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. Author manuscript; available in PMC: 2024 Sep 4.
Published in final edited form as: Cancer Immunol Res. 2024 Mar 4;12(3):350–362. doi: 10.1158/2326-6066.CIR-23-0496

Fig. 3∣. Galectin-4 binds LILRB3 and induces downstream signaling in an ITIM-dependent manner.

Fig. 3∣

(a) Co-immunoprecipitation (Co-IP) demonstrates that recombinant Fc-tagged extracellular domain of LILRB3 (LILRB3-ECD) but not human IgG (hIgG) attached to Protein A Dynabeads interacts with recombinant human galectin-4 (hGal4). (b) Binding kinetics of His-tagged human galectin-4 to immobilized LILRB3-ECD were measured using biolayer interferometry and determined using a 1:1 binding curve. (c) Representative Co-IP of LILRB3-specific phosphotyrosine (pY) Western blot of THP1-LILRB3 lysates after 10 minutes on coated HLA-DR (20 μg/mL) and coated human galectin-2 (20 μg/mL) or human galectin-4 (20 μg/mL). (d) Representative Co-IP of LILRB3-specific pY Western blot of THP1-LILRB3 lysates after indicated time points with soluble LPS (200 ng/mL) and coated and soluble human galectin-4 (20 μg/mL). (e) Representative Co-IP of LILRB3-specific pY and SHP1 Western blot of 293T cells transfected with SHP1/SHP2/hGal4/Lyn and LILRB3-WT or LILRB3 ITIM-mutated plasmids after 48 hours. Three technical replicates were performed for each condition and at least two independent experiments were performed for each condition.