Spliceosomal snRNPs are multicomponent particles that undergo a complex maturation pathway. Human Sm-class snRNAs are generated as 3'-end extended precursors, which are exported to the cytoplasm and assembled together with Sm proteins into core RNPs by the SMN complex. Here, we provide evidence that these pre-snRNA substrates contain compact, evolutionarily conserved secondary structures that overlap with the Sm binding site. These structural motifs in pre-snRNAs are predicted to interfere with Sm core assembly. We model structural rearrangements that lead to an open pre-snRNA conformation compatible with Sm protein interaction. The predicted rearrangement pathway is conserved in Metazoa and requires an external factor that initiates snRNA remodeling. We show that the essential helicase Gemin3, which is a component of the SMN complex, is crucial for snRNA structural rearrangements during snRNP maturation. The SMN complex thus facilitates ATP-driven structural changes in snRNAs that expose the Sm site and enable Sm protein binding.

• MeSH
• HeLa buňky MeSH
• jádro snRNP - proteiny genetika   MeSH
• lidé MeSH
• prekurzory RNA * metabolismus   MeSH
• proteinový komplex SMN metabolismus   MeSH
• ribonukleoproteiny malé jaderné metabolismus   MeSH
• RNA malá jaderná * metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• jádro snRNP - proteiny MeSH
• prekurzory RNA * MeSH
• proteinový komplex SMN MeSH
• ribonukleoproteiny malé jaderné MeSH
• RNA malá jaderná * MeSH

SART3 is a multifunctional protein that acts in several steps of gene expression, including assembly and recycling of the spliceosomal U4/U6 small nuclear ribonucleoprotein particle (snRNP). In this work, we provide evidence that SART3 associates via its N-terminal HAT domain with the 12S U2 snRNP. Further analysis showed that SART3 associates with the post-splicing complex containing U2 and U5 snRNP components. In addition, we observed an interaction between SART3 and the RNA helicase DHX15, which disassembles post-splicing complexes. Based on our data, we propose a model that SART3 associates via its N-terminal HAT domain with the post-splicing complex, where it interacts with U6 snRNA to protect it and to initiate U6 snRNA recycling before a next round of splicing.

• Klíčová slova
• Recycling, Splicing, U2 snRNP, U6 snRNA,
• MeSH
• malý jaderný ribonukleoprotein U2 genetika   metabolismus   MeSH
• malý jaderný ribonukleoprotein U4-U6 genetika   metabolismus   MeSH
• malý jaderný ribonukleoprotein U5 genetika   metabolismus   MeSH
• ribonukleoproteiny malé jaderné genetika   metabolismus   MeSH
• RNA malá jaderná genetika   metabolismus   MeSH
• sestřih RNA * genetika   MeSH
• spliceozomy * genetika   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• malý jaderný ribonukleoprotein U2 MeSH
• malý jaderný ribonukleoprotein U4-U6 MeSH
• malý jaderný ribonukleoprotein U5 MeSH
• ribonukleoproteiny malé jaderné MeSH
• RNA malá jaderná MeSH

The coordination of cell division with stress response is essential for maintaining genome stability in plant meristems. Proteins involved in pre-mRNA splicing are important for these processes in animal and human cells. Based on its homology to the splicing factor SART1, which is implicated in the control of cell division and genome stability in human cells, we analyzed if MDF has similar functions in plants. We found that MDF associates with U4/U6.U5 tri-snRNP proteins and is essential for correct splicing of 2,037 transcripts. Loss of MDF function leads to cell division defects and cell death in meristems and was associated with up-regulation of stress-induced genes and down-regulation of mitotic regulators. In addition, the mdf-1 mutant is hypersensitive to DNA damage treatment supporting its role in coordinating stress response with cell division. Our analysis of a dephosphomutant of MDF suggested how its protein activity might be controlled. Our work uncovers the conserved function of a plant splicing factor and provides novel insight into the interplay of pre-mRNA processing and genome stability in plants.

• MeSH
• Arabidopsis * genetika   metabolismus   MeSH
• buněčné dělení genetika   MeSH
• lidé MeSH
• malý jaderný ribonukleoprotein U4-U6 genetika   metabolismus   MeSH
• malý jaderný ribonukleoprotein U5 * genetika   metabolismus   MeSH
• nestabilita genomu MeSH
• prekurzory RNA genetika   metabolismus   MeSH
• sestřihové faktory genetika   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• AT5G16780 protein, Arabidopsis MeSH Prohlížeč
• malý jaderný ribonukleoprotein U4-U6 MeSH
• malý jaderný ribonukleoprotein U5 * MeSH
• milk-derived factor MeSH Prohlížeč
• prekurzory RNA MeSH
• sestřihové faktory MeSH

RNA splicing, the process of intron removal from pre-mRNA, is essential for the regulation of gene expression. It is controlled by the spliceosome, a megadalton RNA-protein complex that assembles de novo on each pre-mRNA intron through an ordered assembly of intermediate complexes1,2. Spliceosome activation is a major control step that requires substantial protein and RNA rearrangements leading to a catalytically active complex1-5. Splicing factor 3B subunit 1 (SF3B1) protein-a subunit of the U2 small nuclear ribonucleoprotein6-is phosphorylated during spliceosome activation7-10, but the kinase that is responsible has not been identified. Here we show that cyclin-dependent kinase 11 (CDK11) associates with SF3B1 and phosphorylates threonine residues at its N terminus during spliceosome activation. The phosphorylation is important for the association between SF3B1 and U5 and U6 snRNAs in the activated spliceosome, termed the Bact complex, and the phosphorylation can be blocked by OTS964, a potent and selective inhibitor of CDK11. Inhibition of CDK11 prevents spliceosomal transition from the precatalytic complex B to the activated complex Bact and leads to widespread intron retention and accumulation of non-functional spliceosomes on pre-mRNAs and chromatin. We demonstrate a central role of CDK11 in spliceosome assembly and splicing regulation and characterize OTS964 as a highly selective CDK11 inhibitor that suppresses spliceosome activation and splicing.

• MeSH
• aktivace enzymů účinky léků   MeSH
• chinolony farmakologie   MeSH
• chromatin metabolismus   MeSH
• cyklin-dependentní kinasy * antagonisté a inhibitory   metabolismus   MeSH
• fosfoproteiny * chemie   metabolismus   MeSH
• fosforylace MeSH
• malý jaderný ribonukleoprotein U2 * chemie   metabolismus   MeSH
• prekurzory RNA * genetika   metabolismus   MeSH
• sestřih RNA * účinky léků   MeSH
• spliceozomy * účinky léků   metabolismus   MeSH
• threonin metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• chinolony MeSH
• chromatin MeSH
• cyklin-dependentní kinasy * MeSH
• fosfoproteiny * MeSH
• malý jaderný ribonukleoprotein U2 * MeSH
• OTS964 MeSH Prohlížeč
• prekurzory RNA * MeSH
• threonin MeSH

U5 snRNP is a complex particle essential for RNA splicing. U5 snRNPs undergo intricate biogenesis that ensures that only a fully mature particle assembles into a splicing competent U4/U6•U5 tri-snRNP and enters the splicing reaction. During splicing, U5 snRNP is substantially rearranged and leaves as a U5/PRPF19 post-splicing particle, which requires re-generation before the next round of splicing. Here, we show that a previously uncharacterized protein TSSC4 is a component of U5 snRNP that promotes tri-snRNP formation. We provide evidence that TSSC4 associates with U5 snRNP chaperones, U5 snRNP and the U5/PRPF19 particle. Specifically, TSSC4 interacts with U5-specific proteins PRPF8, EFTUD2 and SNRNP200. We also identified TSSC4 domains critical for the interaction with U5 snRNP and the PRPF19 complex, as well as for TSSC4 function in tri-snRNP assembly. TSSC4 emerges as a specific chaperone that acts in U5 snRNP de novo biogenesis as well as post-splicing recycling.

• MeSH
• down regulace MeSH
• elongační faktory MeSH
• enzymy opravy DNA metabolismus   MeSH
• HeLa buňky MeSH
• interakční proteinové domény a motivy MeSH
• jaderné proteiny metabolismus   MeSH
• lidé MeSH
• malý jaderný ribonukleoprotein U5 chemie   metabolismus   MeSH
• nádorové supresorové proteiny chemie   genetika   metabolismus   MeSH
• proteinové domény MeSH
• proteiny vázající RNA metabolismus   MeSH
• rekombinantní fúzní proteiny MeSH
• ribonukleoproteiny malé jaderné chemie   metabolismus   MeSH
• sestřih RNA MeSH
• sestřihové faktory metabolismus   MeSH
• spliceozomy metabolismus   MeSH
• transkripční faktory MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• EFTUD2 protein, human MeSH Prohlížeč
• elongační faktory MeSH
• enzymy opravy DNA MeSH
• jaderné proteiny MeSH
• malý jaderný ribonukleoprotein U5 MeSH
• nádorové supresorové proteiny MeSH
• proteiny vázající RNA MeSH
• PRPF19 protein, human MeSH Prohlížeč
• PRPF6 protein, human MeSH Prohlížeč
• PRPF8 protein, human MeSH Prohlížeč
• rekombinantní fúzní proteiny MeSH
• ribonukleoproteiny malé jaderné MeSH
• sestřihové faktory MeSH
• SNRNP200 protein, human MeSH Prohlížeč
• transkripční faktory MeSH
• TSSC4 protein, human MeSH Prohlížeč

A leading pharmacological strategy toward HIV cure requires "shock" or activation of HIV gene expression in latently infected cells with latency reversal agents (LRAs) followed by their subsequent clearance. In a screen for novel LRAs, we used fungal secondary metabolites as a source of bioactive molecules. Using orthogonal mass spectrometry (MS) coupled to latency reversal bioassays, we identified gliotoxin (GTX) as a novel LRA. GTX significantly induced HIV-1 gene expression in latent ex vivo infected primary cells and in CD4+ T cells from all aviremic HIV-1+ participants. RNA sequencing identified 7SK RNA, the scaffold of the positive transcription elongation factor b (P-TEFb) inhibitory 7SK small nuclear ribonucleoprotein (snRNP) complex, to be significantly reduced upon GTX treatment of CD4+ T cells. GTX directly disrupted 7SK snRNP by targeting La-related protein 7 (LARP7), releasing active P-TEFb, which phosphorylated RNA polymerase II (Pol II) C-terminal domain (CTD), inducing HIV transcription.

• MeSH
• gliotoxin * metabolismus   MeSH
• HeLa buňky MeSH
• HIV infekce * farmakoterapie   MeSH
• HIV-1 * metabolismus   MeSH
• lidé MeSH
• proteiny vázající RNA metabolismus   MeSH
• ribonukleoproteiny malé jaderné chemie   MeSH
• ribonukleoproteiny MeSH
• transkripční faktor B aktivující elongaci genetika   metabolismus   MeSH
• transkripční faktory metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• gliotoxin * MeSH
• Larp7 protein, human MeSH Prohlížeč
• proteiny vázající RNA MeSH
• ribonukleoproteiny malé jaderné MeSH
• ribonukleoproteiny MeSH
• transkripční faktor B aktivující elongaci MeSH
• transkripční faktory MeSH

Although atomistic explicit-solvent Molecular Dynamics (MD) is a popular tool to study protein-RNA recognition, satisfactory MD description of protein-RNA complexes is not always achieved. Unfortunately, it is often difficult to separate MD simulation instabilities primarily caused by the simple point-charge molecular mechanics (MM) force fields from problems related to the notorious uncertainties in the starting structures. Herein, we report a series of large-scale QM/MM calculations on the U1A protein-RNA complex. This experimentally well-characterized system has an intricate protein-RNA interface, which is very unstable in MD simulations. The QM/MM calculations identify several H-bonds poorly described by the MM method and thus indicate the sources of instabilities of the U1A interface in MD simulations. The results suggest that advanced QM/MM computations could be used to indirectly rationalize problems seen in MM-based MD simulations of protein-RNA complexes. As the most accurate QM method, we employ the computationally demanding meta-GGA density functional TPSS-D3(BJ)/def2-TZVP level of theory. Because considerably faster methods would be needed to extend sampling and to study even larger protein-RNA interfaces, a set of low-cost QM/MM methods is compared to the TPSS-D3(BJ)/def2-TZVP data. The PBEh-3c and B97-3c density functional composite methods appear to be suitable for protein-RNA interfaces. In contrast, HF-3c and the tight-binding Hamiltonians DFTB3-D3 and GFN-xTB perform unsatisfactorily and do not provide any advantage over the MM description. These conclusions are supported also by similar analysis of a simple HutP protein-RNA interface, which is well-described by MD with the exception of just one H-bond. Some other methodological aspects of QM/MM calculations on protein-RNA interfaces are discussed.

• MeSH
• Bacillus subtilis chemie   metabolismus   MeSH
• bakteriální proteiny chemie   metabolismus   MeSH
• kvantová teorie MeSH
• lidé MeSH
• malý jaderný ribonukleoprotein U1 chemie   metabolismus   MeSH
• proteiny vázající RNA chemie   metabolismus   MeSH
• RNA chemie   metabolismus   MeSH
• simulace molekulární dynamiky * ekonomika   MeSH
• software MeSH
• vodíková vazba MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bakteriální proteiny MeSH
• malý jaderný ribonukleoprotein U1 MeSH
• proteiny vázající RNA MeSH
• RNA MeSH
• U1A protein MeSH Prohlížeč

PUF60 is a splicing factor that binds uridine (U)-rich tracts and facilitates association of the U2 small nuclear ribonucleoprotein with primary transcripts. PUF60 deficiency (PD) causes a developmental delay coupled with intellectual disability and spinal, cardiac, ocular and renal defects, but PD pathogenesis is not understood. Using RNA-Seq, we identify human PUF60-regulated exons and show that PUF60 preferentially acts as their activator. PUF60-activated internal exons are enriched for Us upstream of their 3' splice sites (3'ss), are preceded by longer AG dinucleotide exclusion zones and more distant branch sites, with a higher probability of unpaired interactions across a typical branch site location as compared to control exons. In contrast, PUF60-repressed exons show U-depletion with lower estimates of RNA single-strandedness. We also describe PUF60-regulated, alternatively spliced isoforms encoding other U-bound splicing factors, including PUF60 partners, suggesting that they are co-regulated in the cell, and identify PUF60-regulated exons derived from transposed elements. PD-associated amino-acid substitutions, even within a single RNA recognition motif (RRM), altered selection of competing 3'ss and branch points of a PUF60-dependent exon and the 3'ss choice was also influenced by alternative splicing of PUF60. Finally, we propose that differential distribution of RNA processing steps detected in cells lacking PUF60 and the PUF60-paralog RBM39 is due to the RBM39 RS domain interactions. Together, these results provide new insights into regulation of exon usage by the 3'ss organization and reveal that germline mutation heterogeneity in RRMs can enhance phenotypic variability at the level of splice-site and branch-site selection.

• MeSH
• aminokyselinové motivy MeSH
• exony * MeSH
• HEK293 buňky MeSH
• HeLa buňky MeSH
• heterogenní jaderné ribonukleoproteiny metabolismus   MeSH
• jaderné proteiny metabolismus   MeSH
• krátké rozptýlené jaderné elementy MeSH
• lidé MeSH
• malý jaderný ribonukleoprotein U1 metabolismus   MeSH
• missense mutace * MeSH
• místa sestřihu RNA * MeSH
• proteiny vázající RNA metabolismus   MeSH
• represorové proteiny chemie   nedostatek   metabolismus   MeSH
• sekvenční analýza RNA MeSH
• sestřihové faktory chemie   nedostatek   metabolismus   MeSH
• sestřihový faktor U2AF MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• HCC1 autoantigen MeSH Prohlížeč
• heterogenní jaderné ribonukleoproteiny MeSH
• jaderné proteiny MeSH
• malý jaderný ribonukleoprotein U1 MeSH
• místa sestřihu RNA * MeSH
• poly-U binding splicing factor 60KDa MeSH Prohlížeč
• proteiny vázající RNA MeSH
• represorové proteiny MeSH
• sestřihové faktory MeSH
• sestřihový faktor U2AF MeSH
• SNRNP70 protein, human MeSH Prohlížeč

Split gene architecture of most human genes requires removal of intervening sequences by mRNA splicing that occurs on large multiprotein complexes called spliceosomes. Mutations compromising several spliceosomal components have been recorded in degenerative syndromes and haematological neoplasia, thereby highlighting the importance of accurate splicing execution in homeostasis of assorted adult tissues. Moreover, insufficient splicing underlies defective development of craniofacial skeleton and upper extremities. This review summarizes recent advances in the understanding of splicing factor function deduced from cryo-EM structures. We combine these data with the characterization of splicing factors implicated in hereditary or somatic disorders, with a focus on potential functional consequences the mutations may elicit in spliceosome assembly and/or performance. Given aberrant splicing or perturbations in splicing efficiency substantially underpin disease pathogenesis, profound understanding of the mis-splicing principles may open new therapeutic vistas. In three major sections dedicated to retinal dystrophies, hereditary acrofacial syndromes, and haematological malignancies, we delineate the noticeable variety of conditions associated with dysfunctional splicing and accentuate recurrent patterns in splicing defects.

• Klíčová slova
• Congenital craniofacial disorders, Haematological malignancies, Mutations, Retinopathy, Spliceosome,
• MeSH
• elektronová kryomikroskopie MeSH
• konformace proteinů MeSH
• lidé MeSH
• mutace MeSH
• nemoc genetika   MeSH
• prekurzory RNA genetika   MeSH
• ribonukleoproteiny malé jaderné chemie   genetika   ultrastruktura   MeSH
• sestřih RNA * MeSH
• spliceozomy genetika   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• prekurzory RNA MeSH
• ribonukleoproteiny malé jaderné MeSH

Kabuki syndrome is mainly caused by dominant de-novo pathogenic variants in the KMT2D and KDM6A genes. The clinical features of this syndrome are highly variable, making the diagnosis of Kabuki-like phenotypes difficult, even for experienced clinical geneticists. Herein we present molecular genetic findings of causal genetic variation using array comparative genome hybridization and a Mendeliome analysis, utilizing targeted exome analysis focusing on regions harboring rare disease-causing variants in Kabuki-like patients which remained KMT2D/KDM6A-negative. The aCGH analysis revealed a pathogenic CNV in the 14q11.2 region, while targeted exome sequencing revealed pathogenic variants in genes associated with intellectual disability (HUWE1, GRIN1), including a gene coding for mandibulofacial dysostosis with microcephaly (EFTUD2). Lower values of the MLL2-Kabuki phenotypic score are indicative of Kabuki-like phenotype (rather than true Kabuki syndrome), where aCGH and Mendeliome analyses have high diagnostic yield. Based on our findings we conclude that for new patients with Kabuki-like phenotypes it is possible to choose a specific molecular testing approach that has the highest detection rate for a given MLL2-Kabuki score, thus fostering more precise patient diagnosis and improved management in these genetically- and phenotypically heterogeneous clinical entities.

• Klíčová slova
• EFTUD2, GRIN1, HUWE1, Intellectual disability, KMT2D, Kabuki syndrome,
• MeSH
• dítě MeSH
• DNA vazebné proteiny genetika   MeSH
• elongační faktory genetika   MeSH
• exom MeSH
• fenotyp * MeSH
• genetická heterogenita * MeSH
• genotyp * MeSH
• histondemethylasy genetika   MeSH
• jaderné proteiny genetika   MeSH
• krevní nemoci diagnóza   genetika   patofyziologie   MeSH
• lidé MeSH
• lidské chromozomy, pár 14 MeSH
• malý jaderný ribonukleoprotein U5 genetika   MeSH
• mandibulofaciální dysostóza genetika   MeSH
• mentální retardace genetika   MeSH
• mikrocefalie genetika   MeSH
• mnohočetné abnormality diagnóza   genetika   patofyziologie   MeSH
• nádorové proteiny genetika   MeSH
• nádorové supresorové proteiny genetika   MeSH
• obličej abnormality   patofyziologie   MeSH
• předškolní dítě MeSH
• proteiny nervové tkáně genetika   MeSH
• receptory N-methyl-D-aspartátu genetika   MeSH
• srovnávací genomová hybridizace MeSH
• ubikvitinligasy genetika   MeSH
• vestibulární nemoci diagnóza   genetika   patofyziologie   MeSH
• vysoce účinné nukleotidové sekvenování MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH
• Názvy látek
• DNA vazebné proteiny MeSH
• EFTUD2 protein, human MeSH Prohlížeč
• elongační faktory MeSH
• GRIN1 protein, human MeSH Prohlížeč
• histondemethylasy MeSH
• HUWE1 protein, human MeSH Prohlížeč
• jaderné proteiny MeSH
• KDM6A protein, human MeSH Prohlížeč
• KMT2D protein, human MeSH Prohlížeč
• malý jaderný ribonukleoprotein U5 MeSH
• nádorové proteiny MeSH
• nádorové supresorové proteiny MeSH
• proteiny nervové tkáně MeSH
• receptory N-methyl-D-aspartátu MeSH
• ubikvitinligasy MeSH