Journal: Acta Neuropathologica Communications
Article Title: 4R-tau isoform induction via TDP-43 in neurons in response to insulin: converging signaling pathways with implications for neurodegenerative disease
doi: 10.1186/s40478-025-02174-x
Figure Lengend Snippet: Inhibition of CK1 reduces insulin-induced 4R-tau expression and TDP-43 phosphorylation in cortical neurons. A Representative Western blot (WB) of cortical neuron lysates pretreated for 1 h with kinase and phosphatase inhibitors prior to insulin (Ins) stimulation. Inhibitors included D4476 (CK1 inhibitor), DMAT (CK2 inhibitor), and okadaic acid (OA; PP1a/PP2a inhibitor), each tested at two concentrations (×1 and ×10). Detection with a 4R-tau –specific antibody reveals a marked reduction in the insulin-induced 4R-tau signal only in D4476 + Ins-treated neurons, with both concentrations showing strong inhibition. D4476 at ×1 significantly reduces 4R tau induction. B WB showing electrophoretic mobility of TDP-43 under the same treatment conditions as in (A). The mobility shift observed in Ins- and OA-treated samples (consistent with phosphorylation) is absent in D4476 + Ins-treated neurons, supporting CK1 involvement in TDP-43 phosphorylation. C Schematic of CK inhibitors used: D4476 selectively inhibits CK1, and DMAT inhibits CK2, both kinases implicated in TDP-43 phosphorylation D Representative WB showing the effect of different pre-incubation times (1, 2, and 4 h) with D4476 (×1) prior to insulin stimulation. A time-dependent decrease in 4R-tau signal is observed, indicating progressive inhibition of insulin-induced expression. Right: Quantification of 4R-tau levels normalized to tubulin from three independent experiments. Insulin treatment significantly increases 4R-tau expression compared to control, and this induction is progressively inhibited by D4476 pretreatment (**** p < 0.0001, one-way ANOVA followed by Tukey’s post hoc test). No 4R-tau signal was detected in untreated (Ctr) or in the 4 h D4476 + Ins condition (nd). E Representative membranes showing 4R-tau (upper panel) and TDP-43 (lower panel) expression in control (Ctr), Ins, and D4476 (4 h pre-incubation) + Ins conditions. While 4R-tau levels increase upon insulin stimulation and are reduced by D4476, TDP-43 levels remain unchanged across conditions
Article Snippet: ; mouse NGF (nerve growth factor 2.5 S Alomone N-100 ) at 100 ng/ml for 60 min as in [ ]; Okadaic Acid ( OA ) as PP1a and PP2a 10 nM and 100 nM 24 h before incubation with insulin as used in [ ] ; CK1 inhibitor D4776 (Medchemexpress, stock 1 mg/25 ml DMSO) 100 μM and 1000 μM 1 h before other treatment was used at the following concentrations before incubation with insulin as used in [ ]; CK2 inhibitor DMAT (Medchemexpress, HY-15535, (stock 1 mg/DMSO) 20 μM and 200 μM, 24 h incubation before other treatment, as previously used in [ ].
Techniques: Inhibition, Expressing, Phospho-proteomics, Western Blot, Mobility Shift, Incubation, Control
MedChemExpress is a verified supplier 
MedChemExpress manufactures this product 
