Fig. 5
From: Circular HER2 RNA positive triple negative breast cancer is sensitive to Pertuzumab
HER2–103 promoted EGFR/HER3 interaction and activation in TNBC. a Circ-HER2 shRNAs or scramble shRNA stably transfected MDA-MB-231 or MDA-MB-468 cells, as well as circ-HER2 and circ-HER2 noATG vector stably transfected MDA-MB-231 or MDA-MB-468 cells were subjected to immunoblotting with indicated antibodies. The transfection efficiency of above cells was verified by q-PCR. b circ-HER2 or circ-HER2 noATG vector stably transfected BT549 cells were subjected to immunoblotting with indicated antibodies. The transfection efficiency was verified both by q-PCR. c Immunoprecipitation using HER3 or EGFR antibodies were performed in circ-HER2 shRNAs or scramble shRNA stably transfected MDA-MB-468 cells with indicated antibodies. d Left, circ-HER2 was stably transfected to MDA-MB-468 cells, HER2–103 was immunoprecipitated, followed by immunoblotting with antibodies against EGFR and HER3; middle, EGFR and HER3 were immunoprecipitated in above mentioned cells, followed by immunoblotting with antibodies against HER2–103. Right, circ-HER2 was stably transfected to MDA-MB-231 cells, HER2–103 or EGFR were immunoprecipitated followed by immunoblotting with antibodies against EGFR and HER2–103. e Upper left, EGFR domains, namely, L1, CR1, L2, CR2, juxta-membrane (JM) segment and internal cellular domain (ICD). Upper right, HA-tagged EGFR domains and Flag-tagged HER2–103 were co-transfected into 293 T cells. Coimmunoprecipitated truncated EGFR protein was detected by anti-HA antibody after immunoprecipitation with anti-Flag antibody. Lower left, HER3 domains, namely, L1, CR1, L2, CR2, juxta-membrane (JM) segment and internal cellular domain (ICD). HA-tagged HER3 domains and Flag tagged HER2–103 were co-transfected into HEK293T cells. Coimmunoprecipitated truncated HER3 protein was detected by anti-HA antibody after immunoprecipitation with anti-Flag antibody. f HER2–103 was dose-dependently transfected into MDA-MB-468 cells. Interaction between EGFR/HER3 was determined mutually by immunoprecipitation, with indicated antibodies. g The kinase activity of EGFR pY1068 was detected following circ-HER2 stably transfection in MDA-MB-468 cells. Lines show the mean ± SD, ***, p < 0.001; **, p < 0.01; *p < 0.05. Data are representative from at least 2–3 experiments with similar results