Nuclear cathepsin D enhances TRPS1 transcriptional repressor function to regulate cell cycle progression and transformation in human breast cancer cells.

Anne-Sophie Bach,Sophie Pattingre,Christian Gespach,Thomas Reinheckel,Béatrice Orsetti,Frank J Kaiser,Guillaume Meurice,Emmanuelle Liaudet-Coopman,Charles Theillet,Laetitia K Linares,Salwa Sebti,Danielle Derocq,Céline Gongora,Pascal Roger,Eva Ibing,Philippe Montcourrier,Valérie Laurent-Matha

doi:10.18632/oncotarget.4394

Abstract

The lysosomal protease cathepsin D (Cath-D) is overproduced in breast cancer cells (BCC) and supports tumor growth and metastasis formation. Here, we describe the mechanism whereby Cath-D is accumulated in the nucleus of ERα-positive (ER+) BCC. We identified TRPS1 (tricho-rhino-phalangeal-syndrome 1), a repressor of GATA-mediated transcription, and BAT3 (Scythe/BAG6), a nucleo-cytoplasmic shuttling chaperone protein, as new Cath-D-interacting nuclear proteins. Cath-D binds to BAT3 in ER+ BCC and they partially co-localize at the surface of lysosomes and in the nucleus. BAT3 silencing inhibits Cath-D accumulation in the nucleus, indicating that Cath-D nuclear targeting is controlled by BAT3. Fully mature Cath-D also binds to full-length TRPS1 and they co-localize in the nucleus of ER+ BCC where they are associated with chromatin. Using the LexA-VP16 fusion co-activator reporter assay, we then show that Cath-D acts as a transcriptional repressor, independently of its catalytic activity. Moreover, microarray analysis of BCC in which Cath-D and/or TRPS1 expression were silenced indicated that Cath-D enhances TRPS1-mediated repression of several TRPS1-regulated genes implicated in carcinogenesis, including PTHrP, a canonical TRPS1 gene target. In addition, co-silencing of TRPS1 and Cath-D in BCC affects the transcription of cell cycle, proliferation and transformation genes, and impairs cell cycle progression and soft agar colony formation. These findings indicate that Cath-D acts as a nuclear transcriptional cofactor of TRPS1 to regulate ER+ BCC proliferation and transformation in a non-proteolytic manner.

Highlights

Cathepsins were originally identified as lysosomal proteases, but recent work highlighted their atypical roles in the extracellular space, cytoplasm and nucleus [1]
As TRPS1 is a nuclear protein [15], we examined the cellular localization of TRPS1 and Cathepsin D (Cath-D) in endoplasmic reticulum (ER)+ breast cancer cells (BCC) lines (T47D, MCF7 and BT474), immortalized human breast epithelial cells (HMT3522-S1) and human breast fibroblasts (HMF) (Fig. 3A)
We found that Cath-D interacts with the GATA family member TRPS1 in the nucleus of ER+ BCC and enhances TRPS1 transcription repressor activity, independently of its proteolytic function

Summary

Introduction

Cathepsins were originally identified as lysosomal proteases, but recent work highlighted their atypical roles in the extracellular space, cytoplasm and nucleus [1]. Cathepsin D (Cath-D) is one of the most abundant lysosomal endoproteinases implicated in protein catabolism. Cath-D is an independent marker of poor prognosis for breast cancer associated with metastasis [2, 3]. Cath-D is overproduced by breast cancer cells (BCC) and the pro-enzyme is abundantly secreted in the tumor microenvironment [4]. Cath-D stimulates BCC proliferation, fibroblast outgrowth, angiogenesis, breast tumor growth and metastasis formation [5,6,7,8,9,10,11,12]. Secreted Cath-D enhances proteolysis in the breast tumor microenvironment by degrading the cysteine cathepsin inhibitor cystatin C [13] and promotes mammary fibroblast outgrowth by binding to LDL receptor-related protein-1 (LRP1) [14]

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Results

Conclusion