Cold-inducible RNA binding protein promotes breast cancer cell malignancy by regulating Cystatin C levels.

Alberto Indacochea,Santiago Guerrero,Matilde E Lleonart,Olga Coll,Ferrán Araujo,Fátima Gebauer,Macarena Ureña

doi:10.1261/rna.076422.120

Abstract

Cold-inducible RNA binding protein (CIRBP) is a stress-responsive protein that promotes cancer development and inflammation. Critical to most CIRBP functions is its capacity to bind and posttranscriptionally modulate mRNA. However, a transcriptome-wide analysis of CIRBP mRNA targets in cancer has not yet been performed. Here, we use an ex vivo breast cancer model to identify CIRBP targets and mechanisms. We find that CIRBP transcript levels correlate with breast cancer subtype and are an indicator of luminal A/B prognosis. Accordingly, overexpression of CIRBP in nontumoral MCF-10A cells promotes cell growth and clonogenicity, while depletion of CIRBP from luminal A MCF-7 cells has opposite effects. We use RNA immunoprecipitation followed by high-throughput sequencing (RIP-seq) to identify a set of 204 high confident CIRBP targets in MCF-7 cells. About 10% of these showed complementary changes after CIRBP manipulation in MCF-10A and MCF-7 cells, and were highly interconnected with known breast cancer genes. To test the potential of CIRBP-mediated regulation of these targets in breast cancer development, we focused on Cystatin C (CST3), one of the most highly interconnected genes, encoding a protein that displays tumor suppressive capacities. CST3 depletion restored the effects of CIRBP depletion in MCF-7 cells, indicating that CIRBP functions, at least in part, by down-regulating CST3 levels. Our data provide a resource of CIRBP targets in breast cancer, and identify CST3 as a novel downstream mediator of CIRBP function.

Highlights

RNA binding proteins orchestrate posttranscriptional control of gene expression and are emerging as important modulators of cancer progression (Wurth and Gebauer 2015; Pereira et al 2017; Moore et al 2018; GarcíaCárdenas et al 2019)
We focus on breast cancer because (i) previous in vitro evidence indicated that Cold-inducible RNA binding protein (CIRBP) could modulate the tumoral properties of MDA-MB-231 breast cancer cells (Chang et al 2016), (ii) immunohistochemistry (IHC) analysis pointed to the presence of CIRBP in the cytoplasm of cells from breast cancer patients (Artero-Castro et al 2009), and (iii) the large number of samples present in databases compared to other tumor types allows for higher statistical relevance
CIRBP is up-regulated in luminal breast cancer and its expression correlates with poor clinical outcome

Summary

Introduction

RNA binding proteins orchestrate posttranscriptional control of gene expression and are emerging as important modulators of cancer progression (Wurth and Gebauer 2015; Pereira et al 2017; Moore et al 2018; GarcíaCárdenas et al 2019). Cold-inducible RNA binding protein (CIRBP, termed CIRP and hnRNP A18) is a stress-responsive protein that partially relocates from the nucleus to the cytoplasm under diverse types of stress such as mild hypothermia, UV-irradiation, endoplasmic reticulum (ER) stress or hypoxia (Lujan et al 2018). CIRBP binds to the coding sequence and 3′-UTRs of target transcripts and promotes or stabilizes mRNA levels. CIRBP regulates circadian gene expression by promoting alternative polyadenylation (APA) of target transcripts (Morf et al 2012; Liu et al 2013). Under UV-irradiation, CIRBP is transiently recruited to sites of DNA damage, where it promotes repair by modulating the association of DNA repair complexes (MRN and RNA (2021) 27:190–201; Published by Cold Spring Harbor Laboratory Press for the RNA Society

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