in animal models of MS (experimental autoimmune encephalomyelitis -EAE- and the cuprizone). The treatment with ApTOLL positively impacted the clinical symptomatology, and this was associated with better preservation plus restoration of myelin and oligodendrocytes in the demyelinated lesions of these animals, which suggests not only an immunomodulatory but also a remyelinating effect of the treatment with ApTOLL. This latter was corroborated on purified cultures of rodent and adult human oligodendrocyte precursor cells (OPCs), confirming the expression of TLR4 in this cell type. Altogether, the molecular nature of ApTOLL and its mechanism/s of action strongly supports this compound as a novel candidate to treat MS and other demyelinating scenarios.

neuroscience↗