Journal: Cells

Article Title: Loss of Ubiquitin Carboxy-Terminal Hydrolase L1 Impairs Long-Term Differentiation Competence and Metabolic Regulation in Murine Spermatogonial Stem Cells

doi: 10.3390/cells10092265

Figure Lengend Snippet: UCH-L1 expression in mouse testis. ( A ) Immunohistochemistry panel of UCH-L1, SOX9, SCP3 and DAPI at 0, 7, 17, 30 and 120 days old. Scale bar = 50 µm. ( B – M ) Immunohistochemistry panel of UCH-L1, LIN28A and DAPI. Arrows indicate Sertoli cells, determined by nuclear morphology, and stars indicate LIN28A positive cells. Scale bars = 50 µm.

Article Snippet: The sections were incubated with primary antibodies against PLZF (HPA001499, Sigma,), LIN28A (8641, Cell Signalling Technologies, Danvers, MA, USA), UCH-L1 (13179, Cell Signalling Technologies) and SOX9 (sc-17341, Santa Cruz Biotehnologies, Dallas, TX, USA,), followed by an incubation with species-specific secondary Alexa Fluor 488, 555 or 647 antibodies.

Techniques: Expressing, Immunohistochemistry

Journal: Cells

Article Title: Loss of Ubiquitin Carboxy-Terminal Hydrolase L1 Impairs Long-Term Differentiation Competence and Metabolic Regulation in Murine Spermatogonial Stem Cells

doi: 10.3390/cells10092265

Figure Lengend Snippet: Phenotype of Uch-l1 +/+ , Uch-l1 +/− and Uch-l1 −/− testes. ( A – C ) H&E histology images of Uch-l1 +/+ , Uch-l1 +/− and Uch-l1 −/− testes at 120 days old. Scale bar = 200 µm. ( D ) Body weight of Uch-l1 +/+ , Uch-l1 +/− and Uch-l1 −/− mice. ( E ) Weight of Uch-l1 +/+ , Uch-l1 −/− and Uch-l1 −/− testes. ( F , G ) 120-day old Uch-l1 +/+ and Uch-l1 −/− testis histology, respectively. Star indicates a degenerating seminiferous tubule. Scale bar = 200 µm. ( H ) Percentage of degenerating seminiferous tubules at 30, 60, 90 and 120 days of age in Uch-l1 +/+ and Uch-l1 −/− testes. N = 6/genotype/time point. ( I ) Litter size from Uch-l1 +/+ and Uch-l1 −/− male matings to Uch-l1 +/+ females. Individual data points represent one litter. * indicates significant differences ( p < 0.05).

Article Snippet: The sections were incubated with primary antibodies against PLZF (HPA001499, Sigma,), LIN28A (8641, Cell Signalling Technologies, Danvers, MA, USA), UCH-L1 (13179, Cell Signalling Technologies) and SOX9 (sc-17341, Santa Cruz Biotehnologies, Dallas, TX, USA,), followed by an incubation with species-specific secondary Alexa Fluor 488, 555 or 647 antibodies.

Techniques:

Journal: Cells

Article Title: Loss of Ubiquitin Carboxy-Terminal Hydrolase L1 Impairs Long-Term Differentiation Competence and Metabolic Regulation in Murine Spermatogonial Stem Cells

doi: 10.3390/cells10092265

Figure Lengend Snippet: Analysis of whole Uch-l1 −/− , Uch-l1 +/− and Uch-l1 +/+ testes. ( A ) Quantitative PCR results of significantly differentially expressed genes in 7, 17 and 120-day old of Uch-l1 −/− relative to Uch-l1 +/+ testes ( B , F ) Flow cytometry analysis of PLZF and c-KIT positive cells in Uch-l1 +/+ , Uch-l1 +/− and Uch-l1 −/− testes at 120 days of age. n = 6/genotype. ( B , C ) Representative dot plot of c-KIT and PLZF positive cells in Uch-l1 +/+ and Uch-l1 −/− testes, respectively. ( D ) Percentage of PLZF+ c-KIT- cells in Uch-l1 +/+ , Uch-l1 +/− and Uch-l1 −/− . ( E ) Percentage of PLZF+ c-KIT+ cells in Uch-l1 +/+ , Uch-l1 +/− and Uch-l1 −/− . ( F ) Percentage of c-KIT+ cells in Uch-l1 +/+ , Uch-l1 +/− and Uch-l1 −/− . * indicates significant differences ( p < 0.05), ** represents significant differences p < 0.01.

Article Snippet: The sections were incubated with primary antibodies against PLZF (HPA001499, Sigma,), LIN28A (8641, Cell Signalling Technologies, Danvers, MA, USA), UCH-L1 (13179, Cell Signalling Technologies) and SOX9 (sc-17341, Santa Cruz Biotehnologies, Dallas, TX, USA,), followed by an incubation with species-specific secondary Alexa Fluor 488, 555 or 647 antibodies.

Techniques: Real-time Polymerase Chain Reaction, Flow Cytometry

Journal: Cells

Article Title: Loss of Ubiquitin Carboxy-Terminal Hydrolase L1 Impairs Long-Term Differentiation Competence and Metabolic Regulation in Murine Spermatogonial Stem Cells

doi: 10.3390/cells10092265

Figure Lengend Snippet: Single-cell RNA sequencing analysis of Uch-l1 +/+ and Uch-l1 −/− spermatogonia. t-SNE analysis plot of single transcriptomes of Uch-l1 +/+ ( A ) and Uch-l1 −/− ( B ), showing unbiased cell clusters distinguished by color according to the key. ( C ) The feature plot shows the expression of some genes that allow the distinction between spermatogonia, spermatocytes and spermatids. ( D ) Heatmaps of the upregulated and downregulated gene sets signatures determined by GSEA in Uchl1 +/+ and Uch-l1 −/− .

Article Snippet: The sections were incubated with primary antibodies against PLZF (HPA001499, Sigma,), LIN28A (8641, Cell Signalling Technologies, Danvers, MA, USA), UCH-L1 (13179, Cell Signalling Technologies) and SOX9 (sc-17341, Santa Cruz Biotehnologies, Dallas, TX, USA,), followed by an incubation with species-specific secondary Alexa Fluor 488, 555 or 647 antibodies.

Techniques: RNA Sequencing, Expressing

Journal: Cells

Article Title: Loss of Ubiquitin Carboxy-Terminal Hydrolase L1 Impairs Long-Term Differentiation Competence and Metabolic Regulation in Murine Spermatogonial Stem Cells

doi: 10.3390/cells10092265

Figure Lengend Snippet: Integrative analysis across Uchl1 +/+ and Uch-l1 −/− undifferentiated spermatogonia transcriptomes. ( A ) UMAP analysis plot of single transcriptome of four merged samples (2 Uch-l1 +/+ and 2 Uch-l1 −/− ), showing unbiased cell clusters distinguished by color according to the key. ( B ) UMAP analysis showing individual samples grouped by color. ( C ) UMAP analysis plot showing the SPG1 (cluster 1) sub-clustered into seven distinct clusters (0 to 6). ( D ) UMAP analysis plot showing the two genotypes Uchl-1 +/+ and Uch-l1 −/− , separately. ( E ) GSEA showing enrichment of OXPHOS pathway in cluster 2 in Uch-l1 −/− . ( F ) Gene network of 11 genes contributing to the enrichment score of OXPHOS.

Article Snippet: The sections were incubated with primary antibodies against PLZF (HPA001499, Sigma,), LIN28A (8641, Cell Signalling Technologies, Danvers, MA, USA), UCH-L1 (13179, Cell Signalling Technologies) and SOX9 (sc-17341, Santa Cruz Biotehnologies, Dallas, TX, USA,), followed by an incubation with species-specific secondary Alexa Fluor 488, 555 or 647 antibodies.

Techniques:

Journal: Cells

Article Title: Loss of Ubiquitin Carboxy-Terminal Hydrolase L1 Impairs Long-Term Differentiation Competence and Metabolic Regulation in Murine Spermatogonial Stem Cells

doi: 10.3390/cells10092265

Figure Lengend Snippet: Transplantation of Uch-l1 +/+ and Uch-l1 −/− SSCs. ( A ) Recipient testis transplanted with Uch-l1 +/+ SSCs collected 2-months post transplantation. ( B ) Representative colony from recipient testis A. ( C ) Recipient testis transplanted with Uch-l1 −/− SSCs two months post transplantation. ( D ) Representative colony from recipient testis B. Scale bar = 1 mm. ( E – H ) Transplanted colony analysis. ( E ) Colony number and ( F ) colony length two months post transplantation ( n = 7 and 6 for Uch-l1 +/+ vs. Uch-l1 −/− SSCs transplantation, respectively). ( G ) Colony number and ( H ) total colonized length 10 months post transplantation ( n = 9 and 6 for Uch-l1 +/+ vs. Uch-l1 −/− SSCs transplantation, respectively). ( I ) Representative transplanted colony cross-sections of dark (left), medium (middle) and light (right) blue intensity of Uch-l1 −/− SSCs 10 months post transplantation. Scale bar = 100 µm. ( J ) Proportion of colonies with different degrees of differentiation (blue intensity) from Uch-l1 +/+ and Uch-l1 −/− SSCs 10 months post transplantation. * represents significant differences p < 0.05.

Article Snippet: The sections were incubated with primary antibodies against PLZF (HPA001499, Sigma,), LIN28A (8641, Cell Signalling Technologies, Danvers, MA, USA), UCH-L1 (13179, Cell Signalling Technologies) and SOX9 (sc-17341, Santa Cruz Biotehnologies, Dallas, TX, USA,), followed by an incubation with species-specific secondary Alexa Fluor 488, 555 or 647 antibodies.

Techniques: Transplantation Assay

Journal: EMBO Reports

Article Title: Muscle‐derived GDF15 drives diurnal anorexia and systemic metabolic remodeling during mitochondrial stress

doi: 10.15252/embr.201948804

Figure Lengend Snippet: A Schematic representation of HSA ‐Ucp1 ‐transgenic (TG) mouse line as model of compromised skeletal muscle‐specific mitochondrial OXPHOS capacity via respiratory uncoupling. B Uncoupled (LEAK) mitochondrial respiration of soleus (SOL) and extensor digitorum longus (EDL) muscle of wild‐type (WT) versus Ucp1 ‐TG (TG) mice (WT n = 9, TG n = 5). C Relative mRNA expression in quadriceps (Quad) of ISR components ( n = 8 per genotype). D, E Representative immunoblots of ISR component eIF2α and quantification of phospho‐eIF2α (p‐eIF2a Ser51 ) relative protein expression in quadriceps skeletal muscle (WT n = 3, TG n = 4). F, G Multi‐tissue transcriptomic profiling of Gdf15 gene expression. Heatmap is shown as raw ct expression values ( n = 4 per genotype). Quantification of Gdf15 mRNA expression in TG mice is shown as fold change compared to WT littermates (WT n = 5, TG n = 4). H Skeletal muscle (Quad and Gastroc) GDF15 protein content normalized to total protein content (WT n = 9, TG n = 18). I Ex vivo secretion of GDF15 from SOL and EDL muscle of WT versus TG mice ( n = 6 per genotype) after 2‐h incubation normalized to muscle wet weight (mg). J Mouse GDF15 plasma levels in post‐absorptive state of male and female WT versus TG mice at 20 weeks of age (male WT n = 8, TG n = 9; female WT n = 5, TG n = 6). K Mouse GDF15 plasma levels in post‐absorptive state of male WT versus TG mice fed low‐fat diet (LFD) or high‐fat diet (HFD) at 24 weeks of age (LFD WT n = 6, TG n = 5; HFD WT n = 5, TG n = 5). L Mouse GDF15 plasma levels in post‐absorptive state of young (10 weeks) versus old (95 weeks) male WT and TG mice (young WT n = 10, TG n = 13; old WT n = 13, TG n = 13). M Representative scheme of in vitro study design in differentiated C2C12 myocytes. N Gdf15 gene expression of differentiated C2C12 myotubes treated with vehicle control (Ctrl) or chemical mitochondrial uncoupler (FCCP, 1 μM versus 5 μM) for 5 h ( n = 3 biological replicates). Data information: Circulating plasma parameters are expressed as interleaved box and whiskers (min to max) plots, and all other data are expressed as means ± SEM; P‐ value calculated by unpaired t‐ test (A‐L) or one‐way ANOVA with Tukey's post hoc test (N); * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The following primary antibodies were used: phospho‐AMPK (Thr172) (#2531, Cell Signaling Technology), total AMPK (#2603, Cell Signaling Technology), phospho‐eIF2α (Ser51) (#3597, Cell Signaling Technology), eIF2α antibodies (#3524, Cell Signaling Technology), OXPHOS antibody (#ab110413, Abcam), slow myosin (#ab11083, Abcam), and UCP1 antibodies (#ab23841, Abcam).

Techniques: Transgenic Assay, Expressing, Western Blot, Gene Expression, Ex Vivo, Incubation, Clinical Proteomics, In Vitro, Control

Journal: EMBO Reports

Article Title: Muscle‐derived GDF15 drives diurnal anorexia and systemic metabolic remodeling during mitochondrial stress

doi: 10.15252/embr.201948804

Figure Lengend Snippet: A Genotyping PCR panel of Gdf15 and HSA‐ Ucp1 loci shown for wild‐type (WT), Gdf15 ‐KO (KO), Ucp1 ‐TG (TG), and Ucp1 ‐TGx Gdf15 ‐KO (TGxKO) mice. B Relative mRNA expression in quadriceps (Quad) of Ucp1 and Gdf15 in 20‐week‐old male WT ( n = 8), TG ( n = 7), and TGxKO ( n = 6) mice. C Plasma mGDF15 levels at 20 weeks of age (WT n = 9, TG n = 10, TGxKO n = 9). D Voluntary wheel running (VWR) activity shown hourly over 24 h at 15 weeks of age ( n = 4 per genotype). E Grip strength at 10–20 weeks of age (WT n = 15, TG n = 14, TGxKO n = 11). F Skeletal muscle mass relative to body lean mass of quadriceps (Quad) and gastrocnemius (Gastroc) from mice at 95 weeks of age (WT n = 8, TG n = 5, TGxKO n = 5). G, H Representative H&E histological staining of tibialis anterior (TA) muscle (G) (scale bars represent 50 μm), and cross‐sectional area (CSA) of myofibers (H) at 95 weeks (WT n = 12, TG n = 8, TGxKO n = 8). I Representative traces of oxygen consumption rate (OCR) during substrate–uncoupler–inhibitor titration (SUIT) protocol for mitochondrial respiratory capacity in permeabilized mouse soleus muscle fibers, PM (pyruvate+malate; LEAK respiration), ADP (OXPHOS capacity), Cyt c (cytochrome c, integrity of outer mt‐membrane), G (glutamate;), S (succinate), U (uncoupler, FCCP), Rot (rotenone), and Ama (antimycin A; less than 2% residual oxygen consumption, ROX). J–L Mitochondrial respiratory capacity (oxygen consumption rate, OCR) from oxidative soleus (SOL) (J) and glycolytic extensor digitorum longus (EDL) muscle fibers (K), and NetOXPHOS control ratio of 95‐week‐old mice (L) ( n = 5 per genotype). M Quadriceps (Quad) relative mRNA expression of ISR components at 95 weeks of age (WT n = 8, TG n = 5, TGxKO n = 5). N Representative immunoblots of ISR component eIF2α in quadriceps (Quad) muscle of 95‐week‐old mice. O Gastrocnemius enzyme activity of NAD(P)H quinone dehydrogenase 1 (NQO1) and total glutathione peroxidase (GPX) in 95‐week‐old male mice (WT n = 8, TG n = 5, TGxKO n = 5). Data information: Circulating plasma parameters are expressed as interleaved box and whiskers (min to max) plots, and all other data are expressed as means ± SEM; P‐ value calculated by one‐way ANOVA with Tukey's post hoc test; * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The following primary antibodies were used: phospho‐AMPK (Thr172) (#2531, Cell Signaling Technology), total AMPK (#2603, Cell Signaling Technology), phospho‐eIF2α (Ser51) (#3597, Cell Signaling Technology), eIF2α antibodies (#3524, Cell Signaling Technology), OXPHOS antibody (#ab110413, Abcam), slow myosin (#ab11083, Abcam), and UCP1 antibodies (#ab23841, Abcam).

Techniques: Expressing, Clinical Proteomics, Activity Assay, Staining, Titration, Membrane, Control, Western Blot

Journal: EMBO Reports

Article Title: Muscle‐derived GDF15 drives diurnal anorexia and systemic metabolic remodeling during mitochondrial stress

doi: 10.15252/embr.201948804

Figure Lengend Snippet: A Plasma GDF15 levels at 20 weeks of age (WT n = 9, KO n = 9). B Voluntary wheel running (VWR) activity shown hourly over 24 h at 15 weeks of age ( n = 4 per genotype). C Grip strength at 10–20 weeks of age (WT n = 15, KO n = 14). D Skeletal muscle mass relative to body lean mass of quadriceps (Quad), gastrocnemius (Gastroc), soleus, and EDL (WT n = 9, KO = 10). E Representative H&E histological staining of tibialis anterior (TA) muscle (G) (scale bars represent 50 μm) F–H Mitochondrial respiratory capacity (oxygen consumption rate, OCR) from oxidative soleus (SOL) (F) and glycolytic extensor digitorum longus (EDL) muscle fibers (G), and NetOXPHOS control ratio (H) ( n = 5 per genotype). I, J Representative immunoblots of OXPHOS in quadriceps (Quad) muscle (I) and their corresponding quantifications normalized to MFN2 (J) ( n = 6 per genotype). K Skeletal muscle (Quad) relative mRNA expression of ISR components (WT n = 8, KO n = 6). L Representative immunoblots of ISR component eIF2α and phospho‐eIF2α (p‐eIF2a Ser51 ), TG sample was included as positive control. M Skeletal muscle (Gastroc) enzyme activity of NQO1 and GPX (WT n = 8, KO n = 6). Data information: Data shown are from male wild‐type (WT) versus Gdf15 ‐KO (KO) mice. Circulating plasma parameters are expressed as interleaved box and whiskers (min to max) plots, and all other data are expressed as means ± SEM; P‐ value calculated by unpaired Student's t ‐test.

Article Snippet: The following primary antibodies were used: phospho‐AMPK (Thr172) (#2531, Cell Signaling Technology), total AMPK (#2603, Cell Signaling Technology), phospho‐eIF2α (Ser51) (#3597, Cell Signaling Technology), eIF2α antibodies (#3524, Cell Signaling Technology), OXPHOS antibody (#ab110413, Abcam), slow myosin (#ab11083, Abcam), and UCP1 antibodies (#ab23841, Abcam).

Techniques: Clinical Proteomics, Activity Assay, Staining, Control, Western Blot, Expressing, Positive Control

Journal: EMBO Reports

Article Title: Muscle‐derived GDF15 drives diurnal anorexia and systemic metabolic remodeling during mitochondrial stress

doi: 10.15252/embr.201948804

Figure Lengend Snippet: A Muscle mass relative to body lean mass of quadriceps (Quad), gastrocnemius (Gastroc), soleus, and extensor digitorum longus (EDL) muscles in 20‐week‐old male mice (WT n = 9, TG, n = 9, TGxKO = 10). B, C Representative H&E histological staining of tibialis anterior (TA) muscle (B), and cross‐sectional area (CSA) of myofibers (C) at 20 weeks (WT n = 8, TG n = 6, TGxKO n = 8), scale bars represent 50 μm. D–I Representative immunoblots and their corresponding quantifications (WT n = 8, TG n = 8, TGxKO n = 8), normalized to ATUB of p‐AMPK Thr172 /t‐AMPK (D, E), slow myosin (F, G), and OXPHOS (H, I) in quadriceps (Quad) muscle at 20 weeks of age. J Quantification of ISR component eIF2α and of phospho‐eIF2α (p‐eIF2a Ser51 ) relative protein expression in quadriceps skeletal muscle at 95 weeks of age (WT n = 8, TG n = 8, TGxKO n = 8). Data information: Data shown are from male wild‐type (WT), Ucp1 ‐TG (TG), and Ucp1 ‐TGx Gdf15 ‐KO (TGxKO) mice. Data are expressed as means ± SEM; P‐ value calculated by one‐way ANOVA with Tukey's post hoc test; * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The following primary antibodies were used: phospho‐AMPK (Thr172) (#2531, Cell Signaling Technology), total AMPK (#2603, Cell Signaling Technology), phospho‐eIF2α (Ser51) (#3597, Cell Signaling Technology), eIF2α antibodies (#3524, Cell Signaling Technology), OXPHOS antibody (#ab110413, Abcam), slow myosin (#ab11083, Abcam), and UCP1 antibodies (#ab23841, Abcam).

Techniques: Muscles, Staining, Western Blot, Expressing

Journal: EMBO Reports

Article Title: Muscle‐derived GDF15 drives diurnal anorexia and systemic metabolic remodeling during mitochondrial stress

doi: 10.15252/embr.201948804

Figure Lengend Snippet: A Relative quadriceps (Quad) mRNA expression of the ISR components Atf4, Atf5, Atf6, and Chop (WT day n = 9, WT night n = 5, TG day n = 20, TG night n = 12). B, C Representative immunoblots of ISR component eIF2α (B) and quantification of phospho‐eIF2α (p‐eIF2a Ser51 ) relative protein expression (C) in skeletal muscle (Gastroc, WT day n = 4, WT night n = 5, TG day n = 9, TG night n = 9). D, E Relative mRNA expression of Rev‐erba (D) and Gdf15 (E) in quadriceps, liver, and eWAT (WT day n = 9, WT night n = 5, TG day n = 20, TG night n = 12). F Skeletal muscle (Quad) GDF15 protein content normalized to total protein content (WT day n = 4, WT night n = 5, TG day n = 14, TG night n = 12) G Circulating GDF15 plasma levels (WT day n = 9, WT night n = 5, TG day n = 18, TG night n = 12). H Graphical summary showing muscle mitochondrial stress induced secretion and diurnal variation of GDF15 to drive a daytime‐restricted anorexia, affecting in turn metabolic flexibility, adiposity, WAT browning, and systemic insulin sensitivity. Parts of this cartoon were created using Servier Medical Art ( http://smart.servier.com ). Data information: Data shown are from male wild‐type (WT) versus Ucp1 ‐TG (TG) mice sacrificed at day (10am) or night (10 pm). Circulating plasma parameters are expressed as interleaved box and whiskers (min to max) plots, and all other data are expressed as means ± SEM; P‐ value calculated by one‐way ANOVA with Tukey's post hoc test; * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The following primary antibodies were used: phospho‐AMPK (Thr172) (#2531, Cell Signaling Technology), total AMPK (#2603, Cell Signaling Technology), phospho‐eIF2α (Ser51) (#3597, Cell Signaling Technology), eIF2α antibodies (#3524, Cell Signaling Technology), OXPHOS antibody (#ab110413, Abcam), slow myosin (#ab11083, Abcam), and UCP1 antibodies (#ab23841, Abcam).

Techniques: Expressing, Western Blot, Clinical Proteomics

Journal: Cell Death & Disease

Article Title: Deubiquitinase UCH-L1 confers paclitaxel resistance via stabilizing PKM2 to promote glycolysis in triple-negative breast cancer

doi: 10.1038/s41419-026-08521-7

Figure Lengend Snippet: A Boxplot of the UCH-L1 mRNA expression levels in four different subtypes of BRCA from the GEPIA2 database. B mRNA expression levels of UCH-L1 in TNBC and normal breast tissues from the GEPIA2 database. C IHC analysis of TNBC ( n = 40) and normal breast tissue samples ( n = 20) was performed using anti-UCH-L1 antibody. Representative images of IHC staining and quantified data presented in box plots are shown. Scale bar, 100 μm. D Western blot analysis of UCH-L1 protein expression level in normal breast cell and multiple TNBC cell lines. β-Actin was used as a loading control. E Kaplan-Meier plots for the overall survival of TNBC patients according to UCH-L1 mRNA expression from the Kaplan–Meier Plotter database. F Kaplan-Meier plots for the relapse-free survival of TNBC patients according to UCH-L1 protein expression from the Kaplan–Meier Plotter database. G Kaplan-Meier plots for the overall survival of chemotherapy-treated TNBC patients according to UCH-L1 mRNA expression from the Kaplan–Meier Plotter database. H The receiver operating characteristic (ROC) curve between UCH-L1 expression and therapeutic responses to taxane-containing NAC in breast cancer cohorts. I The correlation analysis between the efficacy of taxane-based chemotherapy and UCH-L1 expression. Chi-square test was performed to calculate statistical significance between two comparator groups. Stacked bar chart showed the proportion of patients with or without response in high or low UCH-L1 groups. * p < 0.05; *** p < 0.001.

Article Snippet: Antibodies used in immunoblotting: UCH-L1 (66230-1-Ig, 1:1000), PKM2 (60268-1-Ig, 1:1000), β-actin (66009-1-Ig, 1:5000), Myc (60003-2-Ig, 1:5000), HA (66006-2-Ig, 1:5000), Flag (66008-4-lg, 1:5000), GST (10000-0-AP, 1:4000) were purchased from Proteintech.

Techniques: Expressing, Immunohistochemistry, Western Blot, Control

Journal: Cell Death & Disease

Article Title: Deubiquitinase UCH-L1 confers paclitaxel resistance via stabilizing PKM2 to promote glycolysis in triple-negative breast cancer

doi: 10.1038/s41419-026-08521-7

Figure Lengend Snippet: A Knockdown efficiency of UCH-L1 in TNBC cells was evaluated by Western blot assay. B HCC1806 and BT549 cells with or without UCH-L1 depletion were incubated with indicated concentrations of PTX for 72 h, and cell viability was measured by CCK-8 assay. C Colony formation of HCC1806 and BT549 cells (500 cells per well) with or without UCH-L1 depletion in the presence of DMSO or 20 nM PTX. D HCC1806 and BT549 cells with or without UCH-L1 depletion were incubated with 20 nM PTX for 24 h, and cell proliferation was measured by EdU assay. Scale bars, 150μm. E HCC1806 and BT549 cells with or without UCH-L1 depletion were incubated with 20 nM PTX for 48 h, and cell apoptosis was measured by TUNEL assay. Scale bars, 150μm. F A schematic view of the animal study plan ( n = 6). G Curves of tumor growth. H Photopraphs of mice tumors of each group at the termination of the experiment. I Tumor weights were measured at the end of the experiment. J Characterization of HCC1806 xenograft tumors with histologic analysis by IHC staining of Ki67 and TUNEL. K The effect of treatment on mice body weight. Data are represented as mean ± SD of biological triplicates. * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: Antibodies used in immunoblotting: UCH-L1 (66230-1-Ig, 1:1000), PKM2 (60268-1-Ig, 1:1000), β-actin (66009-1-Ig, 1:5000), Myc (60003-2-Ig, 1:5000), HA (66006-2-Ig, 1:5000), Flag (66008-4-lg, 1:5000), GST (10000-0-AP, 1:4000) were purchased from Proteintech.

Techniques: Knockdown, Western Blot, Incubation, CCK-8 Assay, EdU Assay, TUNEL Assay, Immunohistochemistry

Journal: Cell Death & Disease

Article Title: Deubiquitinase UCH-L1 confers paclitaxel resistance via stabilizing PKM2 to promote glycolysis in triple-negative breast cancer

doi: 10.1038/s41419-026-08521-7

Figure Lengend Snippet: A Volcano plot of quantitative proteomics showed the differentially expressed genes between UCH-L1 knockdown and control HCC1806 cells. B KEGG pathway analysis for differentially expressed proteins in HCC1806 cells with or without UCH-L1 knockdown. C Effect of UCH-L1 knockdown on the ECAR of HCC1806 cells was detected using the Seahorse analyzer. D Quantification analysis of glycolysis, glycolytic capacity and glycolytic reserve levels in the ECAR assays. E - G Glucose consumption, lactate and ATP production were determined in control and UCH-L1 knockdown HCC1806 and BT549 cells. H - J Glucose consumption, lactate and ATP production were determined in MDA-MB-231 cells with or without UCH-L1 overexpression. K - L Colony formation of MDA-MB-231 cells with or without UCH-L1 overexpression in the presence of DMSO or PTX and 2-DG. M MDA-MB-231 cells from the indicated groups were incubated with PTX for 72 h, and cell viability was measured by CCK-8 assay. N MDA-MB-231 cells stably expressing empty vector (EV) or flag-tagged UCH-L1 were incubated with DMSO or PTX and 2-DG for 24 h, and cell proliferation was measured using EdU. Scale bars, 150 μm. O MDA-MB-231 cells with or without UCH-L1 overexpression were incubated with DMSO or PTX and 2-DG, and cell apoptosis was measured using TUNEL assay. Scale bars, 100 μm. Results shown are representative of three independent experiments. Data are represented as mean ± SD of biological triplicates. * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: Antibodies used in immunoblotting: UCH-L1 (66230-1-Ig, 1:1000), PKM2 (60268-1-Ig, 1:1000), β-actin (66009-1-Ig, 1:5000), Myc (60003-2-Ig, 1:5000), HA (66006-2-Ig, 1:5000), Flag (66008-4-lg, 1:5000), GST (10000-0-AP, 1:4000) were purchased from Proteintech.

Techniques: Quantitative Proteomics, Knockdown, Control, Over Expression, Incubation, CCK-8 Assay, Stable Transfection, Expressing, Plasmid Preparation, TUNEL Assay

Journal: Cell Death & Disease

Article Title: Deubiquitinase UCH-L1 confers paclitaxel resistance via stabilizing PKM2 to promote glycolysis in triple-negative breast cancer

doi: 10.1038/s41419-026-08521-7

Figure Lengend Snippet: A The top twenty UCH-L1-interacting proteins according to the number of identified unique peptides by Co-IP-MS assay. B HEK293T cells were transfected with Flag-PKM2 and Myc-UCH-L1 plasmids, and then subjected to immunoprecipitation with anti-Flag or anti-Myc antibodies. The lysates and immunoprecipitates were then blotted. C Immunoprecipitation analysis with the indicated antibodies was performed to detect endogenous UCH-L1 and PKM2 interaction in BT549 cells. D Immunofluorescence assay to confirm the colocalization of UCH-L1 and PKM2 in HCC1806 cells. E Purified recombinant GST-UCH-L1 interacts with PKM2. GST-UCH-L1 and GST proteins were pulled down with glutathione beads. PKM2 was detected by Western blot. F Schematic representation of full-length (FL) UCH-L1, PKM2 and different truncation mutants. G Myc-UCH-L1 FL or indicated truncation mutants were co-expressed with Flag-PKM2 in HEK293T cells. Flag-PKM2 was immunoprecipitated with anti-Flag antibody, followed by immunoblotting analysis of Myc-UCH-L1 using Myc antibody. H Flag-PKM2 FL or indicated truncation mutants were co-expressed with Myc-UCH-L1 in HEK293T cells. Myc-UCH-L1 was immunoprecipitated with anti-Myc antibody, followed by immunoblotting analysis of Flag-PKM2 using Flag antibody. I MDA-MB-231 cells transfected with the indicated plasmid were incubated with PTX for 72 h, and cell viability was measured by CCK-8 assay. J , K MDA-MB-231 cells with UCH-L1 overexpression were transfected with PKM2 siRNA, and cell viability was detected by CCK-8 assay after 72 h incubation of the indicated concentrations of PTX. Results shown are representative of three independent experiments. Data are represented as mean ± SD of biological triplicates. * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: Antibodies used in immunoblotting: UCH-L1 (66230-1-Ig, 1:1000), PKM2 (60268-1-Ig, 1:1000), β-actin (66009-1-Ig, 1:5000), Myc (60003-2-Ig, 1:5000), HA (66006-2-Ig, 1:5000), Flag (66008-4-lg, 1:5000), GST (10000-0-AP, 1:4000) were purchased from Proteintech.

Techniques: Co-Immunoprecipitation Assay, Transfection, Immunoprecipitation, Immunofluorescence, Purification, Recombinant, Western Blot, Plasmid Preparation, Incubation, CCK-8 Assay, Over Expression

Journal: Cell Death & Disease

Article Title: Deubiquitinase UCH-L1 confers paclitaxel resistance via stabilizing PKM2 to promote glycolysis in triple-negative breast cancer

doi: 10.1038/s41419-026-08521-7

Figure Lengend Snippet: A Immunoblotting analysis of PKM2 and UCH-L1 in UCH-L1 knockdown TNBC cells. B Immunoblotting analysis of PKM2 and UCH-L1 in MDA-MB-231 and HEK293T cells transfected with increasing amounts of UCH-L1 WT or C90S. C HCC1806 cells with UCH-L1 depletion were transfected with UCH-L1 WT or C90S, and PKM2 expression was detected. D Immunoblotting analysis of PKM2 and UCH-L1 in UCH-L1 knockdown TNBC cells treated with or without MG132. E MDA-MB-231 cells transfected with UCH-L1 WT or C90S were treated with CHX for the indicated time. F Immunoblotting analysis of PKM2 and UCH-L1 in UCH-L1 knockdown TNBC cells treated with or without CHX for the indicated time. PKM2 expression was detected by Western blot. Quantitation of PKM2 protein level based on band intensity was shown. Results shown are representative of three independent experiments.

Article Snippet: Antibodies used in immunoblotting: UCH-L1 (66230-1-Ig, 1:1000), PKM2 (60268-1-Ig, 1:1000), β-actin (66009-1-Ig, 1:5000), Myc (60003-2-Ig, 1:5000), HA (66006-2-Ig, 1:5000), Flag (66008-4-lg, 1:5000), GST (10000-0-AP, 1:4000) were purchased from Proteintech.

Techniques: Western Blot, Knockdown, Transfection, Expressing, Quantitation Assay

Journal: Cell Death & Disease

Article Title: Deubiquitinase UCH-L1 confers paclitaxel resistance via stabilizing PKM2 to promote glycolysis in triple-negative breast cancer

doi: 10.1038/s41419-026-08521-7

Figure Lengend Snippet: A Flag-PKM2 and HA-ubiquitin (HA-Ub) were co-expressed with Myc-UCH-L1 WT or C90S in HEK293T cells. After MG132 treatment, IP was performed with Flag antibody, followed by immunoblotting with the indicated antibodies. B HA-Ub was co-expressed with Myc-UCH-L1 WT or C90S in MDA-MB-231 cells. After MG132 treatment, IP was performed with PKM2 antibody, followed by immunoblotting with indicated antibodies. C HCC1806 cells with or without UCH-L1 depletion were transfected with HA-Ub. IP was performed with PKM2 antibody after MG132 treatment, followed by immunoblotting with indicated antibodies. D Ubiquitinated PKM2 was purified from HEK293T cells transfected with HA-Ub, and then incubated with GST-tagged wild-type UCH-L1, and UCH-L1 C90S proteins in reaction buffer. The reactions were terminated by boiling in 1X SDS sample buffer and analyzed using Western blot with anti-HA antibody. E HA-WT, K6, K11, K27, K29, K33, K48, or K63 Ub were co-transfected with Flag-PKM2 and Myc-UCH-L1 into HEK293T cells. Cell lysates were collected after MG132 treatment for 8 h and subjected to ubiquitination assay, and the ubiquitination level of PKM2 was detected by HA antibody. F Flag-PKM2 (WT or indicated KR mutants) and HA-Ub were co-expressed in 293T cells with or without UCH-L1 knockdown. IP was performed with Flag antibody after MG132 treatment for 8 h, followed by immunoblotting with anti-HA. G HCC1806 cells expressing Flag-PKM2 WT or indicated KR mutants were transfected with UCH-L1 siRNA, followed by immunoblotting with indicated antibodies. H HCC1806 cells expressing indicated KR mutants of Flag-PKM2 were transfected with UCH-L1 siRNA, and then treated with CHX for the indicated time. Cell lysates were collected and subjected to immunoblotting analysis. Quantitation of Flag-PKM2 protein level based on band intensity was shown. Results shown are representative of three independent experiments.

Article Snippet: Antibodies used in immunoblotting: UCH-L1 (66230-1-Ig, 1:1000), PKM2 (60268-1-Ig, 1:1000), β-actin (66009-1-Ig, 1:5000), Myc (60003-2-Ig, 1:5000), HA (66006-2-Ig, 1:5000), Flag (66008-4-lg, 1:5000), GST (10000-0-AP, 1:4000) were purchased from Proteintech.

Techniques: Ubiquitin Proteomics, Western Blot, Transfection, Purification, Incubation, Knockdown, Expressing, Quantitation Assay

Journal: Cell Death & Disease

Article Title: Deubiquitinase UCH-L1 confers paclitaxel resistance via stabilizing PKM2 to promote glycolysis in triple-negative breast cancer

doi: 10.1038/s41419-026-08521-7

Figure Lengend Snippet: A Dose–response curves of HCC1806 and HCC1806/TaxR cells. The IC50 was determined according to a dose vs. response curve by GraphPad Prism. B Immunoblotting analysis of PKM2 and UCH-L1 in HCC1806 and HCC1806/TaxR cells. C The glucose consumption, lactate and ATP production were determined and compared in HCC1806 and HCC1806/TaxR cells. D HCC1806/TaxR cells from the indicated groups were incubated with PTX for 72 h, and cell viability was measured by CCK-8 assay. E Cell apoptosis of HCC1806/TaxR cells with indicated treatment was measured by TUNEL assay. Scale bars, 150 μm. F HCC1806/TaxR cells with or without UCH-L1 depletion were incubated with 20 nM PTX for 24 h, and cell proliferation was measured by EdU assay. Scale bars, 150 μm. G Colony formation of HCC1806/TaxR cells (300 cells per well) with or without UCH-L1 depletion in the presence of DMSO or 20 nM PTX. HCC1806/TaxR cells were subcutaneously injected into 6-week-old female nude mice and subjected to PTX treatment as planned ( n = 5). Tumor sizes were measured and calculated as curves shown ( H ). The mice tumors of each group were photographed at the termination of the experiment (I). Tumor weights were measured at the end of the experiment ( J ). Characterization of HCC1806/TaxR xenograft tumors with histologic analysis by IHC staining of Ki67 and TUNEL ( K ). Data are represented as mean ± SD of biological triplicates. * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: Antibodies used in immunoblotting: UCH-L1 (66230-1-Ig, 1:1000), PKM2 (60268-1-Ig, 1:1000), β-actin (66009-1-Ig, 1:5000), Myc (60003-2-Ig, 1:5000), HA (66006-2-Ig, 1:5000), Flag (66008-4-lg, 1:5000), GST (10000-0-AP, 1:4000) were purchased from Proteintech.

Techniques: Western Blot, Incubation, CCK-8 Assay, TUNEL Assay, EdU Assay, Injection, Immunohistochemistry

Journal: Cell Death & Disease

Article Title: Deubiquitinase UCH-L1 confers paclitaxel resistance via stabilizing PKM2 to promote glycolysis in triple-negative breast cancer

doi: 10.1038/s41419-026-08521-7

Figure Lengend Snippet: A Boxplot of the PKM2 mRNA expression levels in TNBC and normal breast tissues from the GEPIA2 database. B Kaplan-Meier plots for the overall survival of chemotherapy-treated TNBC patients according to PKM2 mRNA expression from the Kaplan–Meier Plotter database. C The PKM2 expressions in non-responder and responder groups in breast cancer cohorts from ROC plotter server. The clinical response was evaluated based on pathological response. D Representative images of IHC of UCH-L1 and PKM2 in 40 TNBC tissues. The correlation analysis between UCH-L1 and PKM2 expression was performed using Fisher’s Exact Test. E Quantitative expression of IHC staining of PKM2 in high or low UCH-L1 groups. F Quantitative expression of IHC staining of UCH-L1 in high or low PKM2 groups. G The positive correlation between UCH-L1 and PKM2. The Pearson correlation test was used. H Mechanistic diagram of UCH-L1-catalyzed PKM2 deubiquitination in the regulation of glycolysis and chemoresistance. Depletion of UCH-L1 led to PKM2 degradation and downregulation, thereby inhibiting glycolysis and re-sensitizing TaxR-TNBC cells to chemotherapy.

Article Snippet: Antibodies used in immunoblotting: UCH-L1 (66230-1-Ig, 1:1000), PKM2 (60268-1-Ig, 1:1000), β-actin (66009-1-Ig, 1:5000), Myc (60003-2-Ig, 1:5000), HA (66006-2-Ig, 1:5000), Flag (66008-4-lg, 1:5000), GST (10000-0-AP, 1:4000) were purchased from Proteintech.

Techniques: Expressing, Immunohistochemistry

Journal: Cell reports

Article Title: Mass Cytometry Reveals Global Immune Remodeling with Multi-lineage Hypersensitivity to Type I Interferon in Down Syndrome

doi: 10.1016/j.celrep.2019.10.038

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Mouse monoclonal anti-human CD45RO (clone UCHL1) , Fluidigm , Cat#3164007B; RRID:AB_2811092.

Techniques: Recombinant, Mass Cytometry, Software