The following text describes a protein:
Function:
Double-stranded RNA (dsRNA) endoribonuclease which cleaves endogenous (endo), exogenous (exo), and viral dsRNAs to produce short interfering RNAs (siRNAs). The generated siRNAs then mediate gene silencing, also called RNA interference (RNAi), by controlling the elimination of endogenous transcripts from mobile and repetitive DNA elements of the genome as well as exogenous RNA of viral origin. Also acts in an RNAi-independent manner to activate translation through cytoplasmic polyadenylation. As well as its role in dsRNA processing, essential in several steps of the siRNA biogenesis pathway, including siRNA loading into the Argonaute 2 (AGO2)-containing RNA-induced silencing complex (siRISC), length-dependent dicing and guide strand selection for target silencing by the siRISC. Cleaves dsRNAs into siRNAs that are predominantly around twenty-one nucleotides in length. Displays a preference for binding and processing blunt termini (BLT), likely non-self dsRNAs, over dsRNAs with 2 nucleotides 3' overhanging (3'ovr) termini, which are typically the structure of endogenous dsRNAs. According to many reports, the cleavage reaction mode of the enzyme changes according to the termini of the dsRNA substrate. BLT dsRNAs undergo an ATP-dependent processive reaction whereby multiple siRNAs of heterogeneous sizes are produced before the enzyme dissociates. In contrast, dsRNAs with 3'ovr termini, which are typically the structure of endogenous dsRNAs, undergo ATP-independent, distributive cleavage whereby the enzyme dissociates after each cleavage to produce siRNAs of around 22 nucleotides. However, according to another report, the mode of cleavage reaction is not affected by the terminal structures of the dsRNAs substrates. This report suggests that the enzyme is able to initiate processive cleavage of both BLT and 3'ovr dsRNA substrates, and only rarely initiates distributive cleavage. During dsRNA processing and AGO2-loading, requires association with dsRNA-binding accessory proteins loqs isoform PD (loqs-PD) and r2d2. Functions with r2d2 to form the siRNA-mediated RISC loading complex (siRLC) which is responsible for Ago2-loading of endo- and exo-siRNAs. Interaction with loqs-PD increases initial binding to dsRNA substrates and promotes processing of a subset of endo- and exo-dsRNAs. In the absence of r2d2, may also form an alternative siRLC with loqs-PD to load siRNAs into the siRISC. Function with loqs-PD allows the dicer enzyme to cleave endogenous dsRNA templates independently of their termini, and is required for ATP-dependent processing of a subset of siRNAs but is not required for antiviral RNAi. This suggests that the enzyme's intrinsic termini preferences function in viral defense, while function with loqs-PD allows processing of endogenous dsRNAs with diverse termini. However, according to another report the mode of cleavage reaction is not affected by the presence or absence of loqs-PD. Loaded siRNAs serve as a guide to direct the siRISC to complementary RNAs to degrade them or prevent their translation. The siRLC plays an important role in the ATP-dependent asymmetry sensing of the duplex, and is therefore also responsible for the selection of the strand that ultimately acts as the guide siRNA for the siRISC. Thermodynamically asymmetric siRNAs are preoriented in the siRLC by either the dsRNA-binding r2d2 protein, or the loqs-PD protein in the alternative siRLC, which preferentially bind to the most thermodynamically stable strand prior to loading onto AGO2. Both r2d2 and Dcr-2 also initiate unwinding of the siRNA duplex, at which point the heterodimer is exchanged by AGO2. The strand that was bound by r2d2 is discarded while the one that was bound by Dcr-2 is loaded onto Ago2 and serves as guide to direct the siRISC to complementary RNAs to degrade them or prevent their translation. Independently of its role in RNAi, acts with the cytoplasmic poly(A) polymerase wisp to promote cytoplasmic polyadenylation and translational activation of certain messenger RNAs including r2d2 and toll (Tl) transcripts. Consequently it is involved in the post-transcriptional regulation of the Toll immune signaling pathway and promoting resistance to fungal and viral infections. As an RNA-binding protein, likely functions in cytoplasmic polyadenylation by recruiting the poly(A) RNA polymerase wisp to target mRNAs.
Subcellular Localization:
Nucleus. Cytoplasm. Cytoplasm, Cytoplasmic ribonucleoprotein granule.
Domain:
When in complex with loqs isoform PD (loqs-PD), undergoes significant conformational changes during the full ATP-dependent dicing reaction cycle for processing a 50 bp dsRNA with a 3' two-nucleotide overhang and a 5' monophosphate terminus. At the initial dsRNA binding stage, the helicase and DUF283 domains transition from an extended to a closed conformation, this anchors the bound dsRNA through major and minor groove recognition and forms the ATP- and 5'-phosphate binding pockets required for dicing activity. In the next ATP hydrolysis-driven steps, the dsRNAs is thread through the helicase domain towards the catalytic center. The overall domain configuration is relatively rigid during the translocation process until the dsRNA terminus reaches the Platform-PAZ domains. During the early-translocation stage, in which about 8 bp of the dsRNA duplex is threaded through the helicase domain towards the catalytic center, interactions between the DUF283 and RIIIDb domains prevent non-specific cleavage by blocking the access of dsRNA to the RNase active center. In the mid-translocation stage, in which about 17 bp of the dsRNA duplex is thread towards the catalytic center, the dsRBD domain binds to the dsRNA, in the process bending and pushing the dsRNA towards the PAZ domain. At the end of the translocation stage, around 21 bp of the dsRNA threads through the helicase domain into the PAZ-platform cassette, in the process disrupting the DUF283-RNaseIIIb interaction, allowing the dsRNA substrate to enter the catalytic active center of the RIIID domains for precise cleavage, and thus achieves the fully active dicing conformation. In this structure, a clear breakage of the dsRNA after the dicing near to the catalytic center occurs exactly 21 bp away from the PAZ-domain-binding terminus. During the post-dicing state, the cleaved siRNA is released, and the remaining dsRNA duplex bound by the helicase domain returns to a conformation similar to the early-translocation state, which enables the complex to start the next dicing cycle; The N-terminal helicase domain, containing the Helicase ATP-binding, Helicase C-terminal and Helicase insertion domains, functions in dsRNA stimulated ATPase activity and RNA translocation during the dicing reaction. Essential for processing endogenous and viral dsRNAs. Binds and hydrolyzes ATP enabling the dicer to translocate along the long dsRNA substrates, and produce siRNAs processively from the end. Some reports suggest the domain is essential for the recognition and initial binding of both blunt (BLT) and 3' overhanging (3'ovr) termini dsRNA substrates. However, another reports found that the helicase domain is only required for binding and correct cleavage of BLT dsRNAs and is dispensable for processing of 3'ovr termini dsRNAs. One of the reports suggest that the helicase domain recognizes dsRNA with BLT termini, and then initiates processive cleavage via a threading mechanism in which the BLT dsRNA is locally unwound and threaded through the helicase domain in an ATP-dependent manner; The PAZ and platform domain is important for ensuring length fidelity of siRNAs, substrate discrimination, cleavage and anchoring of 3' overhanging (3'ovr) termini substrates, and the positive regulation of Tl. The platform-PAZ domains are important for mediating the ATP-independent cleavage of dsRNAs with 3' overhanging (3'ovr) dsRNAs into 22mer siRNAs. Substrate dsRNAs with blunt termini, are threaded through the helicase domain, cleaved and then the 2-nt 3' overhang of the dsRNA is recognized and anchored by the 5'-pocket in the platform-PAZ domain. The PAZ domain recognizes and anchors the 5'-monophosphate of long dsRNA substrates, positioning the substrate so that the RNase III domain can cleave the dsRNAs 21 nt away from their 5' end. Although it is important for ensuring length fidelity of 21-nt siRNA production, it is not required for efficient siRNA production. Important for substrate discrimination. Inorganic phosphate appears to occupy the same binding pocket in the PAZ domain as the 5' monophosphorylated end of short dsRNAs, pre-miRNAs or hairpin RNA substrates, and so once bound, the inorganic phosphate likely blocks binding and thus cleavage of the dsRNAs. Physiological concentrations of inorganic phosphate inhibit processing of the inappropriate substrates microRNAs (pre-miRNAs) and short dsRNAs, whereas cleavage of long dsRNAs is not inhibited possibly because they are recognized by a different domain/s i.e. the helicase domain and/or the central dsRNA binding domain. This suggests that binding to inorganic phosphate may block binding to nonphysiological substrates, such as pre-miRNAs function to prevent the enzyme from processing nonphysiological dsRNAs substrates. This domain is also important for the ATP-dependent cleavage reaction. The PAZ domain also interacts with the Tl mRNA 3'UTR, and is therefore important for the positive regulation of Tl at the post-transcriptional level, and thus mediating Toll signaling; The DRBM domain or RNA binding domain (RBD), is important for efficient and high-fidelity production of 21 nucleotide siRNAs, and siRNA loading onto AGO2; Interactions between the DUF283 and RIIIDb domains prevent non-specific cleavage by blocking the access of dsRNA to the RNase active center.
Sequence:
MEDVEIKPRGYQLRLVDHLTKSNGIVYLPTGSGKTFVAILVLKRFSQDFDKPIESGGKRALFMCNTVELARQQAMAVRRCTNFKVGFYVGEQGVDDWTRGMWSDEIKKNQVLVGTAQVFLDMVTQTYVALSSLSVVIIDECHHGTGHHPFREFMRLFTIANQTKLPRVVGLTGVLIKGNEITNVATKLKELEITYRGNIITVSDTKEMENVMLYATKPTEVMVSFPHQEQVLTVTRLISAEIEKFYVSLDLMNIGVQPIRRSKSLQCLRDPSKKSFVKQLFNDFLYQMKEYGIYAASIAIISLIVEFDIKRRQAETLSVKLMHRTALTLCEKIRHLLVQKLQDMTYDDDDDNVNTEEVIMNFSTPKVQRFLMSLKVSFADKDPKDICCLVFVERRYTCKCIYGLLLNYIQSTPELRNVLTPQFMVGRNNISPDFESVLERKWQKSAIQQFRDGNANLMICSSVLEEGIDVQACNHVFILDPVKTFNMYVQSKGRARTTEAKFVLFTADKEREKTIQQIYQYRKAHNDIAEYLKDRVLEKTEPELYEIKGHFQDDIDPFTNENGAVLLPNNALAILHRYCQTIPTDAFGFVIPWFHVLQEDERDRIFGVSAKGKHVISINMPVNCMLRDTIYSDPMDNVKTAKISAAFKACKVLYSLGELNERFVPKTLKERVASIADVHFEHWNKYGDSVTATVNKADKSKDRTYKTECPLEFYDALPRVGEICYAYEIFLEPQFESCEYTEHMYLNLQTPRNYAILLRNKLPRLAEMPLFSNQGKLHVRVANAPLEVIIQNSEQLELLHQFHGMVFRDILKIWHPFFVLDRRSKENSYLVVPLILGAGEQKCFDWELMTNFRRLPQSHGSNVQQREQQPAPRPEDFEGKIVTQWYANYDKPMLVTKVHRELTPLSYMEKNQQDKTYYEFTMSKYGNRIGDVVHKDKFMIEVRDLTEQLTFYVHNRGKFNAKSKAKMKVILIPELCFNFNFPGDLWLKLIFLPSILNRMYFLLHAEALRKRFNTYLNLHLLPFNGTDYMPRPLEIDYSLKRNVDPLGNVIPTEDIEEPKSLLEPMPTKSIEASVANLEITEFENPWQKYMEPVDLSRNLLSTYPVELDYYYHFSVGNVCEMNEMDFEDKEYWAKNQFHMPTGNIYGNRTPAKTNANVPALMPSKPTVRGKVKPLLILQKTVSKEHITPAEQGEFLAAITASSAADVFDMERLEILGDSFLKLSATLYLASKYSDWNEGTLTEVKSKLVSNRNLLFCLIDADIPKTLNTIQFTPRYTWLPPGISLPHNVLALWRENPEFAKIIGPHNLRDLALGDEESLVKGNCSDINYNRFVEGCRANGQSFYAGADFSSEVNFCVGLVTIPNKVIADTLEALLGVIVKNYGLQHAFKMLEYFKICRADIDKPLTQLLNLELGGKKMRANVNTTEIDGFLINHYYLEKNLGYTFKDRRYLLQALTHPSYPTNRITGSYQELEFIGDAILDFLISAYIFENNTKMNPGALTDLRSALVNNTTLACICVRHRLHFFILAENAKLSEIISKFVNFQESQGHRVTNYVRILLEEADVQPTPLDLDDELDMTELPHANKCISQEAEKGVPPKGEFNMSTNVDVPKALGDVLEALIAAVYLDCRDLQRTWEVIFNLFEPELQEFTRKVPINHIRQLVEHKHAKPVFSSPIVEGETVMVSCQFTCMEKTIKVYGFGSNKDQAKLSAAKHALQQLSKCDA