Intrinsically disordered RNA-binding motifs cooperate to catalyze RNA folding and drive phase separation.

RNA-binding proteins are essential for gene regulation and the spatial organization of cells. Here, we report that the yeast ribosome biogenesis factor Loc1p is an intrinsically disordered RNA-binding protein with eight repeating positively charged, unstructured nucleic acid binding (PUN) motifs. While a single of these previously undefined motifs stabilizes folded RNAs, multiple copies strongly cooperate to catalyze RNA folding.

A naturally occurring 22-amino acid fragment of human hemoglobin A inhibits autophagy and HIV-1.

Autophagy is an evolutionarily ancient catabolic pathway and has recently emerged as an integral part of the innate immune system. While the core machinery of autophagy is well defined, the physiological regulation of autophagy is less understood. Here, we identify a C-terminal fragment of human hemoglobin A (HBA1, amino acids 111-132) in human bone marrow as a fast-acting non-inflammatory inhibitor of autophagy initiation.

PURA syndrome-causing mutations impair PUR-domain integrity and affect P-body association.

Mutations in the human gene cause the neurodevelopmental PURA syndrome. In contrast to several other monogenetic disorders, almost all reported mutations in this nucleic acid-binding protein result in the full disease penetrance. In this study, we observed that patient mutations across PURA impair its previously reported co-localization with processing bodies.

Human TRMT2A methylates tRNA and contributes to translation fidelity.

5-Methyluridine (m5U) is one of the most abundant RNA modifications found in cytosolic tRNA. tRNA methyltransferase 2 homolog A (hTRMT2A) is the dedicated mammalian enzyme for m5U formation at tRNA position 54. However, its RNA binding specificity and functional role in the cell are not well understood.

Formation of orogenic gold deposits by progressive movement of a fault-fracture mesh through the upper crustal brittle-ductile transition zone.

Orogenic gold deposits are comprised of complex quartz vein arrays that form as a result of fluid flow along transcrustal fault zones in active orogenic belts. Mineral precipitation in these deposits occurs under variable pressure conditions, but a mechanism explaining how the pressure regimes evolve through time has not previously been proposed. Here we show that extensional quartz veins at the Garrcon deposit in the Abitibi greenstone belt of Canada preserve petrographic characteristics suggesting that the three recognized paragenetic stages formed within different pressure regimes.

Compound heterozygous variants in OTULIN are associated with fulminant atypical late-onset ORAS.

Autoinflammatory diseases are a heterogenous group of disorders defined by fever and systemic inflammation suggesting involvement of genes regulating innate immune responses. Patients with homozygous loss-of-function variants in the OTU-deubiquitinase OTULIN suffer from neonatal-onset OTULIN-related autoinflammatory syndrome (ORAS) characterized by fever, panniculitis, diarrhea, and arthritis. Here, we describe an atypical form of ORAS with distinct clinical manifestation of the disease caused by two new compound heterozygous variants (c.

Disrupting Roquin-1 interaction with Regnase-1 induces autoimmunity and enhances antitumor responses.

Roquin and Regnase-1 proteins bind and post-transcriptionally regulate proinflammatory target messenger RNAs to maintain immune homeostasis. Either the sanroque mutation in Roquin-1 or loss of Regnase-1 cause systemic lupus erythematosus-like phenotypes. Analyzing mice with T cells that lack expression of Roquin-1, its paralog Roquin-2 and Regnase-1 proteins, we detect overlapping or unique phenotypes by comparing individual and combined inactivation.

OpenPepXL: An Open-Source Tool for Sensitive Identification of Cross-Linked Peptides in XL-MS.

Cross-linking MS (XL-MS) has been recognized as an effective source of information about protein structures and interactions. In contrast to regular peptide identification, XL-MS has to deal with a quadratic search space, where peptides from every protein could potentially be cross-linked to any other protein. To cope with this search space, most tools apply different heuristics for search space reduction.

Combining dehydration, construct optimization and improved data collection to solve the crystal structure of a CRM1-RanGTP-SPN1-Nup214 quaternary nuclear export complex.

High conformational flexibility is an intrinsic and indispensable property of nuclear transport receptors, which makes crystallization and structure determination of macromolecular complexes containing exportins or importins particularly challenging. Here, the crystallization and structure determination of a quaternary nuclear export complex consisting of the exportin CRM1, the small GTPase Ran in its GTP-bound form, the export cargo SPN1 and an FG repeat-containing fragment of the nuclear pore complex component nucleoporin Nup214 fused to maltose-binding protein is reported. Optimization of constructs, seeding and the development of a sophisticated protocol including successive PEG-mediated crystal dehydration as well as additional post-mounting steps were essential to obtain well diffracting crystals.

Structural and Functional Characterization of CRM1-Nup214 Interactions Reveals Multiple FG-Binding Sites Involved in Nuclear Export.

CRM1 is the major nuclear export receptor. During translocation through the nuclear pore, transport complexes transiently interact with phenylalanine-glycine (FG) repeats of multiple nucleoporins. On the cytoplasmic side of the nuclear pore, CRM1 tightly interacts with the nucleoporin Nup214.

Structural Basis of Targeting the Exportin CRM1 in Cancer.

Recent studies have demonstrated the interference of nucleocytoplasmic trafficking with the establishment and maintenance of various cancers. Nucleocytoplasmic transport is highly regulated and coordinated, involving different nuclear transport factors or receptors, importins and exportins, that mediate cargo transport from the cytoplasm into the nucleus or the other way round, respectively. The exportin CRM1 (Chromosome region maintenance 1) exports a plethora of different protein cargoes and ribonucleoprotein complexes.

Allosteric control of the exportin CRM1 unraveled by crystal structure analysis.

Nucleocytoplasmic trafficking in eukaryotic cells is a highly regulated and coordinated process which involves an increasing variety of soluble nuclear transport receptors. Generally, transport receptors specifically bind their cargo and facilitate its transition through nuclear pore complexes, aqueous channels connecting the two compartments. Directionality of such transport events by receptors of the importin β superfamily requires the interaction with the small GTPase Ras-related nuclear antigen (Ran).

Translation initiation factor eIF3b contains a nine-bladed β-propeller and interacts with the 40S ribosomal subunit.

The multisubunit eukaryotic translation initiation factor 3, among which the subunit b (eIF3b) is a major scaffold protein, plays essential roles in protein synthesis. Here, we report the crystal structure of the WD40 domain of Chaetomium thermophilum eIF3b, revealing a nine-bladed β-propeller fold. Sequence analysis indicates that this propeller architecture is common to all eIF3b orthologs.

Crystal structures of the novel cytosolic 5'-nucleotidase IIIB explain its preference for m7GMP.

5'-nucleotidases catalyze the hydrolytic dephosphorylation of nucleoside monophosphates. As catabolic enzymes they contribute significantly to the regulation of cellular nucleotide levels; misregulation of nucleotide metabolism and nucleotidase deficiencies are associated with a number of diseases. The seven human 5'-nucleotidases differ with respect to substrate specificity and cellular localization.

Structural determinants and mechanism of mammalian CRM1 allostery.

Proteins carrying nuclear export signals cooperatively assemble with the export factor CRM1 and the effector protein RanGTP. In lower eukaryotes, this cooperativity is coupled to CRM1 conformational changes; however, it is unknown if mammalian CRM1 maintains its compact conformation or shows similar structural flexibility. Here, combinations of small-angle X-ray solution scattering and electron microscopy experiments with molecular dynamics simulations reveal pronounced conformational flexibility in mammalian CRM1 and demonstrate that RanGTP binding induces association of its N- and C-terminal regions to form a toroid structure.

Structural basis for cooperativity of CRM1 export complex formation.

In eukaryotes, the nucleocytoplasmic transport of macromolecules is mainly mediated by soluble nuclear transport receptors of the karyopherin-β superfamily termed importins and exportins. The highly versatile exportin chromosome region maintenance 1 (CRM1) is essential for nuclear depletion of numerous structurally and functionally unrelated protein and ribonucleoprotein cargoes. CRM1 has been shown to adopt a toroidal structure in several functional transport complexes and was thought to maintain this conformation throughout the entire nucleocytoplasmic transport cycle.

NES consensus redefined by structures of PKI-type and Rev-type nuclear export signals bound to CRM1.

Classic nuclear export signals (NESs) confer CRM1-dependent nuclear export. Here we present crystal structures of the RanGTP-CRM1 complex alone and bound to the prototypic PKI or HIV-1 Rev NESs. These NESs differ markedly in the spacing of their key hydrophobic (Φ) residues, yet CRM1 recognizes them with the same rigid set of five Φ pockets.

Crystal structure of the nuclear export receptor CRM1 in complex with Snurportin1 and RanGTP.

CRM1 mediates nuclear export of numerous unrelated cargoes, which may carry a short leucine-rich nuclear export signal or export signatures that include folded domains. How CRM1 recognizes such a variety of cargoes has been unknown up to this point. Here we present the crystal structure of the SPN1.

Structural basis for m7G-cap hypermethylation of small nuclear, small nucleolar and telomerase RNA by the dimethyltransferase TGS1.

The 5'-cap of spliceosomal small nuclear RNAs, some small nucleolar RNAs and of telomerase RNA was found to be hypermethylated in vivo. The Trimethylguanosine Synthase 1 (TGS1) mediates this conversion of the 7-methylguanosine-cap to the 2,2,7-trimethylguanosine (m(3)G)-cap during maturation of the RNPs. For mammalian UsnRNAs the generated m(2,2,7)G-cap is one part of a bipartite import signal mediating the transport of the UsnRNP-core complex into the nucleus.

Structure analysis of the conserved methyltransferase domain of human trimethylguanosine synthase TGS1.

Methyltransferases play an important role in the post-transcriptional maturation of most ribonucleic acids. The modification of spliceosomal UsnRNAs includes N2-dimethylation of the m(7)G cap catalyzed by trimethylguanosine synthase 1 (TGS1). This 5'-cap hypermethylation occurs during the biogenesis of UsnRNPs as it initiates the m(3)G cap-dependent nuclear import of UsnRNPs.

Crystal structure of the RRM domain of poly(A)-specific ribonuclease reveals a novel m(7)G-cap-binding mode.

Poly(A)-specific ribonuclease (PARN) is a processive 3'-exoribonuclease involved in the decay of eukaryotic mRNAs. Interestingly, PARN interacts not only with the 3' end of the mRNA but also with its 5' end as PARN contains an RRM domain that specifically binds both the poly(A) tail and the 7-methylguanosine (m(7)G) cap. The interaction of PARN with the 5' cap of mRNAs stimulates the deadenylation activity and enhances the processivity of this reaction.