Calaluce et al. BMC Cancer 2010, 10:126 http://www.biomedcentral.com/1471-2407/10/126 R E S E A R C H A R T I C L E Open Access The RNA binding protein HuR differentially regulates unique subsets of mRNAs in estrogen receptor negative and estrogen receptor positive breast cancer Robert Calaluce1, Matthew M Gubin1, J Wade Davis2,3, Joseph D Magee1, Jing Chen1, Yuki Kuwano6, Myriam Gorospe6, Ulus Atasoy1,4,5* Abstract Background: The discordance between steady-state levels of mRNAs and protein has been attributed to posttranscriptional control mechanisms affecting mRNA stability and translation. Traditional methods of genome wide microarray analysis, profiling steady-state levels of mRNA, may miss important mRNA targets owing to significant posttranscriptional gene regulation by RNA binding proteins (RBPs). Methods: The ribonomic approach, utilizing RNA immunoprecipitation hybridized to microarray (RIP-Chip), provides global identification of putative endogenous mRNA targets of different RBPs. HuR is an RBP that binds to the AU-rich elements (ARE) of labile mRNAs, such as proto-oncogenes, facilitating their translation into protein. HuR has been shown to play a role in cancer progression and elevated levels of cytoplasmic HuR directly correlate with increased invasiveness and poor prognosis for many cancers, including those of the breast. HuR has been described to control genes in several of the acquired capabilities of cancer and has been hypothesized to be a tumor-maintenance gene, allowing for cancers to proliferate once they are established. Results: We used HuR RIP-Chip as a comprehensive and systematic method to survey breast cancer target genes in both MCF-7 (estrogen receptor positive, ER+) and MDA-MB-231 (estrogen receptor negative, ER-) breast cancer cell lines. We identified unique subsets of HuR-associated mRNAs found individually or in both cell types. Two novel HuR targets, CD9 and CALM2 mRNAs, were identified and validated by quantitative RT-PCR and biotin pull- down analysis. Conclusion: This is the first report of a side-by-side genome-wide comparison of HuR-associated targets in wild type ER+ and ER- breast cancer. We found distinct, differentially expressed subsets of cancer related genes in ER+ and ER- breast cancer cell lines, and noted that the differential regulation of two cancer-related genes by HuR was contingent upon the cellular environment. Background Over the past decade array technologies have provided several new means for profiling global changes in gene expression. The power of DNA microarrays is perhaps best illustrated in the way it has been used to differenti- ate treatment responses in patient populations. Indivi- dualized and targeted therapy for several tumors, based * Correspondence: atasoyu@missouri.edu 1Departments of Surgery, University of Missouri, One Hospital Drive, Columbia, Missouri 65212, USA upon underlying differences at the molecular level among gene expression profiles, is beginning to replace the traditional morphological-based treatment paradigm [1-3]. Genome wide microarray analyses, however, are inherently flawed since they globally profile the steady- state levels of mRNA, referred to as the transcriptome. Cellular protein expression levels, however, do not directly correlate with steady-state levels of mRNAs. It is well accepted in the RNA field that there is a poor correlation between steady-state RNA levels and protein. © 2010 Calaluce et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.