Profilin-1 deficiency leads to SMAD3 upregulation and impaired 3D outgrowth of breast cancer cells

Souvik Chakraborty,Andreas Vogt,William Schiemann,Partha Roy,Michael Oddo,Zhijie Ding,Marion Joy,Laura Vollmer,David Gau,Donna Beer Stolz,Chang Jiang,Sujoy Ghosh

doi:10.1038/s41416-018-0284-6

Abstract

BackgroundAdhesion-mediated activation of FAK/ERK signalling pathway, enabled by the formation of filopodial protrusions (FLP), has been shown to be an important event for triggering of dormancy-to-proliferation switch and metastatic outgrowth of breast cancer cells (BCC). We studied the role of actin-binding protein profilin1 (Pfn1) in these processes.MethodsQuantitative immunohistochemistry (IHC) of BC tissue microarray (TMA) and survival analyses of curated transcriptome datasets of BC patients were performed to examine Pfn1’s association with certain clinicopathological features. FLP formation and single cell outgrowth of BCC were assessed using a 3D matrigel culture that accurately predicts dormant vs metastatic outgrowth phenotypes of BCC in certain microenvironment. Gene expression studies were performed to identify potential biological pathways that are perturbed under Pfn1-depleted condition.ResultsLower Pfn1 expression is correlated with lower nuclear grade of breast tumours and longer relapse-free survival of BC patients. Pfn1 depletion leads to defects in FLP and outgrowth of BCC but without impairing either FAK or ERK activation. Guided by transcriptome analyses, we further showed that Pfn1 depletion is associated with prominent SMAD3 upregulation. Although knockdown and overexpression experiments revealed that SMAD3 has an inhibitory effect on the outgrowth of breast cancer cells, SMAD3 knockdown alone was not sufficient to enhance the outgrowth potential of Pfn1-depleted BCC suggesting that other proliferation-regulatory pathways in conjunction with SMAD3 upregulation may underlie the outgrowth-deficient phenotype of BCC cells upon depletion of Pfn1.ConclusionOverall, these data suggest that Pfn1 may be a novel biomarker for BC recurrence and a possible target to reduce metastatic outgrowth of BCC.

Highlights

Tumour metastasis is a multistep process that requires cancer cells to escape from the primary tumour, survive in the circulation, invade the target organs and reinitiate secondary tumour outgrowth at the metastatic sites
Using a three-dimensional (3D) basement membrane matrix (BME or matrigel) culture ( known as MoT culture: it closely approximates the compliant mechanical microenvironment of lung and mammary gland) that accurately predicts the metastatic outgrowth competency of breast cancer cells (BCC) in the lungs,[6,7,15] we show that Pfn[1] deficiency causes defects in filopodial protrusions (FLP) abundance and reduces single cell outgrowth ability of triple-negative BC (TNBC) cells
Nuclear grade (NG) of BC is a good predictor for growth aggressiveness of tumour cells and high nuclear grade (NG) tumours have a higher propensity to relapse

Summary

Introduction

Tumour metastasis is a multistep process that requires cancer cells to escape from the primary tumour, survive in the circulation, invade the target organs and reinitiate secondary tumour outgrowth at the metastatic sites. Oncogenic transformation causes alterations of the actin cytoskeleton through deregulating expression and/or activity of a wide range of actin-regulatory proteins. These changes are often correlated with acquisition of a motile phenotype of cancer cells. Preventing actin stress-fibre assembly or blocking integrin signalling dramatically inhibits FAK-Src-ERK signalling and subsequent metastatic outgrowth of disseminated BCC cells.[1,2,3,4,5] Adhesion-mediated triggering of proliferation switch of BCC is enabled by the formation of F-actin-rich filopodial-like protrusions (FLPs). Adhesion-mediated activation of FAK/ERK signalling pathway, enabled by the formation of filopodial protrusions (FLP), has been shown to be an important event for triggering of dormancy-to-proliferation switch and metastatic outgrowth of breast cancer cells (BCC). CONCLUSION: Overall, these data suggest that Pfn[1] may be a novel biomarker for BC recurrence and a possible target to reduce metastatic outgrowth of BCC

Methods

Results

Conclusion