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

Andrej Gorbatenko,Joaquin Arribas,Stine Falsig Pedersen,Ester E Sorensen,Thomas Litman,Beatriz Morancho,Helena Persson,Inga Newie,Rolf Søkilde,Carlos Rovira

doi:10.1038/s41598-019-39733-x

Abstract

The HER2 oncogene and its truncated form p95HER2 play central roles in breast cancer. Here, we show that although HER2 and p95HER2 generally elicit qualitatively similar changes in miRNA profile in MCF-7 breast cancer cells, a subset of changes are distinct and p95HER2 shifts the miRNA profile towards the basal breast cancer subtype. High-throughput miRNA profiling was carried out 15, 36 and 60 h after HER2 or p95HER2 expression and central hits validated by RT-qPCR. miRNAs strongly regulated by p95HER2 yet not by HER2, included miR-221, miR-222, miR-503, miR-29a, miR-149, miR-196 and miR-361. Estrogen receptor-α (ESR1) expression was essentially ablated by p95HER2 expression, in a manner recapitulated by miR-221/-222 mimics. c-Myb family transcription factors MYB and MYBL1, but not MYBL2, were downregulated by p95HER2 and by miR-503 or miR-221/-222 mimics. MYBL1 3′UTR inhibition by miR-221/222 was lost by deletion of a single putative miR-221/222 binding sites. p95HER2 expression, or knockdown of either MYB protein, elicited upregulation of tissue inhibitor of matrix metalloprotease-2 (TIMP2). miR-221/222 and -503 mimics increased, and TIMP2 knockdown decreased, cell migration and invasion. A similar pathway was operational in T47D- and SKBr-3 cells. This work reveals important differences between HER2- and p95HER2- mediated miRNA changes in breast cancer cells, provides novel mechanistic insight into regulation of MYB family transcription factors by p95HER2, and points to a role for a miR-221/222– MYB family–TIMP2 axis in regulation of motility in breast cancer cells.

Highlights

The receptor tyrosine kinase HER2 (ErbB2) is overexpressed or amplified in 20–30% of breast cancer patients, correlating with cancer aggressiveness and reduced patient survival[1]
We employed generation sequencing to study the miRNA expression profiles induced by these HER2 variants and we interrogated whether p95HER2-induced cellular traits associated with breast cancer aggressiveness involve p95HER2 mediated regulation of miR-221/222 and miR-503 and in turn myeloblastosis viral oncogene homolog (MYB) family transcription factors
Our findings identify a novel role for p95HER2-specific miRNA changes in distinguishing p95HER2 positive cancers from those overexpressing only HER2, and uncover a signaling axis from miR-221/222 and -503 over MYB proteins to tissue inhibitor of matrix metalloproteases-2 (TIMP2) important for controlling cell motility in breast cancer

Summary

Introduction

The receptor tyrosine kinase HER2 (ErbB2) is overexpressed or amplified in 20–30% of breast cancer patients, correlating with cancer aggressiveness and reduced patient survival[1]. We employed generation sequencing to study the miRNA expression profiles induced by these HER2 variants and we interrogated whether p95HER2-induced cellular traits associated with breast cancer aggressiveness involve p95HER2 mediated regulation of miR-221/222 and miR-503 and in turn MYB family transcription factors. P95HER2-upregulated miRNAs include miR-221-/222, resulting in essential ablation of ESR1 expression in p95HER2-expressing cells, and miR-503, which together with miR-221/222 elicit downregulation of MYB and MYBL1. This in turn leads to upregulation of of the putative MYB family target gene, tissue inhibitor of matrix metalloproteases-2 (TIMP2). Our findings identify a novel role for p95HER2-specific miRNA changes in distinguishing p95HER2 positive cancers from those overexpressing only HER2, and uncover a signaling axis from miR-221/222 and -503 over MYB proteins to TIMP2 important for controlling cell motility in breast cancer

Objectives

Methods

Results

Conclusion