porcine tubulin  (Cytoskeleton Inc)

Journal: Nature Communications

Article Title: CPT1A-mediated IDO1 succinylation shapes EGFRvIII-driven resistance to tumor electric field therapy in glioblastoma

doi: 10.1038/s41467-026-77927-w

Figure Lengend Snippet: A Western blot analysis of AhR in cytoplasmic and nuclear fractions of TBD0220 and LN229-EGFRvIII cells. TEFT treatment promotes AhR nuclear translocation, B Immunofluorescence staining of AhR (red) and DAPI (blue). TEFT induces AhR nuclear accumulation, which is reversed by IDO1 knockdown or osimertinib. Scale bars, 10 μm, C ATAC-seq footprinting showing increased chromatin accessibility at the AhR binding motif in EGFRvIII cells versus wild-type, D GO enrichment analysis of AhR target genes (identified by intersecting ATAC-seq and ChIP-Atlas), highlighting “Mitotic Cell Cycle” and “Microtubule Cytoskeleton” as top pathways, E Representative immunofluorescence images of mitotic spindles (α-tubulin, green) and chromosomes (DAPI, blue) in LN229 cells. TEFT causes severe spindle disruption and multipolar mitosis in wild-type cells, whereas EGFRvIII cells maintain bipolar spindles. This protection is lost upon IDO1 knockdown or osimertinib treatment, F Quantification of cells with abnormal nuclei following TEFT treatment ( n = 3 biological replicates per group), G Venn diagram intersecting RNA-seq (upregulated), ATAC-seq (accessible peaks), and ChIP-Atlas (AhR binding targets) data, identifying DCLK1 as a key downstream target, H Western blot confirming the upregulation of DCLK1 protein in EGFRvIII cells upon TEFT, I Immunofluorescence images of mitotic spindles in DCLK1 -knockdown cells. Silencing DCLK1 sensitizes EGFRvIII cells to TEFT-induced spindle defects, J Quantification of abnormal nuclear morphology in DCLK1-depleted cells ( n = 3 biological replicates per group), K Venn diagram integrating multi-omics data with microtubule/GTPase-related genes, identifying ARHGEF2 (GEF-H1) as another target, L In vitro tubulin polymerization assay in TBD0220 cells under the indicated conditions. IDO1 knockdown, ARHGEF2 knockdown, osimertinib, or epacadostat reduced the microtubule-polymerizing activity associated with EGFRvIII-mediated TEFT resistance. Data are presented as mean ± SD. The exact n values and definitions of biological replicates are specified for the relevant panels. Statistical tests were as follows: two-tailed unpaired Student’s t test for panel J ; ordinary one-way ANOVA followed by two-sided Šídák-adjusted multiple-comparisons tests for panel F . Exact P values, adjusted for multiple comparisons where applicable, are shown in the corresponding panels. Source data are provided as a Source Data file.

Article Snippet: Tubulin polymerization kinetics were monitored using the Tubulin Polymerization Assay Kit (BK006P, Cytoskeleton, Inc.) according to the manufacturer’s protocol.

Techniques: Western Blot, Analysis, Translocation Assay, Immunofluorescence, Staining, Knockdown, Footprinting, Binding Assay, Disruption, RNA Sequencing, Morphology, Biomarker Discovery, In Vitro, Polymerization Assay, Activity Assay, Two Tailed Test

Journal: bioRxiv

Article Title: Programming Brain Cell-Type-Selective Delivery In Vivo with Transporter-Guided Therapeutics

doi: 10.64898/2026.06.14.732141

Figure Lengend Snippet: a. Confocal images of α-tubulin immunofluorescence (green) in NIH3T3 cells treated with vehicle, eExACT-2, Colchicine, or Colchicine–eExACT-2 at two concentrations (100 nM and 500 nM). The normal filamentous microtubule pattern seen in vehicle- and eExACT-treated cells is disrupted following treatment with Colchicine or Colchicine–eExACT-2, resulting in diffuse tubulin staining (green), altered cell morphology, and increased multinucleation (Hoechst, blue). Scale bars, 20 μm. b. Quantification of multinucleated cells across treatments and concentrations (N = 3–5 wells per group, with 3–5 images analyzed per well; total cells per group >50). ** indicates P<0.01, **** indicates P<0.0001, and ns indicates not significant (unpaired t-test). c. Fluorescence-based cell-free tubulin polymerization assay comparing the effects of Vehicle, eExACT-2, Colchicine–eExACT-2, and Colchicine (3 µM). Curves show mean fluorescence intensity over time (N = 3 technical replicates per group). For statistical analysis of panel C, Two-sample Kolmogorov-Smirnov test was used to assess overall differences between groups where ** indicates P=0.013 and **** indicates P<0.0001.

Article Snippet: To evaluate the effects of eExACT compounds on microtubule dynamics, we performed a one-step fluorescence-based tubulin polymerization assay (Cytoskeleton, Inc., #BK011P) as previously described .

Techniques: Immunofluorescence, Staining, Fluorescence, Polymerization Assay