Journal: bioRxiv

Article Title: Catestatin ameliorates tauopathy and amyloidogenesis via adrenergic inhibition

doi: 10.64898/2026.01.04.697519

Figure Lengend Snippet: CST levels are reduced across Tauopathies, and AD-spectrum disorders, and CST supplementation suppresses pathological Tau accumulation. CST quantification in human Alzheimer’s disease (AD) cortex shows significantly reduced CST in the (A) prefrontal cortex (PFC) and (C) hippocampus/entorhinal cortex (HC/EC) of Braak VI (n=16) vs. Braak 0–II (n=14) subjects. CST levels positively correlate with Mini-Mental State Examination (MMSE) scores in (B) PFC and (D) HC/EC, linking CST deficiency to cognitive decline. (E&F) Immunostaining of CST in postmortem tissue reveals reduced CST level in Braak VI (n=8) hippocampus compared with Braak 0–II (n=8); quantification shown in ( F ). Scale bar = 100µm (G&H) CST levels are also significantly reduced in cortical tissue from ( G ) corticobasal degeneration (CBD; Braak 0: n=16, Braak 3: n=19) and ( H ) basal ganglia from progressive supranuclear palsy (PSP; Braak 0: n=15, Braak 3: n=19), indicating CST loss as a shared feature of 4R-Tauopathies. (I) In cortical neuron cultures transduced with AAV2–P301S hTau, CST treatment decreases hyperphosphorylated Tau species (S202/T205 and S396/S404) without altering total Tau (n=3). (J–K) Immunocytochemistry confirms CST-mediated suppression of MC1+ misfolded Tau in P301S hTau–expressing neurons; quantified as MC1/MAP2+ area in (n=4). Scale bar = 50µm. (L–N) In organotypic hippocampal slice cultures (OTSC) expressing AAV2–P301S hTau (n=4), ( L ) CST reduces pTau (S202) and pTau (S396/S404) accumulation and ( M&N ) diminishes MC1+ Tau pathology in DG and CA3 regions. Scale bar = 200µm *Data are mean ± SEM; statistical significance indicated as *p < 0.05, **p < 0.01, ***p < 0.001, ***p < 0.0001.

Article Snippet: HEK 293 cells expressing the human Tau RD P301S FRET biosensor (ATCC CRL-3275) were plated onto Millipore EZ chambered slides at a confluency of 1 × 10 3 cells per well in complete DMEM (DMEM supplemented with 10% Fetal bovine serum and 100μg/mL penicillin-streptomycin).

Techniques: Immunostaining, Transduction, Immunocytochemistry, Expressing

Journal: bioRxiv

Article Title: Catestatin ameliorates tauopathy and amyloidogenesis via adrenergic inhibition

doi: 10.64898/2026.01.04.697519

Figure Lengend Snippet: (A) Cortical epinephrine (EPI) levels are markedly elevated in Sal/PS19 compared with nTg controls, and CST treatment significantly lowers EPI concentrations (nTg: n=7, Sal/PS19: n=18, CST/PS19: n=15). (B) Cortical EPI levels are markedly elevated in Sal/5xFAD compared with nTg controls, and CST treatment significantly lowers EPI concentrations (nTg: n=7, Sal/5xFAD: n=6, CST/5xFAD: n=6). (C–E) CST suppresses pathological PKA signaling in the PS19 hippocampus. ( C ) Immunoblots show increased phospho-PKA substrates and elevated p-Tau (S202/T205) in Sal/PS19 mice; CST treatment reduces both, while total Tau and PKA-Cα remain unchanged. ( D&E ) Quantification confirms CST-mediated reduction of PKA activity and Tau phosphorylation (nTg: n=3, Sal/PS19: n=6, CST/PS19: n=5). (F) Schematic illustrating CST blockade ADR–driven PKA activation. In AAV-MAPT (P301S/K18)–transduced hippocampal slices, EPI strongly activates PKA signaling and enhances Tau phosphorylation, whereas CST antagonizes adrenergic/PKA hyperactivation and prevents formation of misfolded Tau species. (G–I) CST counteracts EPI-induced PKA activation and Tau pathology. ( G ) Immunoblot analysis shows that EPI markedly increases phospho-PKA substrates and p-Tau species (S202, S396/404), CST alone has minimal effect on PKA activity. Still, CST prevents EPI-induced PKA hyperactivation (PBS: n=3, CST: n=3, EPI: n=3, CST+EPI: n=3). ( H&I ) Quantification demonstrates CST’s ability to blunt adrenergic/PKA overactivation and its downstream Tau phosphorylation signatures. (J) Immunofluorescence staining (CA3, DG) reveals substantial accumulation of misfolded Tau in EPI-treated slices, which is significantly reduced by CST and nearly abolished by CST+EPI (PBS: n=3, CST: n=3, EPI: n=3, CST+EPI: n=3). Scale bar = 200µM. A.U.: Arbitrary Unit. Data shown as mean ± SEM; significance indicated as *p < 0.05, **p < 0.01, *** p < 0.001, ****p < 0.0001; NS, not significant.

Article Snippet: HEK 293 cells expressing the human Tau RD P301S FRET biosensor (ATCC CRL-3275) were plated onto Millipore EZ chambered slides at a confluency of 1 × 10 3 cells per well in complete DMEM (DMEM supplemented with 10% Fetal bovine serum and 100μg/mL penicillin-streptomycin).

Techniques: Western Blot, Activity Assay, Phospho-proteomics, Activation Assay, Immunofluorescence, Staining

Journal: iScience

Article Title: Robust calibration and quantification of FRET signals using multiplexed biosensor barcoding

doi: 10.1016/j.isci.2025.113743

Figure Lengend Snippet: Construction of FRET calibration standards (A) Construction of FRET-ON calibration standard by Y349F mutation of Cyto-FAK. (B) Plot of YFP/CFP ratio of CytoFAK and FRET-ON (mean ± SEM, n = 20 cells) in response to 100 ng/mL EGF added at 6 min. (C) Plot of YFP/CFP ratio of CytoFAK and FRET-ON (mean ± SEM, n = 20 cells) in response to FAK inhibitor VS-6063 over 24 h. (D) Construction of FRET-OFF calibration standard by removal of the H3 domain of the H3K9me9 (W45A) biosensor. (E) Plot of YFP/CFP ratio of FRET-OFF and H3K9me3 biosensor (mean ± SEM, n = 20 cells) in response to 5 μM TCP over 48 h.

Article Snippet: H3K9me3 (W45A) biosensor , Addgene , Cat#120808.

Techniques: Mutagenesis

Journal: iScience

Article Title: Robust calibration and quantification of FRET signals using multiplexed biosensor barcoding

doi: 10.1016/j.isci.2025.113743

Figure Lengend Snippet: Construction of FRET calibration standards (A) Construction of FRET-ON calibration standard by Y349F mutation of Cyto-FAK. (B) Plot of YFP/CFP ratio of CytoFAK and FRET-ON (mean ± SEM, n = 20 cells) in response to 100 ng/mL EGF added at 6 min. (C) Plot of YFP/CFP ratio of CytoFAK and FRET-ON (mean ± SEM, n = 20 cells) in response to FAK inhibitor VS-6063 over 24 h. (D) Construction of FRET-OFF calibration standard by removal of the H3 domain of the H3K9me9 (W45A) biosensor. (E) Plot of YFP/CFP ratio of FRET-OFF and H3K9me3 biosensor (mean ± SEM, n = 20 cells) in response to 5 μM TCP over 48 h.

Article Snippet: H3K9me3 biosensor , Addgene , Cat#120802.

Techniques: Mutagenesis

Journal: Cell death & disease

Article Title: RhoA balances microglial reactivity and survival during neuroinflammation.

doi: 10.1038/s41419-023-06217-w

Figure Lengend Snippet: Fig. 5 Inflammation disrupts Ca2+, pH, and mitochondrial homeostasis in RhoA-deficient microglia. A Lifeact fluorescence labeling in RhoA WT, RhoA T19N, or RhoA KO HMC3 microglia (n = 3 independent experiments). B Cell counting in RhoA WT, RhoA Q63L, RhoA T19N, or RhoA KO HMC3 microglia (n = 8 independent experiments). Graph displays mean with SD. ****p < 0.0001 (Two-way ANOVA). RhoA WT, RhoA T19N, or RhoA KO HMC3 microglia expressing a global Ca2+ biosensor (C), mitochondrial Ca2+ biosensor (D), endoplasmic reticulum Ca2+ biosensor (E), pH biosensor (F), MitoTimer biosensor (G), or mitophagy biosensor (H) and exposed to LPS (1 µg/ml; 20 min (C-F) or 60 min (G and H)). Graphs (mean and SD) display fluorescence changes (n = 15 cells per group from 3 independent experiments for each biosensor). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 (Two-way ANOVA). Scale bars: 20 µm.

Article Snippet: Raichu-RhoA (provided by M. Matsuda [27]), pTriEx-RhoA FLARE.sc Biosensor WT (RRID:Addgene_12150), pTriEx-RhoA FLARE.sc Biosensor Q63L (RRID:Addgene_12151), pTriEx-RhoA FLARE.sc Biosensor T19N (RRID:Addgene_12152), pRK5-myc-RhoA WT (RRID:Addgene_12962), pRK5-mycRhoA Q63L (RRID:Addgene_12964), pRK5-myc-RhoA T19N (RRID:Addgene_12963), EGFP-p65 (RRID:Addgene_111190), GW1-pHRed (RRID:Addgene_31473), GW1CMV-Perceval (RRID:Addgene_21737), Laconic/pcDNA3.1 (+) (RRID:Addgene_118627), Pyronic /pcDNA3.1 (+) (RRID:Addgene_51308), pcDNA3.1 FLII12Pglu-700uDelta6 (RRID:Addgene_17866), Cyto-ABKAR (RRID:Addgene_61510), pLentiEKAR2G2 (RRID:Addgene_40178), Kras-Src FRET biosensor (RRID:Addgene_78302), pFRET-HSP33 cys (RRID:Addgene_16076), pGP-CMV-GCaMP6F (RRID:Addgene_40755), mCherry-Lifeact-7 (RRID:Addgene_54491), pMitoTimer (RRID:Addgene_52659), pCLBW cox8 EGFP mCherry (RRID:Addgene_78520).

Techniques: Labeling, Cell Counting, Expressing