Journal: Frontiers in Oncology

Article Title: Chondroitin Sulfates Control Invasiveness of the Basal-Like Breast Cancer Cell Line MDA-MB-231 Through ROR1

doi: 10.3389/fonc.2022.914838

Figure Lengend Snippet: CS-E-elicited invasiveness is enhanced by the absence of DKK1. (A) Raw sensor grams. ROR1 was immobilized in a flow cell of a CM5 sensor chip. DKK1 alone (a) , DKK1 premixed with CS-E at a 1:7 molar ratio (b) , and DKK1 premixed with CS-A at a 1:7 molar ration (c) were used as analytes. (B) Response-unit quantification of binding. (C) DKK1 mRNA expression in MDA-MB-231 cells transfected with siDKK1 or control siRNA ( siCont ) measured using qPCR (n=4). Expression data were normalized to those of GAPDH . (D) Invasiveness of DKK1 knocked down MDA-MB-231 cells ( siDKK1 ) or control cells ( siCont ) treated with or without CS-E (n>5). Data were analyzed using a Tukey–Kramer multiple comparison.

Article Snippet: For ROR1 binding assays, WNT5A (0, 0.038, 0.075, 0.15, 0.30, and 0.60 μM), WNT5A/CS-E (0, 0.036/0.25, 0.071/0.50, 0.15/1.0, 0.29/2.0, and 0.57/4.0 μM), recombinant human DKK1 (Cat. No. 5439-DK/CF, R&D Systems) (0, 0.038, 0.075, 0.15, and 0.3 μM), or DKK1/CS-E (0, 0.038/0.28, 0.075/0.55, 0.15/1.1, and 0.3/2.1 μM) were sequentially injected at a flowrate of 30 μl/min for 120 s at 25°C; the dissociation time was set for 130 s. Binding reactions were performed in 50 mM Tris–HCl buffer (pH 7.5).

Techniques: Binding Assay, Expressing, Transfection, Control, Comparison

Journal: Frontiers in Oncology

Article Title: Chondroitin Sulfates Control Invasiveness of the Basal-Like Breast Cancer Cell Line MDA-MB-231 Through ROR1

doi: 10.3389/fonc.2022.914838

Figure Lengend Snippet: Schematic of CS-E enhancement of invasive activity of the triple-negative breast cancer MDA-MB-231 cell line. CS chains bind WNT5A and ROR1 through E units, signaling cancer cells to activate JNK1. Decreasing E units by knockdown of CHST11 and CHST15 inhibits WNT5A−ROR1−JNK signaling. DKK1 suppresses CS tumor promoting activity by binding to E units.

Article Snippet: For ROR1 binding assays, WNT5A (0, 0.038, 0.075, 0.15, 0.30, and 0.60 μM), WNT5A/CS-E (0, 0.036/0.25, 0.071/0.50, 0.15/1.0, 0.29/2.0, and 0.57/4.0 μM), recombinant human DKK1 (Cat. No. 5439-DK/CF, R&D Systems) (0, 0.038, 0.075, 0.15, and 0.3 μM), or DKK1/CS-E (0, 0.038/0.28, 0.075/0.55, 0.15/1.1, and 0.3/2.1 μM) were sequentially injected at a flowrate of 30 μl/min for 120 s at 25°C; the dissociation time was set for 130 s. Binding reactions were performed in 50 mM Tris–HCl buffer (pH 7.5).

Techniques: Activity Assay, Knockdown, Binding Assay