Journal: Nature cell biology

Article Title: The LINK-A lncRNA activates normoxic HIF1α signalling in triple-negative breast cancer.

doi: 10.1038/ncb3295

Figure Lengend Snippet: Figure 3 LINK-A mediates recruitment of BRK to GPNMB for kinase activation. (a) Immunofluorescence detection using the indicated antibodies in MDA-MB-231 cells harbouring control (upper panel) or LINK-A shRNA, followed by HB-EGF stimulation (upper panel). Scale bars, 20 µm. (b) Graphic illustration of the BRK– and LRRK2–LINK-A interactions (upper panel) and the corresponding deletion abolishing these interactions (lower panel). (c,d) Immunofluorescence imaging (c, scale bars, 20 µm) or immunoblot (IB) detection (d) was performed using the indicated antibodies in MDA-MB-231 cells transfected with LNA against LINK-A followed by overexpression of the indicated rescue plasmids with HB- EGF stimulation. The dotted line on the blots of d indicates the position where the images of single blots were vertically cropped to juxtapose non-adjacent lanes. (e) In vitro kinase assay using recombinant BRK and

Article Snippet: Wild-type or mutant substrate proteins were incubated with 50 μl of in vitro kinase assay buffer II (SignalChem) containing 100 μM ATP (cold reaction) or 10 μCi [γ-32P]ATP and the indicated protein kinase for 1 h at 30 ◦C.

Techniques: Activation Assay, Control, shRNA, Imaging, Western Blot, Transfection, Over Expression, In Vitro, Kinase Assay, Recombinant