Reactome is pathway database which provides intuitive bioinformatics tools for the visualisation, interpretation and analysis of pathway knowledge.
Four separate CETP gene mutations have been published previously. Of the four mutations, the intron 14 donor splice site mutation4 and the Asp442-to-Gly mutation in exon 1528 are both known to be very common in the Japanese population.7 8 9 10 29 The other two mutations, the Gln309-to-Stop mutation in exon 1026 and a 1-bp insertion in intron 14,7 are probably sporadic mutations. Except for the Asp442-to-Gly mutation that causes partial CETP deficiency, the other three seemed to have null allelic effects, although exact mechanisms underlying the null effects have not been studied. In the present study, we demonstrated that the primary abnormality due to the intron 14 donor splice site mutation is the exon skipping of mRNA, which decreases the level of mRNA and produces a truncated protein that should be degraded intracellularly. These observations clearly explain the molecular basis of the complete CETP deficiency found not only in patients with the common intron 14 splicing mutation but also in ...
BioMed Research International is a peer-reviewed, Open Access journal that publishes original research articles, review articles, and clinical studies covering a wide range of subjects in life sciences and medicine. The journal is divided into 55 subject areas.
Somatically acquired mutations in components of the RNA processing pathway in CLL. Presented is an overview illustrating the individual components of the RNA-processing pathway, with those components identified as being somatically mutated highlighted (*) and the mutated protein listed in red. Initially, nascent pre-mRNA transcripts undergo 5′ capping and binding of the cap-binding complex (CBC), followed by the formation of the major spliceosome, the machinery responsible for the removal of pre-mRNA introns via a stepwise mechanism. Initial assembly steps include formation of pre-spliceosome complex A (top left nuclear complex) involving recognition of the 5′ splice site by U1 snRNP (an interaction stabilized by members of the serine-arginine-rich (SR) protein family) and recognition of the 3′ SS region by the U2 Auxiliary factor U2AF and by U2snRNP. U2AF binds to the intronic polypyrimidine tract and 3′SS, and facilitates binding of U2 snRNP to the branch-point sequence. Stable U2 ...
DNA- and RNA binding protein, involved in several nuclear processes. Essential pre-mRNA splicing factor required early in spliceosome formation and for splicing catalytic step II, probably as a heteromer with NONO. Binds to pre-mRNA in spliceosome C complex, and specifically binds to intronic polypyrimidine tracts. Involved in regulation of signal-induced alternative splicing. During splicing of PTPRC/CD45, a phosphorylated form is sequestered by THRAP3 from the pre-mRNA in resting T-cells; T-cell activation and subsequent reduced phosphorylation is proposed to lead to release from THRAP3 allowing binding to pre-mRNA splicing regulatotry elements which represses exon inclusion. Interacts with U5 snRNA, probably by binding to a purine-rich sequence located on the 3 side of U5 snRNA stem 1b. May be involved in a pre-mRNA coupled splicing and polyadenylation process as component of a snRNP-free complex with SNRPA/U1A. The SFPQ-NONO heteromer associated with MATR3 may play a role in nuclear ...
Plays a role in preventing exon skipping, ensuring the accuracy of splicing and regulating alternative splicing. Interacts with other spliceosomal components, via the RS domains, to form a bridge between the 5- and 3-splice site binding components, U1 snRNP and U2AF. Can stimulate binding of U1 snRNP to a 5-splice site-containing pre-mRNA. Binds to purine-rich RNA sequences, either the octamer, 5-RGAAGAAC-3 (r=A or G) or the decamers, AGGACAGAGC/AGGACGAAGC. Binds preferentially to the 5-CGAGGCG-3 motif in vitro. Three copies of the octamer constitute a powerful splicing enhancer in vitro, the ASF/SF2 splicing enhancer (ASE) which can specifically activate ASE-dependent splicing. Isoform ASF-2 and isoform ASF-3 act as splicing repressors. May function as export adapter involved in mRNA nuclear export through the TAP/NXF1 pathway ...
1. Mushroom: From Audobon Society Field Guide; mushrooms described in terms of physical characteristics; classification: poisonous or edible. 2. Plants: Data has been extracted from the USDA plants database. It contains all plants (species and genera) in the database and the states of USA and Canada where they occur.. 3. Molecular Biology (Splice-junction Gene Sequences): Primate splice-junction gene sequences (DNA) with associated imperfect domain theory ...
Group II Self-Splicing Introns. -pre-rRNA of fungal and plant mitochondria -majority of chloroplast introns -several classes -require Mg 2+ -no cofactor. Domain Structure of a Group II Intron. A. 5 exon. 3 exon. Chemistry of Group II Self-Splicing. 1st step. 2nd step. lariat. Slideshow 3387240 by guang
A splice site mutation is a genetic mutation that inserts, deletes or changes a number of nucleotides in the specific site at which splicing takes place during the processing of precursor messenger RNA into mature messenger RNA. Splice site consensus sequences that drive exon recognition are located at the very termini of introns. The deletion of the splicing site results in one or more introns remaining in mature mRNA and may lead to the production of abnormal proteins. When a splice site mutation occurs, the mRNA transcript possesses information from these introns that normally should not be included. Introns are supposed to be removed, while the exons are expressed. The mutation must occur at the specific site at which intron splicing occurs: within non-coding sites in a gene, directly next to the location of the exon. The mutation can be an insertion, deletion, frame shift, etc. The splicing process itself is controlled by the given sequences, known as splice-donor and splice-acceptor ...
The current model of spliceosome assembly was developed principally from the in vitro pattern of small nuclear ribonucleoprotein (snRNP) particle association with synthetic splicing substrates (reviewed in Moore et al., 1993; Madhani and Guthrie, 1994; Krämer, 1996). In mammals and yeast, spliceosome assembly progresses by the sequential addition of the U1 snRNP→U2 snRNP→U4/U6.U5 tri‐snRNP particles to the pre‐mRNA. Before 5′ splice‐site cleavage (chemical step I in splicing), the affinities of the U1 and U4 snRNAs for the splicing complex are greatly reduced and, under many (Pikielny et al., 1986; Cheng and Abelson, 1987; Konarska and Sharp, 1987) although not all (Blencowe et al., 1989) isolation conditions, the U4 snRNA is lost from the spliceosome. This model of spliceosome assembly is supported by the abridged spliceosome assembly profiles observed when splicing is inhibited by specific mutations in the pre‐mRNA or when one of the many trans‐acting components of splicing is ...
Pre-mRNA splicing takes place in an RNA machine known as the spliceosome, which consists of small nuclear ribonucleoprotein particles (snRNPs)1 and non-snRNP protein factors. The RNA components in the spliceosome form the catalytic core through a series of dynamic RNA-RNA interactions which are likely to be mediated and/or stabilized by non-snRNP protein factors (for recent reviews see Nilsen, 1998; Staley and Guthrie, 1998). Among the best characterized non-snRNP splicing factors are SR proteins which contain one or two RNA recognition motifs and a signature RS domain containing multiple serine and arginine repeats (for reviews see Fu, 1995; Manley and Tacke, 1996). The RNA recognition motifs bind to RNA sequences in a coordinated fashion to determine splicing specificity (Chandler et al., 1997) and commit pre-mRNA substrates to the splicing pathway (Fu, 1993), whereas the RS domains mediate specific protein- protein interactions in a number of spliceosomal assembly steps (Wu and Maniatis, ...
During spliceosome assembly, protein-protein interactions (PPI) are sequentially formed and disrupted to accommodate the spatial requirements of pre-mRNA substrate recognition and catalysis. Splicing activators and repressors, such as SR proteins and hnRNPs, modulate spliceosome assembly and regulate alternative splicing. However, it remains unclear how they differentially interact with the core spliceosome to perform their functions. Here, we investigate the protein connectivity of SR and hnRNP proteins to the core spliceosome using probabilistic network reconstruction based on the integration of interactome and gene expression data. We validate our model by immunoprecipitation and mass spectrometry of the prototypical splicing factors SRSF1 and hnRNPA1. Network analysis reveals that a factors properties as an activator or repressor can be predicted from its overall connectivity to the rest of the spliceosome. In addition, we discover and experimentally validate PPIs between the oncoprotein SRSF1 and
Many functional RNAs are required to fold into specific three-dimensional structures. A fundamental property of RNA is that its secondary structure and even some tertiary contacts are highly stable, which gives rise to independent modular RNA motifs and makes RNAs prone to adopting misfolded intermediates. Consequently, in addition to stabilizing the native structure relative to the unfolded species (defined here as stability), RNAs are faced with the challenge of stabilizing the native structure relative to alternative structures (defined as structural specificity). How RNAs have evolved to overcome these challenges is incompletely understood. Self-splicing group I introns have been used to study RNA structure and folding for decades. Among them, the Tetrahymena intron was the first discovered and has been studied extensively. In this work, we found that a version of the intron that was generated by in vitro selection for enhanced stability also displayed enhanced specificity against a stable ...
Assembly of the spliceosome by the stepwise binding of the snRNPs to the pre-mRNA. In the early phase of spliceosome assembly, the U1 snRNP binds to the 5 splice site (5 SS: where exon 1 ends and the intron begins), and the U2 snRNP binds to the so-called branch point (BP: near the 3 end of the intron). This spliceosome assembly intermediate is called the A complex. The subsequent binding of the U4/U6.U5 tri snRNP complex gives rise to the precatalytic B complex. The catalytic activation of the spliceosome takes place in two steps. In the first, the RNA helicase Brr2 acts to produce the Bact complex and in the second, the RNA helicase Prp2 facilitates the formation of the B* complex. This has a functional active site and, following the recruitment of the protein Cwc25, the first step of splicing takes place. In this step, the phosphodiester bond at the 5 splice site is cleaved and, at the same time, the 5 end of the intron becomes linked to the 2 hydroxyl group of an adenosine at the ...
A tripartite motif located in the centre of the 7.5kb exon 26 of apolipoprotein B (apoB) mRNA directs editosome assembly and site-specific cytidine-to-uridine editing at nucleotide 6666. apoB mRNA editing is a post-transcriptional event, occurring primarily at the time exon 26 is spliced or at a time after splicing, but before nuclear export. We show, through reporter RNA constructs, that RNA splice sites suppress editing of precursor RNAs when placed proximal or distal to the editing site. Processed RNAs were edited more efficiently than precursor RNAs. Mutation of both the splice donor and acceptor sites was necessary for RNAs to be edited efficiently. The results suggested that commitment of pre-mRNA to the splicing and/or nuclear-export pathways may play a role in regulating editing-site utilization. The HIV-1 Rev-Rev response element (RRE) interaction was utilized to uncouple the commitment of precursor RNAs to the spliceosome assembly pathway and associated nuclear-export pathway. Under ...
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Lucero Rogel. PREP Program. Lab Group: Alan Zahler. PREP Research: The Zahler lab works to investigate the mechanism of RNA splicing by utilizing C. elegans as a model organism. Accurate splicing of pre-mRNA is a critical step in the gene expression pathway in eukaryotes carried out by a large ribonucleoprotein complex known as the spliceosome. The spliceosome consists of 5 RNAs (U1, U2, U4, U5, and U6) that assemble onto the pre-mRNA by recognizing conserved sequence elements that define the beginning and ends of introns, known as 5 and 3 splice sites. To facilitate the interactions between the pre-mRNA and the 5 RNAs, over 150 proteins are employed to aid in the process. Currently in the lab, I am working on elucidating the role of spliceosomal protein SNRP-27 in 5 splice site selection by uncovering differential interactions that a mutant and wild type form of this factor have with the splicing machinery.. Undergraduate Major: Biochemistry and Molecular Biology. Undergraduate Institution: ...
HI! Tetrahymena RNA self-splices and does it in a very specific series of steps. You find an RNA that self-splices in the same way. It also loops before splicing and the 5 end of the intron to the 3 end of the exon. You were able to figure out the six nucleotide sequence of the 3 end of the exon. Reading 5 to 3, what is the sequence of the base-pairing nucleotides on the intron starting at position #1? 5 TTTCGG 3 (No, this is a DNA sequence.) 5 GGCUUU 3 (No, this is the right sequence but it is in the wrong direction.) 5 GGCTTT 3 (No, this is a DNA sequence.) 5 UUUCGG 3 (That is correct.) G can hydrogen bond with C, and A can hydrogen bond with U. Reading 5 to 3 the complementary sequence is UUUCGG. You think that this base pairing is important for the self-splicing reaction. To prove this theory, you insert two Uracils into the intron and you find that self-splicing decreases dramatically. Other than deleting the 2 Us, how might you restore the self-splicing reaction to this RNA? ...
Fibrillarin is thought to shuttle between the nucleolus and the cytoplasm of complicated Cajal bodies (CBs) and interchromatin granule clusters (IGCs) in oocytes , shared with a COOH-terminal P40 capsid protein encoded with HSV1. The coiled body is a nuclear organelle that contains snRNPs involved in splicing, in both animal and plant cells can undergo…
During pre-mRNA splicing, exons in the primary transcript are precisely connected to generate an mRNA. Intron lariat RNAs are formed as by-products of this process. In addition, some exonic circular RNAs (circRNAs) may also result from exon skipping as by-products. Lariat RNAs and circRNAs are both RNase R resistant RNAs. RNase R is a strong 3 to 5 exoribonuclease, which efficiently degrades linear RNAs, such as mRNAs and rRNAs; therefore, the circular parts of lariat RNAs and the circRNAs can be segregated from eukaryotic total RNAs by their RNase R resistance. Thus, RNase R resistant RNAs could provide unexplored splicing information not available from mRNAs. Analyses of these RNAs identified repeating splicing phenomena, such as re-splicing of mature mRNAs and nested splicing. Moreover, circRNA might function as microRNA sponges. There is an enormous variety of endogenous circRNAs, which are generally synthesized in cells and tissues.
Several predictions come out of these models including the lack of introns in the 3′ UTR and that the average length of exons should be correlated with the window that the proofreading mechanism can operate on. These are discussed in several papers out of Mike Lynchs lab including (Lynch and Connery 2003), (Lynch and Kewalramani, 2003), (Lynch and Richardson, 2002) and recently (Scofield et al, 2007).. Efforts to understand the splicing machinery, particularly in S. cerevisiae have led to the discovery of numerous genes that code for proteins that make up the spliceosome. Some of these include small RNAs as well as protein coding genes. The SR proteins are serine-arginine rich proteins that regulate splicing and are found in almost all eukaryotes including most fungi (even those with few introns, such as S. cerevisiae). SR proteins play a role in splicing and in nuclear export (Masuyama et al, 2004, Sanford et al, 2004) indicating that a coupling of these processes may explain why genes with ...
Isolation and Characterization of Functional Tripartite Group II Introns Using a Tn5-Based Genetic Screen. . Biblioteca virtual para leer y descargar libros, documentos, trabajos y tesis universitarias en PDF. Material universiario, documentación y tareas realizadas por universitarios en nuestra biblioteca. Para descargar gratis y para leer online.
cDNA analysis, involving fragment analysis and cloning, indicated that the p. T560R mutation created a novel 5 splice donor site, which led to a novel transcript with a 32 nucleotide deletion in exon 11 ...
mzef = Bio::Tools::MZEF-,new(-file =, result.mzef); # filehandle: $mzef = Bio::Tools::MZEF-,new( -fh =, \*INPUT ); # to indicate that the sequence was reversed prior to feeding it to MZEF # and that you want to have this reflected in the strand() attribute of # the exons, as well have the coordinates translated to the non-reversed # sequence $mzef = Bio::Tools::MZEF-,new( -file =, result.mzef, -strand =, -1 ); # parse the results # note: this class is-a Bio::Tools::AnalysisResult which implements # Bio::SeqAnalysisParserI, i.e., $genscan-,next_feature() is the same while($gene = $mzef-,next_prediction()) { # $gene is an instance of Bio::Tools::Prediction::Gene # $gene-,exons() returns an array of # Bio::Tools::Prediction::Exon objects # all exons: @exon_arr = $gene-,exons(); # internal exons only @intrl_exons = $gene-,exons(Internal); # note that presently MZEF predicts only internal exons! } # essential if you gave a filename at initialization (otherwise the file # will stay open) ...
Splicing is initiated by a productive interaction between the pre-mRNA and the U1 snRNP, in which a short RNA duplex is established between the 5 splice site ...
With just 20,000 genes but more than 100,000 proteins, human beings have become increasingly unique because of alternative RNA splicing in evolution.
The 5 and 3 ends of each intron are marked with GU and AG dinucleotide sequences; a short tract of poly-pyrimidines (C or T) also occurs near the 3 end ahead of the AG singal. ...
Burge Lab MaxEntScan::score3ss scores 23 mers using different 3ss models To score 5 splice sites go to MaxEntScan::score5ss To build your own MaxEntScan models as described in the paper (below) refer to MaxEntScan::build Reference ...
SRSF5 overexpression lysate, 0.1 mg. Transient overexpression lysate of splicing factor, arginine/serine-rich 5 (SFRS5), transcript variant 2
CDC2-related protein kinase containing an arginine- and serine-rich (SR) domain, a characteristic of the SR protein family of splicing factors, and may be involved in RNA ...
Dengan adanya Permenkes ini, baik perawat maupun masyarakat sebagai pengguna jasa pelayanan terlindungi dari segi hukum. Pada Permenkes ini, disebutkan bahwa perawat yang menjalankan praktik mandiri, harus memiliki SIPP. Sementara untuk mengurus SIPP, perawat tersebut harus melampirkan STR (Surat Tanda Registrasi) yang dapat diperoleh setelah melalui Uji Kompetensi ...
[Ive combined all the previous intron entries together to make it easier to read. However, did not have the time to thoroughly edit, so some parts might seem a little repetitive.] Since I will be discussing introns, let me begin with a few points of clarification. First, I will be focusing on introns found in…
Choroidal vascular remodelling in central serous chorioretinopathy after indocyannie green guided photodynamic therapy with verteporfin: a novel treatment at the primary desease level. Chan, W-M.; Lam, D. S. C.; Lai, T. Y. Y.; Tam, B. S. M.; Liu, D. T. L.; Chan, C. K. M. // British Journal of Ophthalmology;Dec2003, Vol. 87 Issue 12, p1453 Aims: To evaluate the changes in the choroidal vasculature in central serous chorioretinopathy (CSC) after photodynamic therapy (PDT) with verteporfin and to assess its potential role as a treatment option. Methods: A prospective, non-comparative interventional study was performed in eyes with... ...
The adenovirus major late transcription unit (MLTU) is an example of a complex alternatively spliced gene, in which more than 15 different 3 splice sites can be joined to a common 5 splice site. Maturation of the full repertoire of possible mRNAs requires late viral protein synthesis and occurs only at late stages of the infectious cycle (16-24 hpi). We are trying to decipher the mechanisms regulating alternative 3 splice site choice during the infectious cycle. Therefore, we examined the splicing activity of several 3 splice sites from the MLTU in vitro in nuclear extracts prepared from adenovirus infected cells (Ad NE) and from uninfected cells. The results suggest that pre-mRNAs with "weak" 3 splice sites (short, atypical polypyrimidine tracts) are activated and pre-mRNAs with long, prototypical polypyrimidine tracts are repressed in Ad NE. In fact, our data show a reciprocal correlation between the strength of a polypyrimidine tract, defined by its affinity for U2AF65K in vitro, and the ...
Pre-mRNA splicing is executed by the spliceosome, which has eight major functional states each with distinct composition. Five of these eight human spliceosomal complexes, all preceding exon ligation, have been structurally characterized. In this study, we report the cryo-electron microscopy structures of the human post-catalytic spliceosome (P complex) and intron lariat spliceosome (ILS) at average resolutions of 3.0 and 2.9 Å, respectively. In the P complex, the ligated exon remains anchored to loop I of U5 small nuclear RNA, and the 3-splice site is recognized by the junction between the 5-splice site and the branch point sequence. The ATPase/helicase Prp22, along with the ligated exon and eight other proteins, are dissociated in the P-to-ILS transition. Intriguingly, the ILS complex exists in two distinct conformations, one with the ATPase/helicase Prp43 and one without. Comparison of these three late-stage human spliceosomes reveals mechanistic insights into exon release and spliceosome ...
The interactions established at the 5-splice site during spliceosome assembly are likely to be important for both precise recognition of the upstream intron boundary and for positioning this site in the active center of the spliceosome. Definition of the RNA-RNA and the RNA-protein interactions at the 5 splice site would be facilitated by the use of a small substrate amenable to modification during chemical synthesis. We describe a trans-splicing reaction performed in Saccharomyces cerevisiae extracts in which the 5 splice site and the 3 splice site are on separate molecules. The RNA contributing the 5 splice site is only 20 nucleotides long and was synthesized chemically. The trans-splicing reaction is accurate and has the same sequence, ATP, and Mg^2+ requirements as cis-splicing. We also report how deoxy substitutions around the 5-splice site affect trans-splicing efficiency. ...
Dinoflagellates are one of the last major lineages of eukaryotes for which little is known about genome structure and organization. We report here the sequence and gene structure of a clone isolated from a cosmid library which, to our knowledge, represents the largest contiguously sequenced dinoflagellate genomic tandem gene array. These data, combined with information from a large transcriptomic library, allowed a high level of confidence of every base pair call. This degree of confidence is not possible with PCR-based contigs. The sequence contains an intron-rich set of five highly-expressed gene repeats arranged in tandem. One of the tandem repeat gene members contains an intron 26,372 bp long. This study characterizes a splice-site consensus sequence for dinoflagellate introns. Two to nine base pairs around the 3 splice site are repeated by an identical two to nine base pairs around the 5 splice site. The 5 and 3 splice sites are in the same locations within each repeat so that the ...
Production of mRNA in eukaryotic cells involves not only transcription but also various processing reactions such as splicing. Recent experiments have indicated that there are direct physical connections between components of the transcription and processing machinery, supporting previous suggestions that pre-mRNA splicing occurs co-transcriptionally. Here we have used a novel functional approach to demonstrate co-transcriptional regulation of alternative splicing. Exon 3 of the alpha-tropomyosin gene is specifically repressed in smooth muscle cells. By delaying synthesis of an essential downstream inhibitory element, we show that the decision to splice or repress exon 3 occurs during a limited window of opportunity following transcription, indicating that splice site selection proceeds rapidly after transcription.
Alternative pre-mRNA splicing is a powerful mechanism that is exploited by higher eukaryotes to diversify their proteomes, and to differentially regulate the expression, function, and localization of mRNA and proteins. Pre-mRNA splicing is typically regulated by RNA-binding proteins that recognize cis-acting RNA elements, and either activate or repress splicing of adjacent exons in a temporal, and tissue specific, manner. Understanding how RNA-binding proteins control the splicing code is fundamental to understanding organismal development and disease. The SR proteins are a well-conserved class of RNA-binding proteins that have an essential role in the regulation of splice site selection, and have also been implicated as key regulators during other stages of RNA metabolism. The complexity of the RNA targets, and specificity of RNA binding location remains poorly understood for many members of the SR protein family. Here, we present a comprehensive study to elucidate how the SR proteins ...
Recognition of the 3′-splice site is a key step in pre-mRNA splicing and accomplished by a dynamic complex comprising splicing factor 1 (SF1) and the U2 snRNP auxiliary factor 65-kDa subunit (U2AF65). Both proteins mediate protein-protein and protein-RNA interactions for cooperative RNA-binding during spliceosome assembly. Here, we report the solution structure of a novel helix-hairpin domain in the N-terminal region of SF1 (SF1NTD). The nuclear magnetic resonance- and small-angle X-ray scattering-derived structure of a complex of the SF1NTD with the C-terminal U2AF homology motif domain of U2AF65 (U2AF65UHM) reveals that, in addition to the known U2AF65UHM-SF1 interaction, the helix-hairpin domain forms a secondary, hydrophobic interface with U2AF65UHM, which locks the orientation of the two subunits. Mutational analysis shows that the helix hairpin is essential for cooperative formation of the ternary SF1-U2AF65-RNA complex. We further show that tandem serine phosphorylation of a conserved ...
Precursor mRNA (pre-mRNA) splicing is catalyzed by a large ribonucleoprotein complex known as the spliceosome. Numerous studies have indicated that aberrant splicing patterns or mutations in spliceosome components, including the splicing factor 3b subunit 1 (SF3B1), are associated with hallmark cancer phenotypes. This has led to the identification and development of small molecules with spliceosome-modulating activity as potential anticancer agents. Jerantinine A (JA) is a novel indole alkaloid which displays potent anti-proliferative activities against human cancer cell lines by inhibiting tubulin polymerization and inducing G2/M cell cycle arrest. Using a combined pooled-genome wide shRNA library screen and global proteomic profiling, we showed that JA targets the spliceosome by up-regulating SF3B1 and SF3B3 protein in breast cancer cells. Notably, JA induced significant tumor-specific cell death and a significant increase in unspliced pre-mRNAs. In contrast, depletion of endogenous SF3B1 ...
Group II introns are large metallo-ribozymes that use divalent metal ions in folding and catalysis. The 3-terminal domain 6 contains a conserved adenosine whose 2-OH acts as the nucleophile in the first splicing step. In the hierarchy of folding, D6 binds last into the active site. In order to investigate and understand the folding process to the catalytically active intron structure, it is important to know the individual binding affinities of Mg2+ ions to D6. We recently studied the solution structure of a 27 nucleotide long domain 6 (D6-27) from the mitochondrial yeast group II intron Sc.ai5, identifying also five Mg2+ binding sites including the one at the 5-terminal phosphate residues. Mg2+ coordination to the 5-terminal di- and triphosphate groups is strongest (e.g., log KA,TP = 4.55 ± 0.10) and could be evaluated here in detail for the first time. The other four binding sites within D6-27 are filled simultaneously (e.g., log KA,BR = 2.38 ± 0.06) and thus compete for the free Mg2+ ...
Nuclear pre-mRNA splicing takes places in a large RNP complex, the spliceosome, which is assembled in an ordered multistep process. It consists of five small nuclear RNAs (the U1, U2, U4, U5, and U6 snRNAs) and more than 100 proteins, as recent proteomic analyses have determined (15, 16, 39). The spliceosome shows characteristic dynamics during assembly and splicing catalysis. For example, only the U2, U5, and U6 snRNAs participate in the catalytic center of the spliceosome, whereas the U1 and U4 snRNAs play essential roles only during the early assembly stages. After completion of the two-step splicing reaction and the release of mRNA and lariat products, the spliceosome disassembles into its components. Before entering a new cycle, at least some the components presumably must be reactivated. However, very little is known about this recycling phase of the spliceosome cycle.. The U4, U5, and U6 snRNAs enter the prespliceosome in the form of the 25S U4/U6.U5 tri-snRNP but are released from the ...
Pre-mRNA splicing or the removal of introns from precursor messenger RNAs depends on the accurate recognition of intron sequences by the plant splicing machinery. The major components of this machinery are small nuclear ribonucleoprotein protein particles (snRNPs) which consist of snRNAs and snRNP proteins. We have analysed various aspects of intron sequence and structure in relation to splice site selection and splicing efficiency and we have cloned snRNA genes and a gene encoding the snRNP protein, U2B". In the absence of an in vitro splicing system for plants, transient expression in protoplasts and stable plant transform ations have been used to analyse splicing of intron constructs. We aim to address the function of the UsnRNP-specific protein, U2B", via the production of transgenic plants expressing antisense U2B" transcripts and epitope-tagged U2B" protein. In addition, we have cloned genes encoding other proteins which potentially interact with RNA, such as RNA helicases, and strategies ...
cytoplasm, cytoplasmic ribonucleoprotein granule, cytosol, exon-exon junction complex, nucleus, perinuclear region of cytoplasm, polysome, telomeric DNA binding, animal organ regeneration, liver development
... , Authors: Dessen P. Published in: Atlas Genet Cytogenet Oncol Haematol.
... , Authors: Jafar Sharif, Asami Tsuboi, Haruhiko Koseki. Published in: Atlas Genet Cytogenet Oncol Haematol.
RNA editing in the mitochondrion of Trypanosoma brucei extensively alters the adenosine triphosphate synthase (ATPase) subunit 6 precursor messenger RNA (pre-mRNA) by addition of 447 uridines and removal of 28 uridines. In vivo, the guide RNA gA6[14] is thought to specify the deletion of two uridines from the editing site closest to the 3 end. In this study, an in vitro system was developed that accurately removed uridines from this editing site in synthetic ATPase 6 pre-mRNA when gA6[14] and ATP were added. Mutations in both the guide RNA and the pre-mRNA editing site suggest that base-pairing interactions control the number of uridines deleted in vitro. Thus, guide RNAs are required for RNA editing and for the transfer of genetic information to pre-mRNAs. ...
Five small nuclear RNAs (snRNAs) (U1, U2, U4, U5, U6) found in the nucleus of eukaryotic cells mediate the excision of introns from pre-mRNAs. The structure and expression of the snRNA components have been well documented in animal and yeast systems but little information has existed on the structure and expression of the splicesomal snRNAs involved in plant intron excision. To further define the snRNA components involved in intron excision, a clone library has been constructed from anti-m$\sb3$G immunoprecipitated snRNAs expressed in pea seedlings. cDNA clones representing U1, U2, U4, and U5 snRNAs expressed in seedling tissue have been isolated and sequenced. Comparison of the pea snRNA variants with other organisms suggest that functionally important primary sequences are conserved phylogenetically even though the overall sequences have diverged substantially. Structural variations occur in regions required for U1-specific protein binding suggesting that alternate U1 snRNP particles may exist ...
The wide, but scattered distribution of group I introns in nature is a result of two processes; the vertical inheritance of introns with or without losses, and the occasional transfer of introns across species barriers. Reversal of the group I intron