Showing papers on "RNA-induced transcriptional silencing published in 2006"
TL;DR: Recent developments in the plant RNAsilencing field that have identified components of specific silencing pathways and have shed light on the mechanisms and biological roles of RNA silencing in plants are reviewed.
777 citations
TL;DR: This study indicates that, unlike other RNase III type proteins, human Dicer may employ two different dsRBD‐containing proteins that facilitate RISC assembly, and describes an additional dsRNA‐binding protein known as PACT, which is significant in RNA silencing.
Abstract: Small RNA‐mediated gene silencing (RNA silencing) has emerged as a major regulatory pathway in eukaryotes. Identification of the key factors involved in this pathway has been a subject of rigorous investigation in recent years. In humans, small RNAs are generated by Dicer and assembled into the effector complex known as RNA‐induced silencing complex (RISC) by multiple factors including hAgo2, the mRNA‐targeting endonuclease, and TRBP (HIV‐1 TAR RNA‐binding protein), a dsRNA‐binding protein that interacts with both Dicer and hAgo2. Here we describe an additional dsRNA‐binding protein known as PACT, which is significant in RNA silencing. PACT is associated with an ∼500 kDa complex that contains Dicer, hAgo2, and TRBP. The interaction with Dicer involves the third dsRNA‐binding domain (dsRBD) of PACT and the N‐terminal region of Dicer containing the helicase motif. Like TRBP, PACT is not required for the pre‐microRNA (miRNA) cleavage reaction step. However, the depletion of PACT strongly affects the accumulation of mature miRNA in vivo and moderately reduces the efficiency of small interfering RNA‐induced RNA interference. Our study indicates that, unlike other RNase III type proteins, human Dicer may employ two different dsRBD‐containing proteins that facilitate RISC assembly.
735 citations
TL;DR: Mutational analyses indicated that the interdomain interactions couple ATP hydrolysis to RNA unwinding, probably through fine positioning of the duplex relative to the wedge helix, which may enable the targeted modulation of intricate RNA structures.
521 citations
TL;DR: It is found that silencing suppressors p19, p21 and HC‐Pro each inhibit the intermediate step of RNA silencing via binding to siRNAs, although the molecular features required for duplex siRNA binding differ among the three proteins.
Abstract: RNA silencing is an evolutionarily conserved system that functions as an antiviral mechanism in higher plants and insects. To counteract RNA silencing, viruses express silencing suppressors that interfere with both siRNA- and microRNA-guided silencing pathways. We used comparative in vitro and in vivo approaches to analyse the molecular mechanism of suppression by three well-studied silencing suppressors. We found that silencing suppressors p19, p21 and HC-Pro each inhibit the intermediate step of RNA silencing via binding to siRNAs, although the molecular features required for duplex siRNA binding differ among the three proteins. None of the suppressors affected the activity of preassembled RISC complexes. In contrast, each suppressor uniformly inhibited the siRNA-initiated RISC assembly pathway by preventing RNA silencing initiator complex formation.
515 citations
TL;DR: The results establish a connection between RNAi components AGO1 and TRBP2, RNAPII transcription and Polycomb-regulated control of gene expression, implicating Polycomb silencing in the mechanism of mammalian TGS.
Abstract: Argonaute proteins are the core components of effector complexes that facilitate RNA interference (RNAi). Small interfering RNAs (siRNAs) targeted to promoter regions mediate transcriptional gene silencing (TGS) in human cells through heterochromatin formation. RNAi effector complexes have yet to be implicated in the mechanism of mammalian TGS. Here we describe the role of the human Argonaute-1 homolog (AGO1) in directing TGS at the promoters for human immunodeficiency virus-1 coreceptor CCR5 and tumor suppressor RASSF1A. AGO1 associates with RNA polymerase II (RNAPII) and is required for histone H3 Lys9 dimethylation and TGS. AGO1, TAR RNA-binding protein-2 (7TRBP2) and Polycomb protein EZH2 colocalize to the siRNA-targeted RASSF1A promoter, implicating Polycomb silencing in the mechanism of mammalian TGS. These results establish a connection between RNAi components AGO1 and TRBP2, RNAPII transcription and Polycomb-regulated control of gene expression.
455 citations
TL;DR: Findings support a model in which targeting of nascent transcripts by RITS mediates chromatin modifications and suggest that cotranscriptional processing events play a primary role in the silencing mechanism.
407 citations
TL;DR: The DExD/H box family of proteins includes a large number of proteins that play important roles in RNA metabolism, but it is clear that several members of this family are multifunctional and, in addition to acting as RNA helicases in processes such as pre-mRNA processing, play important role in transcriptional regulation.
Abstract: The DExD/H box family of proteins includes a large number of proteins that play important roles in RNA metabolism. Members of this family have been shown to act as RNA helicases or unwindases, using the energy from ATP hydrolysis to unwind RNA structures or dissociate RNA–protein complexes in cellular processes that require modulation of RNA structures. However, it is clear that several members of this family are multifunctional and, in addition to acting as RNA helicases in processes such as pre-mRNA processing, play important roles in transcriptional regulation. In this review I shall concentrate on RNA helicase A (Dhx9), DP103 (Ddx20), p68 (Ddx5) and p72 (Ddx17), proteins for which there is a strong body of evidence showing that they play important roles in transcription, often as coactivators or corepressors through their interaction with key components of the transcriptional machinery, such as CREB-binding protein, p300, RNA polymerase II and histone deacetylases.
384 citations
TL;DR: It is found that synthetic antigene RNAs (agRNAs) complementary to transcription start sites or more upstream regions of gene promoters inhibit gene transcription, which occurs in the nucleus, requires high promoter activity and does not necessarily require histone modification.
Abstract: Duplex RNAs complementary to messenger RNA inhibit translation in mammalian cells by RNA interference (RNAi). Studies have reported that RNAs complementary to promoter DNA also inhibit gene expression. Here we show that the human homologs of Argonaute-1 (AGO1) and Argonaute-2 (AGO2) link the silencing pathways that target mRNA with pathways mediating recognition of DNA. We find that synthetic antigene RNAs (agRNAs) complementary to transcription start sites or more upstream regions of gene promoters inhibit gene transcription. This silencing occurs in the nucleus, requires high promoter activity and does not necessarily require histone modification. AGO1 and AGO2 associate with promoter DNA in cells treated with agRNAs, and inhibiting expression of AGO1 or AGO2 reverses transcriptional and post-transcriptional silencing. Our data indicate key linkages and important mechanistic distinctions between transcriptional and post-transcriptional silencing pathways in mammalian cells.
339 citations
TL;DR: A 2.9 angstrom structure of a complex containing 10 molecules of the N protein and 90 bases of RNA is determined, which serves to protect the RNA in the absence of polynucleotide synthesis.
Abstract: Vesicular stomatitis virus is a negative-stranded RNA virus. Its nucleoprotein (N) binds the viral genomic RNA and is involved in multiple functions including transcription, replication, and assembly. We have determined a 2.9 angstrom structure of a complex containing 10 molecules of the N protein and 90 bases of RNA. The RNA is tightly sequestered in a cavity at the interface between two lobes of the N protein. This serves to protect the RNA in the absence of polynucleotide synthesis. For the RNA to be accessed, some conformational change in the N protein should be necessary.
297 citations
TL;DR: A→I RNA editing seems to have additional functions, including the regulation of retrotransposons and gene silencing, which adds a new urgency to the challenges of fully understanding ADAR functions.
Abstract: The most prevalent type of RNA editing is mediated by ADAR (adenosine deaminase acting on RNA) enzymes, which convert adenosines to inosines (a process known as A-->I RNA editing) in double-stranded (ds)RNA substrates. A-->I RNA editing was long thought to affect only selected transcripts by altering the proteins they encode. However, genome-wide screening has revealed numerous editing sites within inverted Alu repeats in introns and untranslated regions. Also, recent evidence indicates that A-->I RNA editing crosstalks with RNA-interference pathways, which, like A-->I RNA editing, involve dsRNAs. A-->I RNA editing therefore seems to have additional functions, including the regulation of retrotransposons and gene silencing, which adds a new urgency to the challenges of fully understanding ADAR functions.
289 citations
TL;DR: It is shown that a single episode of RNAi in the nematode Caenorhabditis elegans can induce transcriptional silencing effects that are inherited indefinitely in the absence of the original trigger.
Abstract: Small RNA molecules participate in a variety of activities in the cell: in a process known as RNA interference (RNAi), double-stranded RNA triggers the degradation of messenger RNA that has a matching sequence; the small RNA intermediates of this process can also modify gene expression in the nucleus Here we show that a single episode of RNAi in the nematode Caenorhabditis elegans can induce transcriptional silencing effects that are inherited indefinitely in the absence of the original trigger Our findings may prove useful in the ongoing development of RNAi to treat disease
TL;DR: It is shown that CaMV-derived siRNAs accumulate in turnip- and Arabidopsis-infected plants and that the leader is a major, albeit not exclusive, source for those molecules, and direct evidence that at least one of those molecules acts as a bona fide siRNA in infected turnip is provided.
Abstract: RNA silencing is an ancient mechanism of gene regulation with important antiviral roles in plants and insects. Although induction of RNA silencing by RNA viruses has been well documented in plants, the interactions between DNA viruses and the host silencing machinery remain poorly understood. We investigate this question with cauliflower mosaic virus (CaMV), a dsDNA virus that expresses its genome through the polycistronic 35S RNA, which carries an unusually extensive secondary structure known as translational leader. We show that CaMV-derived siRNAs accumulate in turnip- and Arabidopsis-infected plants and that the leader is a major, albeit not exclusive, source for those molecules. Biogenesis of leader-derived siRNA requires the coordinated and hierarchical action of the four Arabidopsis Dicer-like (DCL) proteins. Our study also uncovers a “facilitating” role exerted by the microRNA biosynthetic enzyme DCL1 on accumulation of DCL2-, DCL3-, and DCL4-dependent siRNAs derived from the 35S leader. This feature of DCL1 defines a small RNA biosynthetic pathway that might have relevance for endogenous gene regulation. Several leader-derived siRNAs were found to bear near-perfect sequence complementarity to Arabidopsis transcripts, and, using a sensor transgene, we provide direct evidence that at least one of those molecules acts as a bona fide siRNA in infected turnip. Extensive bioinformatics searches identified >100 transcripts potentially targeted by CaMV-derived siRNAs, several of which are effectively down-regulated during infection. The implications of virus-directed silencing of host gene expression are discussed.
TL;DR: Recent advances in the understanding of different geminivirus silencing suppressors indicate that they can affect all three pathways, suggesting that multiple aspects of silencing impact geminavirus replication.
TL;DR: It is shown that H3K9me2 spreads into silent reporter genes when they are embedded within these transcripts and that spreading requires read-through transcription, as well as slicing by Argonaute, thus, siRNA guides histone modification by basepairing interactions with RNA.
Abstract: Small interfering RNA (siRNA) guides dimethylation of histone H3 lysine-9 (H3K9me2) via the Argonaute and RNA-dependent RNA polymerase complexes, as well as base-pairing with either RNA or DNA. We show that Argonaute requires the conserved aspartate-aspartate-histidine motif for heterochromatic silencing and for ribonuclease H-like cleavage (slicing) of target messages complementary to siRNA. In the fission yeast Schizosaccharomyces pombe, heterochromatic repeats are transcribed by polymerase II. We show that H3K9me2 spreads into silent reporter genes when they are embedded within these transcripts and that spreading requires read-through transcription, as well as slicing by Argonaute. Thus, siRNA guides histone modification by basepairing interactions with RNA.
TL;DR: Enhanced silencing phenotype (esp) mutants in Arabidopsis thaliana are described that reveal how proteins associated with RNA processing and 3′ end formation can influence RNA silencing, and it is proposed that the ESP-containing 3″ end formation complexes prevent transgene and endogenous mRNAs from entering RNA-silencing pathways.
Abstract: Many eukaryotic cells use RNA-directed silencing mechanisms to protect against viruses and transposons and to suppress endogenous gene expression at the posttranscriptional level. RNA silencing also is implicated in epigenetic mechanisms affecting chromosome structure and transcriptional gene silencing. Here, we describe enhanced silencing phenotype (esp) mutants in Arabidopsis thaliana that reveal how proteins associated with RNA processing and 3′ end formation can influence RNA silencing. These proteins were a putative DEAH RNA helicase homologue of the yeast PRP2 RNA splicing cofactor and homologues of mRNA 3′ end formation proteins CstF64, symplekin/PTA1, and CPSF100. The last two proteins physically associated with the flowering time regulator FY in the 3′ end formation complex AtCPSF. The phenotypes of the 3′ end formation esp mutants include impaired termination of the transgene transcripts, early flowering, and enhanced silencing of the FCA-β mRNA. Based on these findings, we propose that the ESP-containing 3′ end formation complexes prevent transgene and endogenous mRNAs from entering RNA-silencing pathways. According to this proposal, in the absence of these ESP proteins, these RNAs have aberrant 3′ termini. The aberrant RNAs would enter the RNA silencing pathways because they are converted into dsRNA by RNA-dependent RNA polymerases.
TL;DR: This review addresses recent findings that appear to blur the definitions of euchromatin and heterochromatin by pointing to the presence of typically heterochROMatic modifications (including H3K9me) in euchromaatin and typically euchROMatic enzymes (including RNA polymerases) in heterochromaatin.
Abstract: Eukaryotic genomes are packaged in two general varieties of chromatin: gene-rich euchromatin and gene-poor heterochromatin. Each type of chromatin has been defined by the presence of distinct chromosomal proteins and posttranslational histone modifications. This review addresses recent findings that appear to blur the definitions of euchromatin and heterochromatin by pointing to the presence of typically heterochromatic modifications (including H3K9me) in euchromatin and typically euchromatic enzymes (including RNA polymerases) in heterochromatin. We discuss the implications of these new findings for the current definition of heterochromatin.
TL;DR: It is shown that activity of siRNA duplexes containing full 2'-OMe substitutions in the sense strand is mediated by the RNA-induced silencing complex (RISC) and that strand-specific loading (or binding) to hAgo2 may be modulated through selective incorporation of these modifications.
Abstract: RNA interference (RNAi) is a process by which short interfering RNAs (siRNAs) direct the degradation of complementary single-strand RNAs. In this study, we investigated the effects of full-strand phosphorothioate (PS) backbone and 2′-O-methyl (2′-OMe) sugar modifications on RNAi-mediated silencing. In contrast to previous reports, we have identified active siRNA duplexes containing full 2′-OMe-modified sense strands that display comparable activity to the unmodified analog of similar sequence. The structure of these modified siRNAs is the predominant determinant of their activity, with sequence and backbone composition being secondary. We further show, by using biotin-tagged siRNAs and affinity-tagged hAgo2/eIF2C2, that activity of siRNA duplexes containing full 2′-OMe substitutions in the sense strand is mediated by the RNA-induced silencing complex (RISC) and that strand-specific loading (or binding) to hAgo2 may be modulated through selective incorporation of these modifications.
TL;DR: The application of tissue-specific or inducible gene silencing, with the use of appropriate promoters, and the ability to silence several genes simultaneously should enhance the authors' ability to create novel traits in plants.
TL;DR: Interestingly, orthologous silencing proteins from different fungal clades are often clustered more closely than paralogous proteins in a fungus, indicating that duplication events occurred before speciation events.
Abstract: Comprehensive phylogenetic analyses of fungal Argonaute, Dicer, and RNA-dependent RNA polymerase-like proteins have been performed to gain insights into the diversification of RNA silencing pathways during the evolution of fungi. A wide range of fungi including ascomycetes, basidiomycetyes, and zygomycetes possesses multiple RNA silencing components in the genome, whereas a portion of ascomycete and basidiomycete fungi apparently lacks the whole or most of the components. The number of paralogous silencing proteins in the genome differs considerably among fungal species, suggesting that RNA silencing pathways have diversified significantly during evolution in parallel with developing the complexity of life cycle or in response to environmental conditions. Interestingly, orthologous silencing proteins from different fungal clades are often clustered more closely than paralogous proteins in a fungus, indicating that duplication events occurred before speciation events. Therefore, the origin of multiple RNA silencing pathways seems to be very ancient, likely having occurred prior to the divergence of the major fungal lineages.
TL;DR: All approaches employed in Chlamydomonas to date to downregulate target gene expression by antisense or inverted repeat constructs are reviewed and discussed critically.
Abstract: The generation of a comprehensive EST library and the sequencing of its genome set the stage for reverse genetics approaches in Chlamydomonas reinhardtii However, these also require tools for the specific downregulation of target gene expression Consequently, a large number of diverse constructs were developed aimed at reducing target gene expression in Chlamydomonas via the stable expression of antisense or inverted repeat-containing RNA Double-stranded RNA (dsRNA) generated by the annealing of antisense and sense RNAs or by hairpin formation of an inverted repeat, feeds into the RNA silencing pathway In this pathway, dsRNA is cleaved into approximately 25-bp small interfering RNAs (siRNAs) by the endonuclease Dicer One of the two complementary strands of a siRNA is then loaded onto an Argonaute-like protein present as core component within larger complexes Guided by this single-stranded RNA, the Argonaute-like protein either detects homologous transcripts and cleaves these endonucleolytically, or initiates transcriptional gene silencing This article summarizes current information derived mainly from the Chlamydomonas genome project on components that are assumed to be involved in RNA silencing mechanisms in Chlamydomonas Furthermore, all approaches employed in Chlamydomonas to date to downregulate target gene expression by antisense or inverted repeat constructs are reviewed and discussed critically
TL;DR: The identification of XRN4 as an integral new component in ethylene signaling adds RNA degradation as another posttranscriptional process that modulates the perception of this plant hormone.
Abstract: EXORIBONUCLEASE4 (XRN4), the Arabidopsis thaliana homolog of yeast XRN1, is involved in the degradation of several unstable mRNAs. Although a role for XRN4 in RNA silencing of certain transgenes has been reported, xrn4 mutant plants were found to lack any apparent visible phenotype. Here, we show that XRN4 is allelic to the unidentified components of the ethylene response pathway ETHYLENE-INSENSITIVE5/ACC-INSENSITIVE1 (EIN5/AIN1) and EIN7. xrn4 mutant seedlings are ethylene-insensitive as a consequence of the upregulation of EIN3 BINDING F-BOX PROTEIN1 (EBF1) and EBF2 mRNA levels, which encode related F-box proteins involved in the turnover of EIN3 protein, a crucial transcriptional regulator of the ethylene response pathway. Epistasis analysis placed XRN4/EIN5/AIN1 downstream of CTR1 and upstream of EBF1/2. XRN4 does not appear to regulate ethylene signaling via an RNA-INDUCED SILENCING COMPLEX–based RNA silencing mechanism but acts by independent means. The identification of XRN4 as an integral new component in ethylene signaling adds RNA degradation as another posttranscriptional process that modulates the perception of this plant hormone.
TL;DR: It is reported that hypovirus CHV1-EP713-encoded p29 can suppress RNA silencing in the natural host, the chestnut blight fungus Cryphonectria parasitica.
Abstract: Virulence-attenuating hypoviruses of the species Cryphonectria hypovirus 1 (CHV1) encode a papain-like protease, p29, that shares similarities with the potyvirus-encoded suppressor of RNA silencing HC-Pro. We now report that hypovirus CHV1-EP713-encoded p29 can suppress RNA silencing in the natural host, the chestnut blight fungus Cryphonectria parasitica. Hairpin RNA-triggered silencing was suppressed in C. parasitica strains expressing p29, and transformation of a transgenic green fluorescent protein (GFP)-silenced strain with p29 resulted in an increased number of transformants with elevated GFP expression levels. The CHV1-EP713 p29 protein was also shown to suppress both virus-induced and agroinfiltration-induced RNA silencing and systemic spread of silencing in GFP-expressing transgenic Nicotiana benthamiana line 16c plants. The demonstration that a mycovirus encodes a suppressor of RNA silencing provides circumstantial evidence that RNA silencing in fungi may serve as an antiviral defense mechanism. The observation that a phylogenetically conserved protein of related plant and fungal viruses functions as a suppressor of RNA silencing in both fungi and plants indicates a level of conservation of the mechanisms underlying RNA silencing in these two groups of organisms.
TL;DR: A highly conserved RNA structure in the 3′ non-translated region of the SARS virus was cleaved preferentially at one of the unpaired uridylate bases, demonstrating that both RNA structure and base-pairing can affect cleavage by Nsp15.
TL;DR: Thermal melting analysis showed that the base pair between rF and adenosine (A) was destabilizing relative to a uridine-adenosine pair, although it was slightly less destabilizing than other mismatches.
Abstract: Recently, chemically synthesized short interfering RNA (siRNA) duplexes have been used with success for gene silencing. Chemical modification is desired for therapeutic applications to improve biostability and pharmacokinetic properties; chemical modification may also provide insight into the mechanism of silencing. siRNA duplexes containing the 2,4-difluorotoluyl ribonucleoside (rF) were synthesized to evaluate the effect of noncanonical nucleoside mimetics on RNA interference. 5′-Modification of the guide strand with rF did not alter silencing relative to unmodified control. Internal uridine to rF substitutions were well-tolerated. Thermal melting analysis showed that the base pair between rF and adenosine (A) was destabilizing relative to a uridine-adenosine pair, although it was slightly less destabilizing than other mismatches. The crystal structure of a duplex containing rF∘A pairs showed local structural variations relative to a canonical RNA helix. As the fluorine atoms cannot act as hydrogen bond...
TL;DR: The 2.3-Å resolution crystal structure of QDE-1, a cRdRP from Neurospora crassa, is reported and it is found that it forms a relatively compact dimeric molecule, each subunit with a catalytic apparatus and protein fold strikingly similar to the catalytic core of the DNA-dependent RNA polymerases responsible for transcription.
Abstract: RNA silencing refers to a group of RNA-induced gene-silencing mechanisms that developed early in the eukaryotic lineage, probably for defence against pathogens and regulation of gene expression. In plants, protozoa, fungi, and nematodes, but apparently not insects and vertebrates, it involves a cell-encoded RNA-dependent RNA polymerase (cRdRP) that produces double-stranded RNA triggers from aberrant single-stranded RNA. We report the 2.3-A resolution crystal structure of QDE-1, a cRdRP from Neurospora crassa, and find that it forms a relatively compact dimeric molecule, each subunit of which comprises several domains with, at its core, a catalytic apparatus and protein fold strikingly similar to the catalytic core of the DNA-dependent RNA polymerases responsible for transcription. This evolutionary link between the two enzyme types suggests that aspects of RNA silencing in some organisms may recapitulate transcription/replication pathways functioning in the ancient RNA-based world.
TL;DR: This work shows that siRNA either selectively localizes in the cytoplasm or translocates into the nucleus, depending on where the silencing target RNA resides, and suggests the existence of a mechanism by which the RNAi machinery orchestrates a target-determined localization of the siRNA and the corresponding RNAi activity.
Abstract: Recent observations of RNA interference (RNAi) in the nuclei of human cells raise key questions about the extent to which nuclear and cytoplasmic RNAi pathways are shared. By directly visualizing the localization of small interfering RNA (siRNA) in live human cells, we show here that siRNA either selectively localizes in the cytoplasm or translocates into the nucleus, depending on where the silencing target RNA resides. Two siRNAs that target the small nuclear 7SK and U6 RNAs localize into the nucleus as duplexes. In contrast, an siRNA targeting the cytoplasmic hepatitis C virus replicon RNA dissociates, and only antisense strand distributes in the cytoplasm of the cells harboring the target RNA, whereas sense strand gets degraded. At the same time, both strands of the latter siRNA are distributed throughout the cytoplasm and nucleus in cells lacking the silencing target RNA. These results suggest the existence of a mechanism by which the RNAi machinery orchestrates a target-determined localization of the siRNA and the corresponding RNAi activity, and also provide evidence for formation of nuclear-programmed active RNA induced silencing complexes directly in the nucleus.
TL;DR: The results implicate 1360 as a target for sequence-specific heterochromatic silencing through an RNAi-dependent mechanism, suggesting that proximity to pericentric heterochromaatin and/or a high local TE density contributes to heterochromeatin formation.
TL;DR: This work has identified a gene (MUT68) in the unicellular green alga Chlamydomonas reinhardtii that is required for the efficient decay of siRNA-targeted transcripts and encodes a noncanonical polyadenylate polymerase that adds untemplated adenines to the 5′ RNA fragments after si RNA-mediated cleavage and appears to stimulate their exosome-dependent degradation.
Abstract: Double-stranded RNA, processed to small interfering RNAs (siRNAs) by Dicer and incorporated into the RNA-induced silencing complex (RISC), triggers gene silencing by a variety of pathways in eukaryotes. RNA interference involving the degradation of homologous transcripts is the best-characterized mechanism. However, the fate of the RNA fragments resulting from siRNA-directed cleavage is poorly understood. We have identified a gene (MUT68) in the unicellular green alga Chlamydomonas reinhardtii that is required for the efficient decay of siRNA-targeted transcripts. MUT68 encodes a noncanonical polyadenylate polymerase that adds untemplated adenines to the 5' RNA fragments after siRNA-mediated cleavage and appears to stimulate their exosome-dependent degradation.
TL;DR: It is demonstrated that the anti-Ago2 monoclonal antibody 4F9 recognized GWBs in a cell cycle dependent manner and was capable of capturing miRNAs associated with Ago2, the effector protein of RNAi.
TL;DR: The latest achievements are explored and future challenges that lie ahead in the RBP world are discussed; a selected group of mRNAs associated with a particular function or biological pathway/process that is regulated by the same RBP.
Abstract: Gene expression is regulated by a complex series of events that take place both at the transcriptional and post-transcriptional levels. The mechanisms and factors involved in transcriptional regulation are relatively well understood, whereas post-transcriptional regulation, in comparison, is still a poorly appreciated process. RNA binding proteins (RBPs) are the key regulators of all post-transcriptional events (RNA splicing, stability, transport and translation). Essentially, in order to improve our knowledge in post-transcriptional regulation, we need to elucidate the mechanisms employed by RBPs to control gene expression. The combination of genomic tools with traditional biochemical approaches generated novel technologies, like ribonomics. The application of these novel technologies not only had a profound impact in the study of RBPs but also created the ground work necessary for the identification of post-transcriptional gene networks; a selected group of mRNAs associated with a particular function or biological pathway/ process that is regulated by the same RBP. In this review article, we explore the latest achievements and discuss future challenges that lie ahead in the RBP world.