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Systematic Analysis of Posttranscriptional Gene Expression Adam R Advanced Review Systematic analysis of posttranscriptional gene expression Adam R. Morris,1 Neelanjan Mukherjee1 and Jack D. Keene1∗ Recent systems studies of gene expression have begun to dissect the layers of regulation that underlie the eukaryotic transcriptome, the combined consequence of transcriptional and posttranscriptional events. Among the regulatory layers of the transcriptome are those of the ribonome, a highly dynamic environment of ribonucleoproteins in which RNA-binding proteins (RBPs), noncoding regulatory RNAs (ncRNAs) and messenger RNAs (mRNAs) interact. While multiple mRNAs are coordinated together in groups within the ribonome of a eukaryotic cell, eachindividualtypeofmRNAconsistsofmultiplecopies,eachofwhichhas an opportunity to be a member of more than one modular group termed a posttranscriptional RNA operon or regulon (PTRO). The mRNAs associated with each PTRO encode functionally related proteins and are coordinated at the levels of RNA stability and translation by the actions of the specific RBPs and noncoding regulatory RNAs. This article examines the methods that led to the elucidation of PTROs and the coordinating mechanisms that appear to regulate the RNA components of PTROs. Moreover, the article considers the characteristics of the dynamic systems that drive PTROs and how mRNA components are bound collectively in physical ‘states’ to respond to cellular perturbations and diseases. In conclusion, these studies have challenged the extent to which cellular mRNA abundance can inform investigators of the functional status of a biological system. We argue that understanding the ribonome has greater potential for illuminating the underlying coordination principles of growth, differentiation, and disease . 2009 John Wiley & Sons, Inc. WIREs Syst Biol Med ukaryotic gene expression is a complex process technology was crucial for the study of global gene Ethat integrates myriad signals to coordinate expression and has led to advances in understanding production of thousands of gene products in and categorizing human disease.5 As discussed effectively precise temporal and spatial patterns.1–4 below, while traditional microarray approaches Its inherent complexity has been addressed in recent have limitations for understanding some aspects of years using a systems approach, thus much of gene expression such as transcription rates, novel systems biology is devoted to the study of gene applications have emerged, which provide insight into expression. It is generally believed that improper underlying mechanisms of gene coordination at the regulation of gene expression can lead to many posttranscriptional level.4,6–8 human defects and disorders. Thus, understanding Many studies of global gene expression focus the underlying mechanisms of gene expression has solely on the transcriptome, and the only factor become an active subfield of genetic diseases and assessed is mRNA abundance, which is but a single medicine. For example, the advent of microarray aspect of gene expression. Several studies have suggested that measuring global mRNA levels to ∗Correspondence to: [email protected] assess the transcriptome using microarrays can be 1University Program in Genetics and Genomics, Department of misleading, as gene expression has multiple layers Molecular Genetics and Microbiology, Duke University Medical that manifest themselves in both the nucleus and the Center, Durham, NC, USA cytoplasm after transcription has ended.2,9–12 Aprime DOI: 10.1002/wsbm.054 example of the importance of the posttranscriptional 2009 John Wiley & Sons, Inc. Advanced Review www.wiley.com/wires/sysbio environment was recently demonstrated in a study control functions to achieve a higher level of har- from the laboratory of David Baltimore, which monized outcome.25 Historically, the study of PTR showed that mRNA stability strongly influences gene has focused on the one-on-one small-scale control expression induction kinetics during the inflammatory functions, which may result in profound outcomes, response, in some cases overriding the effects of but does not address overall RNA coordination. For transcription.13 Regulation of transcription is also example, an RBP or microRNA may affect an mRNA a relatively slow process for responding to cellular sequence element within a Luciferase reporter, demon- perturbations; in the sea urchin embryo, the average strating control of the expression of that RNA. In gene is translated seven times more rapidly than it addition, those same trans-acting factors may also be is transcribed (56 min for transcription versus 8 min found to alter the phenotype of a cell or organism, and for translation).14 In addition, some human genes could be presumed to do so by affecting the same single can take as long as 16 h to be transcribed.15 In mRNA target from which the sequence present in the order to rapidly produce a protein, it is advantageous reporter system was derived in vivo. However, the phe- to increase the rate of translation immediately, notypic change is just as likely to result from the com- without waiting for transcription and subsequent bined effects of that trans-acting RBP or microRNA mRNA export and cytoplasmic regulation.11,16 Even on coordinating multiple mRNA targets. In addressing in the case of genes that are shorter and thus this issue, advances in molecular biological techniques transcribed more quickly, translational control of and detection methods have allowed study of con- existing mRNA provides a rapid means to generate trol on a wider basis, often global, thus leading to necessary proteins in response to cellular signals. a greater understanding of RNA coordination. Gao Moreover, repression of transcription can be a et al. (1994) found that the ELAV/HuB RBP can tar- relatively slow means to cease protein production get multiple mRNAs in vitro using total brain mRNA if not coupled to rapid mRNA decay and/or a and suggested that this could represent a coordinat- 26 decrease in translation. In addition, many studies ing function for posttranscriptional gene regulation. have shown that transcription may be more stochastic Subsequently, Tenenbaum et al. (2000) demonstrated than was previously believed, and therefore the that the HuR and HuB RBPs target multiple mRNAs in newly synthesized mRNA populations created by vivo during neuronal differentiation in mouse embry- 7 transcription may be altered correspondingly in the onic carcinoma P19 cells. Similar multiple targeting posttranscriptional environment.17–21 The underlying interactions by microRNAs were predicted using com- mechanisms of posttranscriptional regulation (PTR) putational algorithms, and it is generally assumed are determined by many factors that may bind to today that microRNAs, like RBPs, can target and 27–29 and regulate an mRNA after transcription and up affect multiple mRNAs in living cells. While RBPs to and during translation, including RNA-binding have been shown in many studies to target functionally proteins (RBPs) and small noncoding RNAs, such related mRNAs, such conclusions have not emerged as microRNAs. Recent studies have shown that for microRNAs. This is consistent with the very broad understanding PTR on a global level provides insights target predictions of microRNAs; however, microR- into the coordination of gene expression and its NAs have been reported to have profound effects 30 implications for disease.16,22,23 on phenotypes. Interestingly, the actions of spe- The life of every copy of an mRNA involves cific microRNAs have been shown to fine-tune the multiple points for regulation (Figure 1), including production of multiple proteins, possibly acting as a splicing, polyadenylation, transport from the nucleus, multi-targeted mRNA rheostat, but functional coher- ence has not been demonstrated among the affected localization within the cytoplasm, translation, and 31,32 decay (for a review of the life of an mRNA we proteins. Indeed, coordination of PTR has best recommend16,22–24). In most of the cases where PTR been demonstrated by identifying the genome-wide mRNAs associated with particular RBPs in ribonucle- has been shown to be important, the regulation 22 involved one or multiple RNA-binding proteins. oprotein (RNP) particles, and therefore the role of Thus, RNA-binding proteins have key roles in RBPs in coordinating gene expression is the focus of posttranscriptional gene expression, coordinating this review. many aspects of the life of an mRNA.22,24 Regulation of gene expression at the post- RIBONOMICS-GLOBAL ANALYSIS transcriptional level involves both control and OF mRNPS coordination. While control describes an individual interaction that results in a specific outcome, coor- The global analysis of mRNA and protein compo- dination describes a process of integrating multiple nents of RNPs has been termed ribonomics because 2009 John Wiley & Sons, Inc. WIREs Systems Biology and Medicine Posttranscriptional gene expression Export FIGURE 1| Coordination of posttranscriptional regulation. New Splicing/poly- degradationStorage/ transcripts (squiggled lines) emerge from adenylation chromosomal DNA and undergo multiple interconnected steps of regulation from Localization/ splicing through translation. RBPs translation coordinately regulate functionally related sub-populations of mRNAs existing in the same state as depicted within different colored shapes, each representing a unique combination of trans-acting factors (e.g., RBPs and microRNAs). The dotted lines depict the ‘regulators of regulators’ concept
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