Tala Bakheet

p38 Mitogen-Activated Protein Kinase-Dependent and -Independent Signaling of mRNA Stability of AU-Rich Element-Containing Transcripts

ABSTRACT Adenylate/uridylate-rich element (ARE)-mediated mRNA turnover is an important regulatory component of gene expression for innate and specific immunity, in the hematopoietic system, in cellular growth regulation, and for many other cellular processes. This diversity is reflected in the distribution of AREs in the human genome, which we have established as a database of more than 900 ARE-containing genes that may utilize AREs as a means of controlling cellular mRNA levels. The p38 mitogen-activated protein kinase (MAP kinase) pathway has been implicated in regulating the stability of nine ARE-containing transcripts. Here we explored the entire spectrum of ARE-containing genes for p38-dependent regulation of ARE-mediated mRNA turnover with a custom cDNA array containing probes for 950 ARE mRNAs. The human monocytic cell line THP-1 treated with lipopolysaccharide (LPS) was used as a reproducible cellular model system that allowed us to precisely control the conditions of mRNA induction and decay in the absence and presence of the p38 inhibitor SB203580. This approach allowed us to establish an LPS-induced ARE mRNA expression profile in human monocytes and determine the half-lives of 470 AU-rich mRNAs. Most importantly, we identified 42 AU-rich genes, previously unrecognized, that show p38-dependent mRNA stabilization. In addition to a number of cytokines, several interesting novel AU-rich transcripts likely to play a role in macrophage activation by LPS exhibited p38-dependent transcript stabilization, including macrophage-specific colony-stimulating factor 1, carbonic anhydrase 2, Bcl2, Bcl2-like 2, and nuclear factor erythroid 2-like 2. Finally, the identification of the p38-dependent upstream activator MAP kinase kinase 6 as a member of this group identifies a positive feedback loop regulating macrophage signaling via p38 MAP kinase-dependent transcript stabilization.

ARED 3.0: the large and diverse AU-rich transcriptome

A comprehensive search that utilized a large set of mRNA data from human genome databases and additionally, expressed sequence tag (EST) database characterized this latest update of AU-rich elements (AREs) containing mRNA database (ARED). A large number of ARE-mRNA, as much as 4000, were recovered and include many of ARE alternative forms. This number represents as much as 5–8% of the human genes depending on the entire number of genes. The new ARED does not contain only larger and diverse number of ARE-mRNAs but additional functionality and enhanced search capabilities are given in the database website . These include class and cluster of AREs, source mRNAs, EST evidence, buildup information, retrieval of lists of genes, and integration with current and new NCBI data, such as Entrez ID and Unigene. Gene Ontology analysis shows there are significant differences in functional diversity of ARED when compared with the overall genome. Many of ARE-genes mediate regulatory processes, reactions to outside stimuli, RNA metabolism, and developmental processes particularly those of early and transient responses. The wide interest in mRNA turnover and importance of AREs in health and disease signify the compilation of ARE-genes.

Systematic Analysis of AU-Rich Element Expression in Cancer Reveals Common Functional Clusters Regulated by Key RNA-Binding Proteins

Defects in AU-rich elements (ARE)-mediated posttranscriptional control can lead to several abnormal processes that underlie carcinogenesis. Here, we performed a systematic analysis of ARE-mRNA expression across multiple cancer types. First, the ARE database (ARED) was intersected with The Cancer Genome Atlas databases and others. A large set of ARE-mRNAs was over-represented in cancer and, unlike non-ARE-mRNAs, correlated with the reversed balance in the expression of the RNA-binding proteins tristetraprolin (TTP, ZFP36) and HuR (ELAVL1). Serial statistical and functional enrichment clustering identified a cluster of 11 overexpressed ARE-mRNAs (CDC6, KIF11, PRC1, NEK2, NCAPG, CENPA, NUF2, KIF18A, CENPE, PBK, TOP2A) that negatively correlated with TTP/HuR mRNA ratios and was involved in the mitotic cell cycle. This cluster was upregulated in a number of solid cancers. Experimentally, we demonstrated that the ARE-mRNA cluster is upregulated in a number of tumor breast cell lines when compared with noninvasive and normal-like breast cancer cells. RNA-IP demonstrated the association of the ARE-mRNAs with TTP and HuR. Experimental modulation of TTP or HuR expression led to changes in the mitosis ARE-mRNAs. Posttranscriptional reporter assays confirmed the functionality of AREs. Moreover, TTP augmented mitotic cell-cycle arrest as demonstrated by flow cytometry and histone H3 phosphorylation. We found that poor breast cancer patient survival was significantly associated with low TTP/HuR mRNA ratios and correlated with high levels of the mitotic ARE-mRNA signature. These results significantly broaden the role of AREs and their binding proteins in cancer, and demonstrate that TTP induces an antimitotic pathway that is diminished in cancer. Cancer Res; 76(14); 4068-80. ©2016 AACR.

ARED-Plus: an updated and expanded database of AU-rich element-containing mRNAs and pre-mRNAs

Abstract Here we present an updated version of the AU-Rich Element Database (ARED-Plus) that is freely available at http://brp.kfshrc.edu.sa/ared. AREs are conserved sequence elements that were first discovered in the 3′UTR of mammalian transcripts. Over the past years, we compiled a series of ARE databases that revealed the extent and wide distribution of ARE-containing genes. For this update, we adopted an optimized search algorithm with improved specificity and sensitivity in ARE selection. The designation of the different ARE clusters was simplified by directly correlating the number of the ARE cluster to the number of overlapping AUUUA pentamers. Additionally, the new database was expanded to include genes with intronic AREs (pre-mRNAs) and their characteristics since recent observations reported their abundance and biological significance. Several enhancements were incorporated such as customized column view, additional search options and live search functionalities. The new version includes links to AREsite and AREScore, two related ARE assessment algorithms for further evaluation of the ARE characteristics. ARED-Plus now contains an updated repertoire of AREs in the human transcriptome that may be useful in several research fields.

Abstract B2-13: AU-rich expression in invasive ductal breast cancer: A systems biology approach in post-transcriptional regulation

AU-Rich element (ARE)-dependent post-transcriptional regulation mechanisms are increasingly emerging as key regulators that are influencing our perception of aberrant gene expression in cancer. AREs are instability determinant sequences located in the 39 UTR of mRNAs and are fundamental to the maintenance of transient expression of gene products that regulate various cellular functions such as cell proliferation, apoptosis, inflammation and immunity. Here we used a systems biology approach to examine the patterns of ARE-containing gene expression that is perturbed in invasive ductal breast cancer (IDBC). Based on IDBC mRNA expression profiles obtained from The Cancer Genome Atlas (TCGA) database, we intersected 2396 over-expressed genes with the AU-rich element database (ARED) and found that 414 comprise IDBC upregulated ARE-containing genes. Functional classification using the gene ontology tool, DAVID, showed the highest enrichment of ARE-genes in IDBC in chromosome segregation (5-fold, p Citation Format: Norah A. Alsouhibani, Edward Hitti, Tala Bakheet, Khalid SA Khabar. AU-rich expression in invasive ductal breast cancer: A systems biology approach in post-transcriptional regulation. [abstract]. In: Proceedings of the AACR Special Conference on Computational and Systems Biology of Cancer; Feb 8-11 2015; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(22 Suppl 2):Abstract nr B2-13.