Journal: Neurotherapeutics
Article Title: Targeting Talin1 to reprogram dendritic cell activation landscapes: A mechanistically grounded cell therapy for multiple sclerosis model
doi: 10.1016/j.neurot.2025.e00723
Figure Lengend Snippet: Talin1 knockdown compromises the key molecules activation or interaction of TLR4/MyD88/NF-κB pathway . BMDCs transduced with control (shN) or Talin1-knockdown (shT) lentiviral vectors were treated with LPS (1 μg/mL) for indicated times. a, Talin1 knockdown declined (in trend) the p-p65 level and IκB degradation, and significantly reduced expressions of p -IκB and p -IRAK4 in BMDCs detected by western blotting. b-c, Co-immunoprecipitation (co-IP) of K63-Ub or TLR4, followed by immunoblot analysis for TRAF6 or TLR4/MyD88/IRAK4. 10 % of the cell lysates used for co-IP were run as input controls. The data revealed that Talin1 knockdown remarkably reduced TRAF6 ubiquitination (K63-Ub) and inhibited IRAK4 recruitment to TLR4 without affecting MyD88 recruitment to TLR4 in BMDCs treated with LPS for 1 h. d-e, Confocal images and colocalization analysis of TLR4/MyD88 and TLR4/IRAK4 in BMDCs with different treatments (n = 6–7 cells/group). The data showed that Talin1 knockdown significantly inhibited TLR4/IRAK4 colocalization without affecting TLR4/MyD88 colocalization. Data are expressed as the mean ± SD. ∗P < 0.05, ∗∗P < 0.01.
Article Snippet: Samples were resolved by SDS-PAGE and immunoblotted using the following antibodies: TLR4 (1:500, Invitrogen), MyD88 (1:500, Novus Biologicals, Centennial, USA), IRAK4(1:500, Abcam), or TRAF6 (1:500, Novus Biologicals).
Techniques: Knockdown, Activation Assay, Transduction, Control, Western Blot, Immunoprecipitation, Co-Immunoprecipitation Assay, Ubiquitin Proteomics
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