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In mammalian cells, spurious transcription results in a vast repertoire of unproductive non-coding RNAs, whose deleterious accumulation is prevented by rapid decay. The nuclear exosome targeting (NEXT) complex plays a central role in directing non-functional transcripts to exosome-mediated degradation, but the structural and molecular mechanisms remain enigmatic. Here, we elucidated the architecture of the human NEXT complex, showing that it exists as a dimer of MTR4-ZCCHC8-RBM7 heterotrimers. Dimerization preconfigures the major MTR4-binding region of ZCCHC8 and arranges the two MTR4 helicases opposite to each other, with each protomer able to function on many types of RNAs. In the inactive state of the complex, the 3' end of an RNA substrate is enclosed in the MTR4 helicase channel by a ZCCHC8 C-terminal gatekeeping domain. The architecture of a NEXT-exosome assembly points to the molecular and regulatory mechanisms with which the NEXT complex guides RNA substrates to the exosome., Competing Interests: Declaration of interests E.C. is a member of the Molecular Cell advisory board., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

The superkiller (SKI) complex is the cytoplasmic co-factor and regulator of the RNA-degrading exosome. In human cells, the SKI complex functions mainly in co-translational surveillance-decay pathways, and its malfunction is linked to a severe congenital disorder, the trichohepatoenteric syndrome. To obtain insights into the molecular mechanisms regulating the human SKI (hSKI) complex, we structurally characterized several of its functional states in the context of 80S ribosomes and substrate RNA. In a prehydrolytic ATP form, the hSKI complex exhibits a closed conformation with an inherent gating system that effectively traps the 80S-bound RNA into the hSKI2 helicase subunit. When active, hSKI switches to an open conformation in which the gating is released and the RNA 3' end exits the helicase. The emerging picture is that the gatekeeping mechanism and architectural remodeling of hSKI underpin a regulated RNA channeling system that is mechanistically conserved among the cytoplasmic and nuclear helicase-exosome complexes., Competing Interests: Declaration of interests E.C. is a member of the Molecular Cell advisory board., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

The RNA-degrading exosome mediates the processing and decay of many cellular transcripts. In the yeast nucleus, the ubiquitous 10-subunit exosome core complex (Exo-9-Rrp44) functions with four conserved cofactors (Rrp6, Rrp47, Mtr4, and Mpp6). Biochemical and structural studies to date have shed insights into the mechanisms of the exosome core and its nuclear cofactors, with the exception of Mpp6. We report the 3.2-Å resolution crystal structure of a S. cerevisiae Exo-9-Mpp6 complex, revealing how linear motifs in the Mpp6 middle domain bind Rrp40 via evolutionary conserved residues. In particular, Mpp6 binds near a tryptophan residue of Rrp40 that is mutated in human patients suffering from pontocerebellar hypoplasia. Using biochemical assays, we show that Mpp6 is required for the ability of Mtr4 to extend the trajectory of an RNA entering the exosome core, suggesting that it promotes the channeling of substrates from the nuclear helicase to the processive RNase., (Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.)

Recent experimental studies revealed structural details of 3' to 5' degradation of RNA molecules, performed by the exosome complex. ssRNA is channeled through its multisubunit ring-like core into the active site tunnel of its key exonuclease subunit Rrp44, which acts both as an enzyme and a motor. Even in isolation, Rrp44 can pull and sequentially cleave RNA nucleotides, one at a time, without any external energy input and release a final 3-5 nucleotide long product. Using molecular dynamics simulations, we identify the main factors that control these processes. Our free energy calculations reveal that RNA transfer from solution into the active site of Rrp44 is highly favorable, but dependent on the length of the RNA strand. While RNA strands formed by 5 nucleotides or more correspond to a decreasing free energy along the translocation coordinate toward the cleavage site, a 4-nucleotide RNA experiences a free energy barrier along the same direction, potentially leading to incomplete cleavage of ssRNA and the release of short (3-5) nucleotide products. We provide new insight into how Rrp44 catalyzes a localized enzymatic reaction and performs an action distributed over several RNA nucleotides, leading eventually to the translocation of whole RNA segments into the position suitable for cleavage.

The MLE helicase remodels the roX lncRNAs, enabling the lncRNA-mediated assembly of the Drosophila dosage compensation complex. We identified a stable MLE core comprising the DExH helicase module and two auxiliary domains: a dsRBD and an OB-like fold. MLEcore is an unusual DExH helicase that can unwind blunt-ended RNA duplexes and has specificity for uridine nucleotides. We determined the 2.1 Å resolution structure of MLEcore bound to a U10 RNA and ADP-AlF4. The OB-like and dsRBD folds bind the DExH module and contribute to form the entrance of the helicase channel. Four uridine nucleotides engage in base-specific interactions, rationalizing the conservation of uridine-rich sequences in critical roX substrates. roX2 binding is orchestrated by MLE's auxiliary domains, which is prerequisite for MLE localization to the male X chromosome. The structure visualizes a transition-state mimic of the reaction and suggests how eukaryotic DEAH/RHA helicases couple ATP hydrolysis to RNA translocation., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The Ccr4-Not complex is involved in several aspects of gene expression, including mRNA decay, translational repression and transcription. We determined the 2.8-Å-resolution crystal structure of a 120-kDa core complex of the Saccharomyces cerevisiae Not module comprising the C-terminal arm of Not1, Not2 and Not5. Not1 is a HEAT-repeat scaffold. Not2 and Not5 have extended regions that wrap around Not1 and around their globular domains, the Not boxes. The Not boxes resemble Sm folds and interact with each other with a noncanonical dimerization surface. Disruption of the interactions within the ternary complex has severe effects on growth in vivo. The ternary complex forms a composite surface that binds poly(U) RNA in vitro, with a site at the Not5 Not box. The results suggest that the Not module forms a versatile platform for macromolecular interactions.

Translational repression and deadenylation of eukaryotic mRNAs result either in the sequestration of the transcripts in a nontranslatable pool or in their degradation. Removal of the 5' cap structure is a crucial step that commits deadenylated mRNAs to 5'-to-3' degradation. Pat1, Edc3 and the DEAD-box protein Dhh1 are evolutionary conserved factors known to participate in both translational repression and decapping, but their interplay is currently unclear. We report the 2.8 Å resolution structure of yeast Dhh1 bound to the N-terminal domain of Pat1. The structure shows how Pat1 wraps around the C-terminal RecA domain of Dhh1, docking onto the Phe-Asp-Phe (FDF) binding site. The FDF-binding site of Dhh1 also recognizes Edc3, revealing why the binding of Pat1 and Edc3 on Dhh1 are mutually exclusive events. Using co-immunoprecipitation assays and structure-based mutants, we demonstrate that the mode of Dhh1-Pat1 recognition is conserved in humans. Pat1 and Edc3 also interfere and compete with the RNA-binding properties of Dhh1. Mapping the RNA-binding sites on Dhh1 with a crosslinking-mass spectrometry approach shows a large RNA-binding surface around the C-terminal RecA domain, including the FDF-binding pocket. The results suggest a model for how Dhh1-containing messenger ribonucleoprotein particles might be remodeled upon Pat1 and Edc3 binding.

The exosome is the major 3'-5' RNA-degradation complex in eukaryotes. The ubiquitous core of the yeast exosome (Exo-10) is formed by nine catalytically inert subunits (Exo-9) and a single active RNase, Rrp44. In the nucleus, the Exo-10 core recruits another nuclease, Rrp6. Here we crystallized an approximately 440-kilodalton complex of Saccharomyces cerevisiae Exo-10 bound to a carboxy-terminal region of Rrp6 and to an RNA duplex with a 3'-overhang of 31 ribonucleotides. The 2.8 Å resolution structure shows how RNA is funnelled into the Exo-9 channel in a single-stranded conformation by an unwinding pore. Rrp44 adopts a closed conformation and captures the RNA 3'-end that exits from the side of Exo-9. Exo-9 subunits bind RNA with sequence-unspecific interactions reminiscent of archaeal exosomes. The substrate binding and channelling mechanisms of 3'-5' RNA degradation complexes are conserved in all kingdoms of life.

The exosome is a conserved macromolecular complex essential for RNA degradation. The nine-subunit core of the eukaryotic exosome shares a similar barrel-like architecture with prokaryotic complexes, but is catalytically inert. Here, we investigate how the Rrp44 nuclease functions in the active ten-subunit exosome. The 3.0 A resolution crystal structure of the yeast Rrp44-Rrp41-Rrp45 complex shows how the nuclease interacts with the exosome core and the relative accessibility of its endoribonuclease and exoribonuclease sites. Biochemical studies indicate that RNAs thread through the central channel of the core to reach the Rrp44 exoribonuclease site. This channeling mechanism involves evolutionary conserved residues. It allows the processive unwinding and degradation of RNA duplexes containing a sufficiently long single-stranded 3' extension, without the requirement for helicase activities. Although the catalytic function of the exosome core has been lost during evolution, the substrate recruitment and binding properties have been conserved from prokaryotes to eukaryotes.

The RNA exosome participates in the degradation and processing of a wide range of RNA molecules. Recent advances in understanding how the exosome is organized and functions largely stem from structural studies. Crystal structures of archaeal exosomes bound to RNA and of the corresponding nine-subunit human exosome core show that the archaeal and eukaryotic complexes have a similar molecular architecture, but have a diverged catalytic mechanism. The crystal structures of two hydrolytic RNases that associate with the exosome provide the framework for their catalytic activity. Negative-stain EM reconstructions give us a first glimpse of how they associate with the core complex. Together, these structural studies have implications for the mechanism of RNA recruitment and degradation by the exosome complexes.

Exosomes and proteasomes are macromolecular complexes that posttranscriptionally regulate gene expression by degrading mRNAs and proteins, respectively. Although the two complexes act on different substrates and are composed of different subunits, they share a similar barrel-like architecture that appears to have evolved to restrict substrate access and prevent indiscriminate degradation.

We talk to authors Achim Keidel and Elena Conti about their paper "Concerted structural rearrangements enable RNA channeling into the cytoplasmic Ski238-Ski7-exosome assembly" (in this issue of Molecular Cell), staying focused on the scientific question while being open to new approaches and their preferred way to celebrate good news. [ABSTRACT FROM AUTHOR]

The RNA exosome is an evolutionarily conserved multi-protein complex involved in the 3′ degradation of a variety of RNA transcripts. In the nucleus, the exosome participates in the maturation of structured RNAs, in the surveillance of pre-mRNAs and in the decay of a variety of noncoding transcripts. In the cytoplasm, the exosome degrades mRNAs in constitutive and regulated turnover pathways. Several structures of subcomplexes of eukaryotic exosomes or related prokaryotic exosome-like complexes are known, but how the complete assembly is organized to fulfil processive RNA degradation has been unclear. An atomic snapshot of a Saccharomyces cerevisiae 420 kDa exosome complex bound to an RNA substrate in the pre-cleavage state of a hydrolytic reaction has been determined. Here, the crystallographic steps towards the structural elucidation, which was carried out by molecular replacement, are presented. [ABSTRACT FROM AUTHOR]

Defective RNAs and proteins are swiftly degraded by cellular quality control mechanisms. A large fraction of their degradation is mediated by the exosome and the proteasome. These complexes have a similar architectural framework based on cylindrical, hollow structures that are conserved from bacteria and archaea to eukaryotes. Mechanistic similarities have also been identified for how RNAs and proteins are channelled into these structures and prepared for degradation. Insights gained from studies of the proteasome should now set the stage for elucidating the regulation, assembly and small-molecule inhibition of the exosome. [ABSTRACT FROM AUTHOR]

Protein and RNA molecules are exchanged between the nucleus and cytoplasm by members of the karyopherin β family of transport factors. Karyopherins adopt a modular HEAT-repeat architecture and are regulated by the GTPase Ran. RanGTP acts as a signal for the nuclear compartment, dissociating molecular cargo from karyopherins that mediate nuclear import and promoting cargo uptake on those mediating nuclear export. After unraveling the mechanisms of nuclear import factors, structural studies have recently provided tremendous insights into nuclear export. The impact of RanGTP binding on the karyopherins ranges from large, global conformational changes to local, allosteric effects. A theme emerges where cargo recognition provides a molecular surveillance mechanism to prevent the transport of macromolecules in an inappropriate state. [Copyright &y& Elsevier]

SMG6 and SMG5 are essential factors in nonsense-mediated mRNA decay, a conserved pathway that degrades mRNAs with premature translation termination codons. Both SMG5 and SMG6 have been predicted to contain a C-terminal PIN (PilT N-terminus) domain, present in proteins with ribonuclease activity. We have determined the structures of human SMG5 and SMG6 PIN domains. Although they share a similar overall fold related to ribonucleases of the RNase H family, they have local differences at the putative active site. SMG6 has the canonical triad of acidic residues that are crucial in RNase H for nuclease activity, while SMG5 lacks key catalytic residues. The structural differences are reflected at the functional level. Only the PIN domain of SMG6 has degradation activity on single-stranded RNA in vitro. This difference in catalytic activity is conserved in Drosophila, where an SMG6 with an inactive PIN domain inhibits NMD in a dominant-negative manner. Our findings suggest that the NMD machinery has intrinsic nuclease activity that is likely to contribute to the rapid decay of mRNAs that terminate translation prematurely. [ABSTRACT FROM AUTHOR]

Summary: The exosome is a macromolecular complex that plays fundamental roles in the biogenesis and turnover of a large number of RNA species. Here we report the crystal structures of the Rrp41-Rrp42 core complex of the S. solfataricus exosome bound to short single-stranded RNAs and to ADP. The RNA binding cleft recognizes four nucleotides in a sequence-unspecific manner, mainly by electrostatic interactions with the phosphate groups. Interactions at the 2′ hydroxyls of the sugars provide specificity for RNA over DNA. The structures show both the bound substrate and the cleaved product of the reaction, suggesting a catalytic mechanism for the 3′-5′ phosphorolytic activity of the exosome. [Copyright &y& Elsevier]

The exosome is a 3′ → 5′ exoribonuclease complex involved in RNA processing. We report the crystal structure of the RNase PH core complex of the Sulfolobus solfataricus exosome determined at a resolution of 2.8 Å. The structure reveals a hexameric ring-like arrangement of three Rrp41–Rrp42 heterodimers, where both subunits adopt the RNase PH fold common to phosphorolytic exoribonucleases. Structure-guided mutagenesis reveals that the activity of the complex resides within the active sites of the Rrp41 subunits, all three of which face the same side of the hexameric structure. The Rrp42 subunit is inactive but contributes to the structuring of the Rrp41 active site. The high sequence similarity of this archaeal exosome to eukaryotic exosomes and its high structural similarity to the bacterial mRNA–degrading PNPase support a common basis for RNA-degrading machineries in all three domains of life. [ABSTRACT FROM AUTHOR]

Nonsense-mediated mRNA decay (NMD) is an mRNA surveillance pathway that ensures the rapid degradation of mRNAs containing premature translation termination codons (PTCs), thereby preventing the synthesis of truncated and potentially harmful proteins. In addition, this pathway regulates the expression of ∼10% of the transcriptome and is essential in mice. Although NMD is conserved in eukaryotes, recent studies in several organisms have revealed that different mechanisms have evolved to discriminate natural from premature stop codons and to degrade the targeted mRNAs. With the elucidation of the first crystal structures of components of the NMD machinery, the way is paved towards a molecular understanding of the protein interaction network underlying this process. [Copyright &y& Elsevier]

The association between Mtr2 and Mex67 is essential for the nuclear export of bulk messenger RNA in yeast. In metazoans, the analogous function is carried out by the TAP-p15 heterodimer. Whereas Mex67 and TAP are highly conserved proteins, their binding partners, Mtr2 and p15 share no sequence similarity, but are nevertheless functionally homologous. Here, we report the 2.8-Å resolution crystal structure of Mtr2 in complex with the NTF2-like domain of Mex67. Mtr2 is a novel member of the NTF2-like family and interacts with Mex67, forming a complex with a similar structure architecture to that of TAP-p15. Mtr2 fulfills an analogous function to that of human p15 in maintaining the structural integrity of the heterodimer. In addition, Mtr2 presents a long internal loop, which contains residues that affect the export of the large ribosomal subunit. [ABSTRACT FROM AUTHOR]

Y14 and Mago are conserved eukaryotic proteins that associate with spliced mRNAs in the nucleus and remain associated at exon junctions during and after nuclear export. In the cytoplasm, Y14 is involved in mRNA quality control via the nonsense-mediated mRNA decay (NMD) pathway and, together with Mago, is involved in localization of osk (oskar) mRNA. We have determined the crystal structure of the complex between Drosophila melanogaster Y14 and Mago at a resolution of 2.5 A. The structure reveals an atypical mode of protein-protein recognition mediated by an RNA-binding domain (RBD). Instead of binding RNA, the RBD of Y14 engages its RNP1 and RNP2 motifs to bind Mago. Using structure-guided mutagenesis, we show that Mago is also a component of the NMD pathway, and that its association with Y14 is essential for function. Heterodimerization creates a single structural platform that interacts with the NMD machinery via phylogenetically conserved residues. [ABSTRACT FROM AUTHOR]

The Ski2-Ski3-Ski8 (Ski238) helicase complex directs cytoplasmic mRNAs toward the nucleolytic exosome complex for degradation. In yeast, the interaction between Ski238 and exosome requires the adaptor protein Ski7. We determined different cryo-EM structures of the Ski238 complex depicting the transition from a rigid autoinhibited closed conformation to a flexible active open conformation in which the Ski2 helicase module has detached from the rest of Ski238. The open conformation favors the interaction of the Ski3 subunit with exosome-bound Ski7, leading to the recruitment of the exosome. In the Ski238-Ski7-exosome holocomplex, the Ski2 helicase module binds the exosome cap, enabling the RNA to traverse from the helicase through the internal exosome channel to the Rrp44 exoribonuclease. Our study pinpoints how conformational changes within the Ski238 complex regulate exosome recruitment for RNA degradation. We also reveal the remarkable conservation of helicase-exosome RNA channeling mechanisms throughout eukaryotic nuclear and cytoplasmic exosome complexes. [Display omitted] • Structures of sequential steps in cytoplasmic exosome-mediated RNA degradation • Concerted binding and detachment events in Ski238 lead to exosome recruitment • RNA is threaded from the Ski2 helicase to the exosome ribonuclease • Conserved RNA channeling mechanisms between cytoplasmic and nuclear exosomes Keidel et al. present a structural view of the interplay between the cytoplasmic RNA exosome and its cofactor, the Ski238 complex, revealing the connection between the helicase activity of Ski2 and the nuclease activity of Rrp44. [ABSTRACT FROM AUTHOR]

The RNA exosome was originally discovered in yeast as an RNA-processing complex required for the maturation of 5.8S ribosomal RNA (rRNA), one of the constituents of the large ribosomal subunit. The exosome is now known in eukaryotes as the major 3 —5' RNA degradation machine involved in numerous processing, turnover, and surveillance pathways, both in the nucleus and the cytoplasm. Yet its role in maturing the 5.8S rRNA in the pre-60S ribosomal particle remains probably the most intricate and emblematic among its functions, as it involves all the RNA unwinding, degradation, and trimming activities embedded in this macromolecular complex. Here, we propose a comprehensive mechanistic model, based on current biochemical and structural data, explaining the dual functions of the nuclear exosome—the constructive versus the destructive mode. [ABSTRACT FROM AUTHOR]

The RNA-binding ARS2 protein is centrally involved in both early RNA polymerase II (RNAPII) transcription termination and transcript decay. Despite its essential nature, the mechanisms by which ARS2 enacts these functions have remained unclear. Here, we show that a conserved basic domain of ARS2 binds a corresponding acidic-rich, short linear motif (SLiM) in the transcription restriction factor ZC3H4. This interaction recruits ZC3H4 to chromatin to elicit RNAPII termination, independent of other early termination pathways defined by the cleavage and polyadenylation (CPA) and Integrator (INT) complexes. We find that ZC3H4, in turn, forms a direct connection to the nuclear exosome targeting (NEXT) complex, hereby facilitating rapid degradation of the nascent RNA. Hence, ARS2 instructs the coupled transcription termination and degradation of the transcript onto which it is bound. This contrasts with ARS2 function at CPA-instructed termination sites where the protein exclusively partakes in RNA suppression via post-transcriptional decay. [Display omitted] • ARS2 transcription termination activity is independent of the INT and CPA pathways • The ZC3H4 restrictor factor interacts with ARS2 via a short linear motif (SLiM) • ARS2 is required for ZC3H4 recruitment to chromatin at a number of loci • ARS2-ZC3H4 triggers transcription termination and decay of the cognate RNA Rouviere et al. show that the RNA-binding protein ARS2, previously shown to act in early transcription termination, does so by recruiting the ZC3H4 "restrictor" factor to chromatin. The ARS2-ZC3H4 axis commits terminated transcripts for degradation by the nuclear exosome via a direct recruitment of the nuclear exosome targeting (NEXT) complex. [ABSTRACT FROM AUTHOR]

The RNA exosome complex processes and degrades a wide range of transcripts, including ribosomal RNAs (rRNAs). We used cryo-electron microscopy to visualize the yeast nuclear exosome holocomplex captured on a precursor large ribosomal subunit (pre-60 S) during 7S-to-5.8S rRNA processing. The cofactors of the nuclear exosome are sandwiched between the ribonuclease core complex (Exo-10) and the remodeled "foot" structure of the pre-60S particle, which harbors the 5.8S rRNA precursor. The exosome-associated helicase Mtr4 recognizes the preribosomal substrate by docking to specific sites on the 25S rRNA, captures the 3′ extension of the 5.8 S rRNA, and channels it toward Exo-10. The structure elucidates how the exosome forms a structural and functional unit together with its massive pre-60 S substrate to process rRNA during ribosome maturation. [ABSTRACT FROM AUTHOR]

Summary The RNA exosome complex associates with nuclear and cytoplasmic cofactors to mediate the decay, surveillance, or processing of a wide variety of transcripts. In the cytoplasm, the conserved core of the exosome (Exo 10 ) functions together with the conserved Ski complex. The interaction of S. cerevisiae Exo 10 and Ski is not direct but requires a bridging cofactor, Ski7. Here, we report the 2.65 Å resolution structure of S. cerevisiae Exo 10 bound to the interacting domain of Ski7. Extensive hydrophobic interactions rationalize the high affinity and stability of this complex, pointing to Ski7 as a constitutive component of the cytosolic exosome. Despite the absence of sequence homology, cytoplasmic Ski7 and nuclear Rrp6 bind Exo 10 using similar surfaces and recognition motifs. Knowledge of the interacting residues in the yeast complexes allowed us to identify a splice variant of human HBS1-Like as a Ski7-like exosome-binding protein, revealing the evolutionary conservation of this cytoplasmic cofactor. [ABSTRACT FROM AUTHOR]

Summary The Cmr complex is an RNA-guided endonuclease that cleaves foreign RNA targets as part of the CRISPR prokaryotic defense system. We investigated the molecular architecture of the P. furiosus Cmr complex using an integrative structural biology approach. We determined crystal structures of P. furiosus Cmr1, Cmr2, Cmr4, and Cmr6 and combined them with known structural information to interpret the cryo-EM map of the complex. To support structure determination, we obtained residue-specific interaction data using protein crosslinking and mass spectrometry. The resulting pseudoatomic model reveals how the superhelical backbone of the complex is defined by the polymerizing principles of Cmr4 and Cmr5 and how it is capped at the extremities by proteins of similar folds. The inner surface of the superhelix exposes conserved residues of Cmr4 that we show are required for target-cleavage activity. The structural and biochemical data thus identify Cmr4 as the conserved endoribonuclease of the Cmr complex. [ABSTRACT FROM AUTHOR]

Summary: The Ski complex is a conserved multiprotein assembly required for the cytoplasmic functions of the exosome, including RNA turnover, surveillance, and interference. Ski2, Ski3, and Ski8 assemble in a tetramer with 1:1:2 stoichiometry. The crystal structure of an S. cerevisiae 370 kDa core complex shows that Ski3 forms an array of 33 TPR motifs organized in N-terminal and C-terminal arms. The C-terminal arm of Ski3 and the two Ski8 subunits position the helicase core of Ski2 centrally within the complex, enhancing RNA binding. The Ski3 N-terminal arm and the Ski2 insertion domain allosterically modulate the ATPase and helicase activities of the complex. Biochemical data suggest that the Ski complex can thread RNAs directly to the exosome, coupling the helicase and the exoribonuclease through a continuous RNA channel. Finally, we identify a Ski8-binding motif common to Ski3 and Spo11, rationalizing the moonlighting properties of Ski8 in mRNA decay and meiosis. [ABSTRACT FROM AUTHOR]

Summary: The eukaryotic exosome is a macromolecular complex essential for RNA processing and decay. It has recently been shown that the RNase activity of the yeast exosome core can be mapped to a single subunit, Rrp44, which processively degrades single-stranded RNAs as well as RNAs containing secondary structures. Here we present the 2.3 Å resolution crystal structure of S. cerevisiae Rrp44 in complex with single-stranded RNA. Although Rrp44 has a linear domain organization similar to bacterial RNase II, in three dimensions the domains have a different arrangement. The three domains of the classical nucleic-acid-binding OB fold are positioned on the catalytic domain such that the RNA-binding path observed in RNase II is occluded. Instead, RNA is threaded to the catalytic site via an alternative route suggesting a mechanism for RNA-duplex unwinding. The structure provides a molecular rationale for the observed biochemical properties of the RNase R family of nucleases. [Copyright &y& Elsevier]