Proceedings of The Physiological Society

Europhysiology 2018 (London, UK) (2018) Proc Physiol Soc 41, PCB112

Poster Communications

HER2 and p95HER2 differentially regulate miRNA expression, downregulate MYB proteins, and increase invasiveness through miR-221/222 and miR-503 in MCF-7 breast cancer cells

A. Gorbatenko1, R. Søkilde3, E. E. Sørensen1, I. Newie2, H. Persson2, T. Litman2, C. Rovira2, S. F. Pedersen1

1. Department of Biology, University of Copenhagen, København Ø, Denmark. 2. Department of Clinical Sciences Lund, Oncology and Pathology,, Lund University, Lund, Sweden. 3. Department of International Health, Immunology and Microbiology, University of Copenhagen, Copenhagen, Denmark.


The HER2 oncogene and its constitutively active, truncated form p95HER2 play central roles in breast cancer1. The phenotypes they elicit are not identical, yet the underlying differences are incompletely understood. The aim of this study was to characterize the impact of HER2 and p95HER2 on the miRNA signature of human breast cancer cells and the resulting downstream signaling events shaping cancer invasiveness. High-throughput miRNA profiling was carried out 15, 36 and 60 h after inducible HER2 or p95HER2 expression in MCF-7 luminal breast cancer cells, and central hits were validated by RT-qPCR. miRNAs which were strongly upregulated by p95HER2, yet not by HER2, included miR-221, miR-222, and miR-503. The c-Myb family transcription factors MYB and MYBL1, but not a third family member, MYBL2, were downregulated by p95HER2 as well as by miR-503- or miR-221/-222 mimics. A single point mutation in the MYBL1 3' untranslated region abolished the upregulation by miR221-/-222. p95HER2 expression, or knockdown (KD) of either MYB protein, elicited upregulation of tissue inhibitor of matrix metalloprotease- 2 (TIMP2), an important regulator of cancer invasiveness. Collectively, this suggests that p95HER2 expression elevates miR-221, -222, and -503 levels, in turn downregulating MYB and MYBL1 protein expression, which in turn favors upregulation of TIMP2. Similar results were obtained in T47D and SKBr-3 human breast cancer cells. To investigate the effect of this pathway on migration and invasion, vector- and p95HER2-overexpressing cells were subjected to Boyden chamber migration and invasion assays. miR-221/-222 and miR-503 mimics elicited markedly increased migration in both cell types. Invasion was also increased by miR-222 (1.5 fold higher), miR-221/-222 (2.5 fold higher) and miR-503 (4 fold higher) in p95HER2 cells (all normalized to vector neg. miR 40 nM), whereas in vector cells, these miRNAs had little or no effect on invasion (n=3). TIMP2 KD decreased invasion of both vector- and p95HER2 overexpressing cells (n=3). Collectively, this work reveals important differences between HER2- and p95HER2- mediated miRNA changes in breast cancer cells, and points to the existence of a signaling pathway from p95HER2 to miR-221/-222 and miR-503 via MYB proteins to TIMP2, which appears to contribute to breast cancer invasiveness.

Where applicable, experiments conform with Society ethical requirements