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1. Introduction
Regulation of gene expression is an essential process through which mammalian cells counter the changes in their microenvironment. These changes drive cells to respond to different stimuli that trigger cellular re-programming towards proliferation, differentiation, development, apoptosis, senescence, carcinogenesis, etc. Once mRNAs are transcribed they are subjected to posttranscriptional events that regulate mRNA metabolism including stability and translation. These two processes normally dictate the protein levels encoded by mRNA. RNA-binding proteins (RPBs) that bind mature mRNA sequences normally have an important regulatory effect on the mRNA.
RBPs are also named mRNA turnover and translation regulator RBPs (TTR-RBPs) since they are capable of regulating both mRNA stability and translation. This family includes numerous members such as Hu proteins [HuA (HuR), HuB, HuC, and HuD] known to bind AU-rich sequences in the 3’ untranslated region (3’UTR) to enhance mRNA translation or to increase its stability [1, 2]. Other RBPs such as T-cell intracellular antigen 1 (TIA-1), TIA-1-related (TIAR), tristetraprolin (TTP), fragile X mental retardation protein (FMRP), polypyrimidine tract-binding protein (PTB), CUG triple repeats RNA-binding protein (CUGBP), nucleolin, and heterogeneous nuclear ribonucleoproteins (hnRNP) A1, A2, and C1/C2 have been shown to influence mRNA stability or translation through interaction with the 3’UTR, CR or the 5’UTR [3-9].
Hu proteins are among the RBPs that are well characterized. While HuR is ubiquitously expressed, HuB, HuC and HuD are primarily neuronal [10]. HuR, also known as embryonic lethal, abnormal vision, Drosophila-like 1 (ELAV L1), binds mRNAs bearing AU- and U-rich sequences, which are considered binding signatures, or RNA-recognition motifs (RRMs) found in numerous mRNAs [11]. HuR binds target mRNA to enhance its stability and/or translation. These targets are involved in several processes such as cell growth, survival, proliferation, stress response, senescence and carcinogenesis [2, 12-17].
The RBP AU-binding factor 1 (AUF1), also known as heterogeneous nuclear ribonucleoprotein D (hnRNPD), is known to bind AU-rich sequences mostly in the 3’UTR of target mRNA. AUF1 belongs to a large family that includes several hnRNPs such as hnRNNP A, B, C, D, E, F, H, I, K, L, M, Q, R and U. AUF1 promotes the degradation of several target transcripts. However, it was also found to enhance the stability and translation of some mRNAs [18-21]. AUF1 is alternatively spliced and all known four isoforms (p37, p40, p42, p45) bind mRNAs but with different binding affinities [22]. AUF1 is thought to recruit mRNAs to the exosome and proteasome for degradation [23, 24]. Target mRNAs are implicated in several processes such as cell cycle, stress response, apoptosis, and carcinogenesis [21].
TIA-1 and TIAR RBPs bind AU/U-rich sequences in the 3’UTR of target transcripts and suppress mRNA translation [25]. However, they can also modulate translation through 5' terminal oligopyrimidine tracts (5'TOP) in response to changes in the cellular environment [26]. Under stress conditions, these proteins are thought to halt mRNA translation in RNA-protein aggregations known as stress granules [27].
The RBP nuclear factor 90 (NF90) interacts with many RNAs bearing AU-rich sequences. NF90 normally suppresses the translation of target mRNAs involved in cell cycle, translation, proliferation and cell division [28]. Although FMRP is expressed in several tissues, it has an essential role in neuronal and intellectual development. FMRP regulates stability and translation of several genes involved in synaptic plasticity [29]. Mutation of FMRP normally results in the inability to inhibit translation and can lead to fragile X syndrome, mental retardation, premature ovarian failure, autism and Parkinson's disease. FMRP is found in RNA granules associated with ribosomal RNA (rRNA) in dendrites [30, 31]. TTP, a zinc finger protein, binds AU-rich sequences in mRNA transcripts to promote their decay. Target mRNAs are involved in cell cycle, inflammation, and carcinogenesis [32]. Nucleolin interacts with mRNAs bearing AU-rich or G-rich sequences to regulate mRNA stability and/or translation. Target transcripts are involved in several processes such as cell cycle, cell morphology, development, growth, proliferation, and carcinogenesis [3]. KH-type splicing regulatory protein (KSRP) RBP binds AU-rich sequences of target transcripts promoting mRNA decay. These targets encode cytokines, chemokines, transcription factors, proto-oncogenes, and cell-cycle regulators [33].
If the RBP functions to promote mRNA degradation such as AUF1 or TTP, then the mRNA half-life is shortened and therefore protein levels will be subsequently low. On the other hand, if the RBP functions to promote mRNA stability such as HuR, then the RBP will subsequently increase protein levels by extending the mRNA half-life. Similarly, if the RBP modulates mRNA translation, protein levels will be influenced accordingly. Figure 1 summarizes these general effects of RBPs as posttranscriptional gene regulators.
This chapter will focus on the effects of RBPs on mRNA stability and translation. It is believed that two or more RBPs may have a functional interplay among themselves and with small RNA molecules known as microRNAs, through binding to the same mRNA. Examples of stabilized and destabilized genes will be indicated, as well as translationally enhanced or suppressed mRNAs and the involvement of encoded proteins in the cellular process.
Figure 1.
Regulation of mRNA stability and translation by RNA-binding proteins (RBPs). They mainly influence the fates of target mRNAs at the post-transcriptional levels. In the cytoplasm, stabilized mRNAs are protected from degradation leading to more protein levels. Destabilized mRNAs are driven to degradation machinery leading to lower protein levels. RBPs can also influence the abundance of mRNAs in the translation machinery (polysomes).
2. Methodology
In this section the methodology of investigating binding of RBPs to target mRNAs as well as the effects on mRNA stability and translation will be explained.
2.1. mRNA-ribonucleoprotein immunoprecipitation (mRNP-IP)
PAS beads from Sigma (P-3391) (or preswollen beads from Sigma) can be used to coat the IgG control antibody or the specific antibody recognizing the RBP. Mix 10 μg antibody, 60 μl volume beads and 200 μl of NT-2 buffer (50 mM Tris, pH 7.4, 150 mM NaCl, 1 mM MgCl2, 0.05% Nonidet P-40). Rotate overnight at 4ºC.
Figure 2.
Schematic illustration of ribonucleoprotein immunoprecipitation (see text).
Harvest and lyse tissue culture cells in ice-cold lysis buffer supplemented with RNAse inhibitors and protease inhibitors for 10 minutes on ice. Lysis buffer is prepared by mixing 100 mM KCl, 5 mM MgCl2, 10 mM Hepes, pH 7.0, 0.5% Nonidet P-40, and 1 mM Dithiothrectol (DTT) added at the time of use
Spin at 14,000 rpm at 4ºC for 30 minutes and discard supernatant. Wash the pellet with 1 ml 70% ethanol and spin at 14.000 rpm at 4ºC for two minutes and then air dry pellet at room temperature for five minutes. Resuspend the pellet in 20-40 μl of RNAse-free water. This RNA can be used as any other RNA for real time PCR analysis or microarrays. If the gene of interest is enriched in RBP-IP which is twofold or higher compared to IgG-IP, then this gene and the RBP do interact. Figure 2 represents a schematic of mRNA-Ribonucleoprotein immunoprecipitation. For experimental examples see references [11, 34, 35].
2.2. Assessing the half-life (t1/2) of target mRNAs
RBPs normally influence either mRNA stability or translation. Downregulation or overexpression of the RBP is helpful to determine whether the RBP affects mRNA stability. This can be achieved by transfection of either siRNA to downregulate, or a construct to overexpress, RBP of interest. Transfected cells are then treated with actinomycin D (2 μg/ml) to inhibit transcription. Cells can be harvested every hour for about six hours followed by isolation of RNA and real time PCR to measure the levels of genes of interest. The ribosomal RNA 18S is normally used for normalization. It is also recommended to measure the levels of a housekeeping gene or a gene that is not targeted by the RBP of interest. If the RBP influences mRNA stability, an increase or a decrease in the t1/2 will be observed and can be calculated using this assay [11, 34, 35].
2.3. Evaluation of mRNA translation by polysome fractions
Prepare sucrose gradient solutions; 10-50% sucrose, 300 mM NaCl, 15mM MgCl2, 15 mM Tris-Cl 7.5, 0.1 mg/ml cycloheximide and 1mg/ml Heparin. Layer the gradient with 2 ml of each solution starting with 10% (top) and ending with 50% (bottom). Be careful not to introduce any air bubbles into the gradient. Leave gradients at 4ºC overnight to allow the step gradient to linearize. Lyse cells as described above (lysis buffer) and place cell lysates slowly at the top of the gradient. Use ultra-centrifugation spin for three hours at 35000 rpm at 4ºC (SW41 rotor). Collect polysome fractions of 1ml each. An example of a polysome profile from HeLa cells is shown in Figure 3. It is important to note that the polysome profile might differ between cell lines and tissues. To analyze the distribution of a particular mRNA in the polysome profile, RNA is isolated from each fraction using Trizol followed by real time PCR and distribution can then be calculated. It is important to note that the sum of the mRNA distribution profile must be 100%. If the RBP affects mRNA translation, one or two observations can be found in the mRNA distribution profile. Significant changes in the amounts of mRNA in heavy translated fraction and a shift in the mRNA distribution profile either towards light or heavy fractions. These examples are illustrated in Figure 3 and reference [35].
3. Hu family
3.1. HuR
As mentioned above, the Hu family includes four RBPs, among them HuR which is ubiquitously expressed. HuR protein recognizes mRNAs through three RNA-recognition motifs [36]. It binds AU-rich sequences present in the 3’UTR or 5’UTR [11, 13]. The interaction of HuR with target mRNA is modulated by HuR phosphorylation through the checkpoint kinase Chk2 [34, 35]. This kinase also regulates HuR levels under heat shock conditions through ubiquitin-proteasome pathway [37]. Protein kinase C (PKC) and mitogen-activated protein kinase (MAPK) p38 also modulate the association of HuR with target mRNA [38-40]. Although HuR is predominantly nuclear, it is capable of shuttling to the cytoplasm through its nucleoyctoplasmic shuttling sequence (HNS) and transport machinery including chromosome region maintenance 1(CRM1), transportins 1 and 2, and importin-1α [41-44]. In addition, under stress conditions such as arsenite or heat shock, HuR aggregates in the cytoplasm with RNAs, known as RNA granules or stress granules, where mRNAs are stored or translationally suppressed [37, 45]. Levels of HuR are regulated by microRNA miR-519 which suppresses HuR mRNA translation [46]. Thus, these and other factors may influence the levels of HuR as a whole or its abundance in the cytoplasm and subsequently affect target mRNAs.
Figure 3.
Schematic representations of translation assay (polysome profiling) to study the influence of RNA-binding proteins (RPBs) on mRNA translation (see text). Left; sucrose gradient, middle; example of a polysome profile using HeLa cells and right; examples of changes in mRNA distribution in polysome profile. Blue represents control, red represents an example of reduced mRNA translation and green represents an example of increased mRNA translation.
3.2. Stabilized HuR targets
Several target mRNAs are stabilized by HuR including cyclins A2, B1, E1, and D1 and p21. Proteins encoded by these mRNAs are involved in cell cycle, proliferation and cell survival. HuR also regulates the stability of genes such as silent mating type information regulation 2 homolog 1 (SIRT1), B-cell lymphoma 2 (BCL-2), epidermal growth factor (EGF), eukaryotic translation initiation factor 4E (eIF4E), and prothymosin α (ProTα) mRNAs. These genes are involved in cell survival and proliferation. HuR promotes carcinogenesis by stabilizing mRNAs that encode for proteins such as Snail, matrix metallopeptidase 9 (MMP-9), urokinase (uPA) and urokinase receptor (uPAR), which enhance tumor invasion. Other genes such as vascular endothelial growth factor
These data suggest that HuR has a protective effect on its target mRNAs. While mRNAs can be recruited to the degradation machinery such as exosome and processing (P) bodies, HuR might be competing for binding to these labile mRNAs preventing or slowing down the degradation process. Since HuR is mostly present in the nucleus and PAR-CLIP identified pre-mRNAs bound to HuR, it is likely that HuR co-transcriptionally binds nuclear RNA substrates. This may also imply that HuR may play roles in splicing or maturation of these RNAs.
3.3. Translationally regulated HuR targets
HuR binds the 3’UTR of several mRNAs to enhance translation. For example, HuR enhances prothymosin α (ProTα), B-cell leukemia (BCL-2), and cyclin A2 mRNA translation. These genes are involved in cell cycle, proliferation, and cell survival [1]. While HuR binds the 3’UTR of wingless-type MMTV integration site family, member 5A (Wnt5a) mRNA to suppress translation, it binds the 3’UTR
3.4. HuD
Among Hu family proteins, HuD is also well studied. This RBP was initially described as neuronal specific [13]; however recent studies have shown HuD to be expressed in other cells such as pancreatic β cells [49, 50]. HuD is essential for neuronal development, identity, and differentiation through stabilization of mRNAs encoding proteins involved in these processes such as growth associated protein 43 (GAP-43), p21, acetylcholinesterase (AchE), and other targets [51-54]. In addition, HuD is involved in Parkinson’s and Alzheimer’s diseases and highly expressed in neuroblastomas [55-58]. Recent studies showed that HuD is regulated by the microRNA miR-375 which suppressed neuritis outgrowth [49]. In pancreatic β cells HuD was also found to bind the 5’UTR of preproinsulin mRNA and negatively regulate mRNA translation [50]. These findings suggest that HuD is expressed in other tissues than neuronal tissues and might have important regulatory functions yet to be investigated.
4. hnRNP family
These RBPs include several members such as hnRNP C, hnRNP D, and hnRNP K. hnRNP D, also known as AUF1, destabilizes mRNAs through binding to the 3’UTR. However some studies have indicated that AUF1 may also stabilize mRNAs and enhance mRNA translation. AUF1 post-transcriptionally regulate the expression of several genes involved in cancer and inflammation [21]. AUF1 is expressed as four isoforms due to alternative splicing of exons 2 and 7. The encoded isoforms are p37AUF1, p40AUF1, p42AUF1, and p45AUF1 according to their molecular masses. AUF1 isoforms contain two RNA recognition motifs (RRMs) that mediate binding to mRNA transcripts [21].
4.1. Influence of AUF1 on target mRNA
Unlike HuR, AUF1 promotes the degradation of the vast majority of its known targets through the recruitment of the mRNA to the exosome and the proteasome [23, 24], for example: cell cycle related genes such as cyclin D1, p21, p27 and p16INK4a [59-62], apoptosis regulators such as B cell leukemia (BCL-2) and growth arrest and DNA-damage-inducible protein alpha (GADD45α) [63, 64]; inflammatory related genes such as granulocyte-macrophage colony-stimulating factor (GM-CSF): and interleukin 6 (IL6) and human inducible nitric oxide synthase (NOS) [65-67] and DNA replication and repair related genes such as thymidylate synthase (TYMS), jun D proto-oncogene (JUND and c-fos (FOS) [68-70]. However, AUF1 can promote the stability and translation of some target transcripts. For example, AUF1 enhances c-MYC mRNA translation and stabilizes interleukin 1β (IL1B) mRNA in LPS stimulated cells [18, 71].
5. Other RBPs
In addition to the abovementioned RBPs, several others are known to post-transcriptionally regulate gene expression in similar fashions. For instance, TTP is known to promote the decay of mRNAs containing ARE sequences including several genes involved in cancer and inflammation such as IL-2, IL-3, IL-6, IL-10, and IL-12 [72-76].
Nucleolin post-transcriptionally modulates the fates of target mRNAs that bear AU-rich and/or G-rich sequences. Nucleolin targets include several genes involved in cellular processes such as proliferation, cell survival, and cell cycle as well as in diseases such as cancer and Alzheimer [3, 77]. For example, nucleolin binds the 3’UTR of BCL-2, GADD45A, gastrin (GAST) and β-globin enhancing mRNA stability. However, nucleolin binds the 3’UTR of APP mRNA, promoting its decay [78-83]. While nucleolin enhances the translation of mRNAs encoding for matrix metallopeptidase 9 (MMP9), AKT1 and cyclin I (CCNI) through binding to the 3’UTR, it suppresses translation of genes encoding for the tumor protein p53 (TP53) and prostaglandin endoperoxide H synthase-1 (PGHS-1) through binding to the 5’UTR [3, 84, 85].
Thus RBPs play an essential role in post-transcriptional gene regulation through binding to different regions of numerous mRNAs encoding for proteins involved in almost all cellular processes impacting cell fates in response to physiological and environmental stimuli.
6. RBPs interplay
Different RBPs are capable of binding to the same RNA inferring diverse effects on the fates of target transcripts. For instance HuR, AUF1, and nucleolin bind BCL-2 mRNA. While nucleolin and HuR promote the stability, AUF1 enhances the degradation of BCL-2 mRNA [63, 86-89]. This implies that HuR and nucleolin have a cooperative effect which is antagonized by AUF1.
Another example is illustrated in the case of GADD45A mRNA. While nucleolin stabilizes GADD45A mRNA, it seems that this effect might be antagonized by AUF1 which promotes its decay and TIAR which suppresses translation [64, 83].
These examples indicate that at least two or more RBPs can bind to the same mRNA molecule in a functional interplay to cooperatively or competitively regulate the fates of target mRNAs, translation and/or stability.
7. Interplay with microRNAs
MicroRNAs (miRNAs) are short RNA molecules, about 22 nt long, that regulate gene expression through RNA-induced silencing complex (RISC) [90, 91]. Targeted mRNAs are silenced either through degradation or translation suppression. RBPs RBPs jointly with miRNAs regulate the fates of mRNAs. While RBPs have diverse effects on target mRNAs, miRNAs only promote mRNA degradation or suppress its translation. In some cases miRNAs and RBPs cooperatively regulate the mRNA to a certain fate. For example, HuR was found to recruit let-7 to suppress c-MYC mRNA translation [92]. HuR competes with miR-494 and miR-548c-3p for the regulation of nucleolin and TOP2A mRNA respectively [93, 94]. These and several other examples indicate that RBPs are capable of competing or cooperating with other RNA binding factors such as miRNA to regulate the expression of target genes.
8. Concluding remarks
RBPs are involved in many cellular processes and pathological conditions such as cancer. Indeed, RBP such as HuR is highly expressed in cancer tissues and is believed to enhance tumorigenesis [2]. AUF1 is also involved in cancer progression through the modulation of neoplastic gene regulatory pathways [21]. Nucleolin is involved in cancer and Alzheimer’s disease, while TTP is involved in cancer and inflammation [32].
Thus RBPs may influence not only gene expression but also diseases and disease-progression. Differential expression or subcellular localization of RBPs in diseases could be useful diagnostic markers and targeted for therapy. Indeed inhibitors of HuR and nucleolin have been reported to influence tumorigenesis in vitro, but their therapeutic usefulness in organisms remains untested [95-100].
In the past decade we have gained useful and specific knowledge about RBPs. We have advanced from <RNA binding> to <sequences specific binding> to <binding signatures/motifs> of RBPs. Nonetheless, it is important to identify the complete set of target RNAs which includes coding and non-coding RNAs in different cellular compartments. For example, the use of techniques such as photoactivatable-ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) can provide a more global spectrum of RNA binding activities of RBPs [47, 101]. In addition, predominantly nuclear or nucleolar RBPs can be also studied by this technology to uncover their binding activities to other RNA species such as microRNAs, long non-coding RNAs, or even nuclear specific RNAs.
This will advance our knowledge in assessing other functions of RBPs in the regulation of gene expression.
References
- 1.
Srikantan S. Gorospe M. Hu R. function in. disease Front. Biosci 2012 189 205 - 2.
Abdelmohsen K. Gorospe M. Posttranscriptional regulation. of cancer. traits by. Hu R. 2010 214 229 - 3.
Abdelmohsen, K., et al., Enhanced translation by Nucleolin via G-rich elements in coding and non-coding regions of target mRNAs. Nucleic Acids Res,2011 8513 8530 - 4.
Kim D. Y. et al. hn R. N. P. Q. mediates a. phase-dependent translation-coupled. m. R. N. A. decay of. mouse Period. 2011 8901 8914 - 5.
Kwon S. Barbarese E. Carson J. H. The cis-acting. R. N. A. trafficking signal. from myelin. basic protein. m. R. N. A. its cognate. trans-acting ligand. hn R. N. P. A. enhance cap-dependent. translation 1999 247 256 - 6.
Hitti, E., et al., Mitogen-activated protein kinase-activated protein kinase 2 regulates tumor necrosis factor mRNA stability and translation mainly by altering tristetraprolin expression, stability, and binding to adenine/uridine-rich element. Mol Cell Biol,2006 2399 2407 - 7.
De Rubeis S. Bagni C. Fragile X. mental retardation. protein control. of neuronal. m. R. N. A. metabolism Insights. into m. R. N. A. stability 2010 43 50 - 8.
Aroca A. Diaz-Quintana A. Diaz-Moreno I. structural A. insight into. the C-terminal. R. N. A. recognition motifs. of T-cell. intracellular antigen. protein F. E. B. 2011 2958 2964 - 9.
Sawicka K. et al. Polypyrimidine-tract-binding protein. a. multifunctional R. N. A-binding protein. 2008 Pt 4):641 647 - 10.
Cuadrado, A., et al., Neuronal HuD gene encoding a mRNA stability regulator is transcriptionally repressed by thyroid hormone. J Neurochem,2003 763 773 - 11.
Lopez de Silanes, I., et al., Identification of a target RNA motif for RNA-binding protein HuR. Proc Natl Acad Sci U S A,2004 2987 2992 - 12.
Abdelmohsen, K., et al., Posttranscriptional gene regulation by RNA-binding proteins during oxidative stress: implications for cellular senescence. Biol Chem,2008 243 255 - 13.
Hinman M. N. Lou H. Diverse molecular. functions of. Hu proteins. 2008 3168 3181 - 14.
Lopez de Silanes. I. Lal A. Gorospe M. Hu R. post-transcriptional paths. to malignancy. R. N. 2005 11 13 - 15.
Kuwano Y. Gorospe M. Protecting the. stress response. guarding the. M. K. P. m. R. N. A. 2008 2640 2642 - 16.
Abdelmohsen, K., et al., Posttranscriptional orchestration of an anti-apoptotic program by HuR. Cell Cycle,2007 1288 1292 - 17.
Yi, J., et al., Reduced nuclear export of HuR mRNA by HuR is linked to the loss of HuR in replicative senescence. Nucleic Acids Res,2010 1547 1558 - 18.
Liao B. Hu Y. Brewer G. Competitive binding. of A. U. F. to T. I. A. R. controls M. Y. C. m. R. N. A. its translation. 2007 511 518 - 19.
Raineri, I., et al., Roles of AUF1 isoforms, HuR and BRF1 in ARE-dependent mRNA turnover studied by RNA interference. Nucleic Acids Res,2004 1279 1288 - 20.
Vazquez-Chantada, M., et al., HuR/methyl-HuR and AUF1 regulate the MAT expressed during liver proliferation, differentiation, and carcinogenesis. Gastroenterology,2010 1943 1953 - 21.
Zucconi, B.E. and G.M. Wilson, Modulation of neoplastic gene regulatory pathways by the RNA-binding factor AUF1. Front Biosci,2012 2307 2325 - 22.
Gratacos F. M. Brewer G. The role. of A. U. F. in regulated. m. R. N. A. decay 2010 457 473 - 23.
Chen, C.Y., et al., AU binding proteins recruit the exosome to degrade ARE-containing mRNAs. Cell,2001 451 464 - 24.
Laroia, G., et al., Control of mRNA decay by heat shock-ubiquitin-proteasome pathway. Science,1999 499 502 - 25.
Kim, H.S., et al., Different modes of interaction by TIAR and HuR with target RNA and DNA. Nucleic Acids Res,2011 1117 1130 - 26.
Damgaard C. K. Lykke-Andersen J. Translational coregulation. of 5’T. O. P. m. R. N. As by T. I. A. T. I. A. R. 2011 2057 2068 - 27.
Gilks, N., et al., Stress granule assembly is mediated by prion-like aggregation of TIA-1. Mol Biol Cell,2004 5383 5398 - 28.
Kuwano, Y., et al., NF90 selectively represses the translation of target mRNAs bearing an AU-rich signature motif. Nucleic Acids Res,2010 225 238 - 29.
Bassell G. J. Warren S. T. Fragile X. syndrome loss. of local. m. R. N. A. regulation alters. synaptic development. function 2008 201 214 - 30.
Antar, L.N., et al., Localization of FMRP-associated mRNA granules and requirement of microtubules for activity-dependent trafficking in hippocampal neurons. Genes Brain Behav,2005 350 359 - 31.
Rooms L. Kooy R. F. Advances in. understanding fragile. X. syndrome related disorders. 2011 601 606 - 32.
Sanduja, S., et al., The role of tristetraprolin in cancer and inflammation. Front Biosci,2012 174 188 - 33.
Briata, P., et al., KSRP, many functions for a single protein. Front Biosci,2011 1787 1796 - 34.
Abdelmohsen, K., et al., Phosphorylation of HuR by Chk2 regulates SIRT1 expression. Mol Cell, 2007 543 557 - 35.
Masuda, K., et al., Global dissociation of HuR-mRNA complexes promotes cell survival after ionizing radiation. EMBO J,2011 1040 1053 - 36.
Burd C. G. Dreyfuss G. Conserved structures. diversity of. functions of. R. N. A-binding proteins. 1994 615 621 - 37.
Abdelmohsen, K., et al., Ubiquitin-mediated proteolysis of HuR by heat shock. EMBO J,2009 1271 1282 - 38.
Doller, A., et al., Posttranslational modification of the AU-rich element binding protein HuR by protein kinase Cdelta elicits angiotensin II-induced stabilization and nuclear export of cyclooxygenase 2 mRNA. Mol Cell Biol,2008 2608 2625 - 39.
Doller A. et al. Protein kinase. C. alpha-dependent phosphorylation. of the. m. R. N. A-stabilizing factor. Hu R. implications for. posttranscriptional regulation. of cyclooxygenase-. 2007 2137 2148 - 40.
Lafarga, V., et al., p38 Mitogen-activated protein kinase- and HuR-dependent stabilization of p21(Cip1) mRNA mediates the G(1)/S checkpoint. Mol Cell Biol,2009 4341 4351 - 41.
Gallouzi, I.E. and J.A. Steitz, Delineation of mRNA export pathways by the use of cell-permeable peptides. Science,2001 1895 1901 - 42.
Fan, X.C. and J.A. Steitz, HNS, a nuclear-cytoplasmic shuttling sequence in HuR. Proc Natl Acad Sci U S A,1998 15293 15298 - 43.
von Roretz, C., A.M. Macri, and I.E. Gallouzi, Transportin 2 regulates apoptosis through the RNA-binding protein HuR. J Biol Chem,2011 25983 25991 - 44.
Rebane A. Aab A. Steitz J. A. Transportins . are . redundant nuclear. import factors. for hn. R. N. P. A. Hu R. R. N. 2004 590 599 - 45.
Kedersha, N., et al., Stress granules and processing bodies are dynamically linked sites of mRNP remodeling. J Cell Biol,2005 871 884 - 46.
Abdelmohsen, K., et al., miR-519 reduces cell proliferation by lowering RNA-binding protein HuR levels. Proc Natl Acad Sci U S A,2008 20297 20302 - 47.
Mukherjee, N., et al., Integrative regulatory mapping indicates that the RNA-binding protein HuR couples pre-mRNA processing and mRNA stability. Mol Cell,2011 327 339 - 48.
Galban, S., et al., RNA-binding proteins HuR and PTB promote the translation of hypoxia-inducible factor 1alpha. Mol Cell Biol,2008 93 107 - 49.
Abdelmohsen, K., et al., miR-375 inhibits differentiation of neurites by lowering HuD levels. Mol Cell Biol,2010 4197 4210 - 50.
Lee, E.K., et al., RNA-Binding Protein HuD Controls Insulin Translation. Mol Cell, 2012 826 835 - 51.
Bolognani F. Contente-Cuomo T. Perrone-Bizzozero N. I. Novel recognition. motifs biological functions. of the. R. N. A-binding protein. Hu D. revealed by. genome-wide identification. of its. targets 2010 117 130 - 52.
Chung, S., et al., The Elav-like proteins bind to a conserved regulatory element in the 3’-untranslated region of GAP-43 mRNA. J Biol Chem,1997 6593 6598 - 53.
Deschenes-Furry J. Perrone-Bizzozero N. Jasmin B. J. The R. N. A-binding protein. Hu D. a. regulator of. neuronal differentiation. maintenance plasticity 2006 822 833 - 54.
Pascale A. Amadio M. Quattrone A. Defining a. neuron neuronal. E. L. A. V. proteins 2008 128 140 - 55.
Amadio, M., et al., nELAV proteins alteration in Alzheimer’s disease brain: a novel putative target for amyloid-beta reverberating on AbetaPP processing. J Alzheimers Dis,2009 409 419 - 56.
Ball, N.S. and P.H. King, Neuron-specific hel-N1 and HuD as novel molecular markers of neuroblastoma: a correlation of HuD messenger RNA levels with favorable prognostic features. Clin Cancer Res,1997 1859 1865 - 57.
DeStefano, A.L., et al., Replication of association between ELAVL4 and Parkinson disease: the GenePD study. Hum Genet,2008 95 99 - 58.
Noureddine, M.A., et al., Association between the neuron-specific RNA-binding protein ELAVL4 and Parkinson disease. Hum Genet,2005 27 33 - 59.
Lal, A., et al., Concurrent versus individual binding of HuR and AUF1 to common labile target mRNAs. EMBO J,2004 3092 3102 - 60.
Trojanowicz, B., et al., The role of AUF1 in thyroid carcinoma progression. Endocr Relat Cancer,2009 857 871 - 61.
Chang, N., et al., HuR uses AUF1 as a cofactor to promote p16INK4 mRNA decay. Mol Cell Biol,2010 3875 3886 - 62.
Wang, W., et al., Increased stability of the p16 mRNA with replicative senescence. EMBO Rep,2005 158 164 - 63.
Ishimaru, D., et al., Mechanism of regulation of BCL-2 mRNA by nucleolin and A+U-rich element-binding factor 1 (AUF1). J Biol Chem,2010 27182 27191 - 64.
Lal, A., et al., Posttranscriptional derepression of GADD45alpha by genotoxic stress. Mol Cell,2006 117 128 - 65.
Sarkar, B., et al., Selective degradation of AU-rich mRNAs promoted by the p37 AUF1 protein isoform. Mol Cell Biol,2003 6685 6693 - 66.
Paschoud, S., et al., Destabilization of interleukin-6 mRNA requires a putative RNA stem-loop structure, an AU-rich element, and the RNA-binding protein AUF1. Mol Cell Biol,2006 8228 8241 - 67.
Pautz, A., et al., Similar regulation of human inducible nitric-oxide synthase expression by different isoforms of the RNA-binding protein AUF1. J Biol Chem,2009 2755 2766 - 68.
Loflin P. Chen C. Y. Shyu A. B. Unraveling a. cytoplasmic role. for hn. R. N. P. D. in the. in vivo. m. R. N. A. destabilization directed. by the. A. U-rich element. 1999 1884 1897 - 69.
Pullmann, R., Jr., et al., Differential stability of thymidylate synthase 3’-untranslated region polymorphic variants regulated by AUF1. J Biol Chem,2006 23456 23463 - 70.
Zou T. et al. Polyamines regulate. the stability. of Jun. D. m. R. N. A. by modulating. the competitive. binding of. its 3’. untranslated region. to Hu. R. A. U. F. 2010 5021 5032 - 71.
Sarkar, S., et al., AUF1 isoform-specific regulation of anti-inflammatory IL10 expression in monocytes. J Interferon Cytokine Res,2008 679 691 - 72.
Ogilvie, R.L., et al., Tristetraprolin down-regulates IL-2 gene expression through AU-rich element-mediated mRNA decay. J Immunol,2005 953 961 - 73.
Stoecklin, G., et al., Somatic mRNA turnover mutants implicate tristetraprolin in the interleukin-3 mRNA degradation pathway. Mol Cell Biol,2000 3753 3763 - 74.
Stoecklin, G., et al., Cellular mutants define a common mRNA degradation pathway targeting cytokine AU-rich elements. RNA,2001 1578 1588 - 75.
Stoecklin, G., et al., Genome-wide analysis identifies interleukin-10 mRNA as target of tristetraprolin. J Biol Chem,2008 11689 11699 - 76.
Jalonen, U., et al., Down-regulation of tristetraprolin expression results in enhanced IL-12 and MIP-2 production and reduced MIP-3alpha synthesis in activated macrophages. Mediators Inflamm,2006 40691 - 77.
Dranovsky, A., et al., Cdc2 phosphorylation of nucleolin demarcates mitotic stages and Alzheimer’s disease pathology. Neurobiol Aging,2001 517 528 - 78.
Chen, C.Y., et al., Nucleolin and YB-1 are required for JNK-mediated interleukin-2 mRNA stabilization during T-cell activation. Genes Dev,2000 1236 1248 - 79.
Otake, Y., et al., Overexpression of nucleolin in chronic lymphocytic leukemia cells induces stabilization of BCL-2 mRNA. Blood,2007 3069 3075 - 80.
Jiang Y. Xu X. S. Russell J. E. nucleolin-binding A. 3’ untranslated. region element. stabilizes beta-globin. m. R. N. A. in vivo. 2006 2419 2429 - 81.
Rajagopalan L. E. et al. hn R. N. P. C. increases amyloid. precursor protein. . A. P. P. production by. stabilizing A. P. P. m. R. N. A. 1998 3418 3423 - 82.
Lee, P.T., et al., Epidermal growth factor increases the interaction between nucleolin and heterogeneous nuclear ribonucleoprotein K/poly(C) binding protein 1 complex to regulate the gastrin mRNA turnover. Mol Biol Cell,2007 5004 5013 - 83.
Zhang, Y., et al., Nucleolin links to arsenic-induced stabilization of GADD45alpha mRNA. Nucleic Acids Res,2006 485 495 - 84.
Takagi, M., et al., Regulation of p53 translation and induction after DNA damage by ribosomal protein L26 and nucleolin. Cell,2005 49 63 - 85.
Bunimov N. et al. Translational regulation. of P. G. H. S. 5’ m. R. N. A. untranslated region. first two. exons conferring. negative regulation. 2007 92 105 - 86.
Sengupta, T.K., et al., Identification of nucleolin as an AU-rich element binding protein involved in BCL-2 mRNA stabilization. J Biol Chem,2004 10855 10863 - 87.
Lossi L. et al. Posttranslational regulation. of B. C. L. levels in. cerebellar granule. cells A. mechanism of. neuronal survival. 2009 855 870 - 88.
Zhang, B., et al., Nucleolin/C23 is a negative regulator of hydrogen peroxide-induced apoptosis in HUVECs. Cell Stress Chaperones,2010 249 257 - 89.
Ishimaru, D., et al., Regulation of BCL-2 expression by HuR in HL60 leukemia cells and A431 carcinoma cells. Mol Cancer Res,2009 1354 1366 - 90.
Shukla G. C. Singh J. Barik S. Micro R. N. As Processing Maturation. Target Recognition. Regulatory Functions. 2011 83 92 - 91.
Bartel, D.P., MicroRNAs: target recognition and regulatory functions. Cell,2009 215 233 - 92.
Kim, H.H., et al., HuR recruits let-7/RISC to repress c-Myc expression. Genes Dev,2009 1743 1748 - 93.
Tominaga, K., et al., Competitive regulation of nucleolin expression by HuR and miR-494. Mol Cell Biol,2011 4219 4231 - 94.
Srikantan, S., et al., Translational control of TOP2A influences doxorubicin efficacy. Mol Cell Biol,2011 3790 3801 - 95.
Meisner, N.C., et al., Identification and mechanistic characterization of low-molecular-weight inhibitors for HuR. Nat Chem Biol,2007 508 515 - 96.
Chae, M.J., et al., Chemical inhibitors destabilize HuR binding to the AU-rich element of TNF-alpha mRNA. Exp Mol Med,2009 824 831 - 97.
Soundararajan, S., et al., The nucleolin targeting aptamer AS1411 destabilizes BCL-2 messenger RNA in human breast cancer cells. Cancer Res,2008 2358 2365 - 98.
Ireson, C.R. and L.R. Kelland, Discovery and development of anticancer aptamers. Mol Cancer Ther,2006 2957 2962 - 99.
Destouches, D., et al., Suppression of tumor growth and angiogenesis by a specific antagonist of the cell-surface expressed nucleolin. PLoS One,2008 e2518 - 100.
Krust, B., et al., Suppression of tumorigenicity of rhabdoid tumor derived G401 cells by the multivalent HB-19 pseudopeptide that targets surface nucleolin. Biochimie,2011 426 433 - 101.
Hafner, M., et al., PAR-CliP--a method to identify transcriptome-wide the binding sites of RNA binding proteins. J Vis Exp,2010