SEARCH · PubMed Health

Results for “RNA Splicing Factors”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 109 records · Page 6Linked to original sources

PubMed2006

A gene similar to the human hyaluronan-mediated motility receptor (RHAMM) gene is upregulated during Porcine Circovirus type 2 infection.

Little is known on the cellular events triggered by the Porcine Circovirus type 2 (PCV2) in Porcine Multisystemic Wasting Syndrome (PMWS). The differential display reverse-transcription PCR (DDRT-PCR) was used to identify cellular target molecules in lymph node tissue that were regulated in PMWS. Comparative profile analysis of a pool of lymph node tissues from PMWS and healthy animals showed that some transcripts were up-regulated in PMWS. Bacterial recombinant clones containing up-regulated transcripts were analyzed by reverse dot blot. Clones showing enhanced hybridization when probed with cDNAs from PMWS animals were sequenced and compared to existing databases. Two of the differentially regulated transcripts displayed homology with human genes such as an RNA splicing factor and hyaluronan-mediated motility receptor (RHAMM). Clones encoding theses genes were subsequently used as probes to analyze their expression pattern in PK15 cells persistently infected with PCV2. Northern blot analyzes indicated that these transcripts were up-regulated in these cells as observed in infected lymph node tissue from PMWS cases. A role for the up-regulation of the RHAMM gene is proposed.

Animals↗

PubMed2000

CROP/Luc7A, a novel serine/arginine-rich nuclear protein, isolated from cisplatin-resistant cell line.

A novel putative SR protein, designated cisplatin resistance-associated overexpressed protein (CROP), has been cloned from cisplatin-resistant cell lines by differential display. The N-half of the deduced amino acid sequence of 432 amino acids of CROP contains cysteine/histidine motifs and leucine zipper-like repeats. The C-half consists mostly of charged and polar amino acids: arginine (58 residues or 25%), glutamate (36 residues or 16%), serine (35 residues or 15%), lysine (30 residues, 13%), and aspartate (20 residues or 9%). The C-half is extremely hydrophilic and comprises domains rich in lysine and glutamate residues, rich in alternating arginine and glutamate residues, and rich in arginine and serine residues. The arginine/serine-rich domain is dominated by a series of 8 amino acid imperfect repetitive motif (consensus sequence, Ser-Arg-Ser-Arg-Asp/Glu-Arg-Arg-Arg), which has been found in RNA splicing factors. The RNase protection assay and Western blotting analysis indicate that the expression of CROP is about 2-3-fold higher in mRNA and protein levels in cisplatin-resistant ACHN/CDDP cells than in host ACHN cells. CROP is the human homologue of yeast Luc7p, which is supposed to be involved in 5'-splice site recognition and is essential for vegetative growth.

Amino Acid Sequence↗

PubMed2004

Nr0b1 and its network partners are expressed early in murine embryos prior to steroidogenic axis organogenesis.

Ahch is an orphan nuclear receptor encoded by Nr0b1 on the murine X chromosome and is the ortholog of human DAX1. Nr0b1/NR0B1 expression at appropriate dosages is required for normal steroidogenic axis development: mutation of the human ortholog, NR0B1, results in adrenal hypoplasia congenita and hypogonadotropic hypogonadism; and duplication or transgenic overexpression in humans or mice, respectively, results in XY phenotypic females, a phenotype known as dosage sensitive sex-reversal. Complete loss of Nr0b1 by targeted deletion has been hypothesized to be lethal in embryonic stem (ES) cells and preliminary evidence suggested that ES cells might express Nr0b1. These investigations examined Nr0b1 expression and its network partners in both cultured ES cells and preimplantation embryos. We cultured ES cells in the absence or presence of differentiation agents and analyzed expression of Nr0b1 and associated network partners by northern blot hybridization and reverse transcriptase-polymerase chain reaction. Nrob1 was highly expressed by totipotent ES cells with reduced expression following induction toward individual germ layer fates. Nr5a1/Sf1, Wt1 and other genes that encode proteins known to interact with Nr0b1 were also expressed. Immunohistochemical analysis of preimplantation embryos for Ahch and key partners confirmed in vivo expression of network components. These findings are consistent with the existence of a potentially functional network of transcription factors, including Ahch, very early in embryonic development. These results validate ES cells as a developmentally dynamic model for mechanistic investigations into this regulatory network early in embryogenesis preceding organogenesis.

Animals↗

PubMed1996

Requirement for SLU7 in yeast pre-mRNA splicing is dictated by the distance between the branchpoint and the 3' splice site.

Yeast pre-mRNA splicing factors SLU7 and PRP16 are required for cleavage of the 3' splice site and exon ligation in vitro. Using natural and model precursor RNAs, we found that SLU7 is dispensable for splicing of RNAs in which the 3' splice site is in close proximity to the branchpoint. SLU7 is only required when the interval between the branchpoint and the 3' splice site is greater than 7 nt. In contrast, PRP16 is essential for splicing of all pre-mRNAs tested. Immunoprecipitation of the products of step 1 by anti-SLU7 antibodies demonstrates that SLU7 is a component of the spliceosome. Recruitment of SLU7 to the spliceosome is greatly enhanced by prior addition of PRP16. PRP16 is liberated from the spliceosome after completion of step 2, whereas SLU7 remains bound to the excised intron and spliced mature RNA until the spliceosome disassembles, in a reaction that requires ATP.

Adenosine Triphosphate↗

PubMed2004

SIPP1, a novel pre-mRNA splicing factor and interactor of protein phosphatase-1.

We have identified a polypeptide that was already known to interact with polyglutamine-tract-binding protein (PQBP)-1/Npw38 as a novel splicing factor and interactor of protein phosphatase-1, hence the name SIPP1 for splicing factor that interacts with PQBP-1 and PP1 (protein phosphotase 1). SIPP1 was inhibitory to PP1, and its inhibitory potency was increased by phosphorylation with protein kinase CK1. Two-hybrid and co-sedimentation analysis revealed that SIPP1 has two distinct PP1-binding domains and that the binding of SIPP1 with PP1 involves a RVXF (Arg-Val-Xaa-Phe) motif, which functions as a PP1-binding sequence in most interactors of PP1. Enhanced-green-fluorescent-protein-tagged SIPP1 was targeted exclusively to the nucleus and was enriched in the nuclear speckles, which represent storage/assembly sites of splicing factors. We have mapped a nuclear localization signal in the N-terminus of SIPP1, while the proline-rich C-terminal domain appeared to be required for its subnuclear targeting to the speckles. Finally, we found that SIPP1 is also a component of the spliceosomes and that a SIPP1-fragment inhibits splicing catalysis by nuclear extracts independent of its ability to interact with PP1.

Amino Acid Motifs↗

PubMed1998

The fission yeast prp10(+) gene involved in pre-mRNA splicing encodes a homologue of highly conserved splicing factor, SAP155.

In the fission yeast Schizosaccharomyces pombe, 14 prp (pre-mRNAprocessing) mutants have been isolated to date. We cloned the prp10(+) gene by complementation of the temperature-sensitive growth of prp10. Five types of transcripts were found that were alternatively spliced with respect to two possible introns located in the 5'-terminal region. Three of them are probably functional and code for putative proteins of approximately 1200 amino acids. Proteins highly homologous to Prp10p are present in other organisms, one of which is a human spliceosome-associated protein SAP155, a subunit of the splicing factor complex SF3. The C-terminal two-thirds of Prp10p is highly conserved among species, and contains consensus repeats for the regulatory subunit A of protein phosphatase PP2A. A gene disruption experiment indicated that the prp10(+) gene is essential for viability in S.pombe. Prp10p tagged with GFP is predominantly localized in the nuclear DNA region. A series of deletions showed that the less conserved N-terminal region of approximately 300 amino acids in Prp10p is dispensable, although the corresponding region was thought to play important roles in the mammalian splicing system.

Cloning, Molecular↗

PubMed2001

[Regulation of elastin synthesis].

Elastin is the main protein of elastic fibers and confers the property of elastic recoil to the tissues such as arteries, lung, elastic cartilage,... Elastin synthesis goes through several steps: gene transcription, alternative splicing of pre-mRNA, mRNA translation, hydroxylation of some proline residues of the newly synthesized protein-tropoelastin-, association of with a 67 kDa chaperone protein, secretion of tropoelastin molecules in the extracellular space, and their deposition on the microfibrillar scaffold which contains fibrillin 1, fibrillin 2, MAGP 1 and MAGP 2,.... After the synthesis of cross-links-lysinonorleucine, desmosine, isodesmosine-, elastin becomes insoluble and elastic. The elastogenic pathway is regulated at many levels. The most recently described regulatory mechanism of elastin synthesis is the control of elastin mRNA stability. Elastogenesis is well controlled during development and aging but remains responsive to external factors such as soluble compounds-cytokines, vitamins, hormones,...- and hemodynamic stress. In order to ensure its function, both quantity and quality of elastin should be and should remain optimal in elastic tissues.

Alternative Splicing↗

PubMed2000

Myotonic dystrophy: the role of the CUG triplet repeats in splicing of a novel DMPK exon and altered cytoplasmic DMPK mRNA isoform ratios.

The mechanism by which (CTG)n expansion in the 3' UTR of the DMPK gene causes myotonic dystrophy (DM) is unknown. We identified four RNA splicing factors--hnRNP C, U2AF (U2 auxiliary factor), PTB (polypyrimidine tract binding protein), and PSF (PTB associated splicing factor)--that bind to two short regions 3' of the (CUG)n, and found a novel 3' DMPK exon resulting in an mRNA lacking the repeats. We propose that the (CUG)n is an essential cis acting element for this splicing event. In contrast to (CUG)n containing mRNAs, the novel isoform is not retained in the nucleus in DM cells, resulting in imbalances in relative levels of cytoplasmic DMPK mRNA isoforms and a new dominant effect of the mutation on DMPK.

3' Untranslated Regions↗

PubMed2005

The Isy1p component of the NineTeen complex interacts with the ATPase Prp16p to regulate the fidelity of pre-mRNA splicing.

Prp16p is a DEAH-box ATPase that transiently associates with the spliceosome to promote the structural transition required for the second chemical step. Yeast strains carrying the cold-sensitive allele prp16-302 stall the release of Prp16p at low temperatures, yet splice precursors with aberrant branchpoints at increased frequency. To identify new factors involved in the regulation of splicing fidelity, we sought suppressors of the prp16-302 growth phenotype. Deletion of the nonessential ISY1 gene (1) improves growth of prp16-302 strains, (2) alleviates stalling, and (3) restores fidelity of branchpoint usage to wild-type levels. Isy1p is a subunit of the NineTeen Complex containing Prp19p, an essential E3 ubiquitin ligase homolog required for splicing. Notably, Deltaisy1 PRP16 strains display reduced fidelity of 3'-splice site selection. Consistent with a recent two-state model of the spliceosome, our genetic and biochemical results suggest that Isy1p acts together with U6 snRNA to promote a spliceosomal conformation favorable for first-step chemistry. We propose that deletion of ISY1 favors the premature release of Prp16p, thus promoting second-step chemistry of precursors with inappropriate 3'-splice sites.

Adenosine Triphosphatases↗

PubMed2000

Revised genomic structure of the human MAGP1 gene and identification of alternate transcripts in human and mouse tissues.

The human MAGP1 (or MFAP2) and mouse Magp1 genes code for the microfibril-associated glycoprotein-1 (MAGP-1), an extracellular matrix protein of microfibrillar structures. We report a revised 5' genomic structure including the use of a single transcription start site that gives rise to a 32-bp 5' exon spanning a segment of the previously described exon B. No evidence of heterogeneous 5' ends from the use of alternative promoters was found in human tissues and cell lines. We located the genetic marker D1S170 to a position 3 kb downstream of the polyadenylation site. Large-scale comparison of the human and mouse genes revealed conservation of sequence outside the coding exons. Although the 5' flanking regions were found to be divergent certain cis-elements for transcription factors are conserved, including Sp1, AP-2, AP-4, NF-kappaB, and c-ETS motifs. We identified a total of five splice variants in addition to the canonical MAGP1A/Magp1A form. These transcripts are species-specific and are generated by different processing mechanisms. The alternate forms MAGP1A', MAGP1B, and MAGP1C are expressed in human tissues; and the two variants Magp1A" and Magp1D were found only in mouse. The alternatively spliced forms show restricted patterns of expression relative to the canonical isoform.

5' Untranslated Regions↗

PubMed2005

The Pso4 mRNA splicing and DNA repair complex interacts with WRN for processing of DNA interstrand cross-links.

DNA interstrand cross-links (ICLs) are perhaps the most formidable lesion encountered by the cellular DNA repair machinery, and the elucidation of the process by which they are removed in eukaryotic cells has proved a daunting task. In particular, the early stages of adduct recognition and uncoupling of the cross-link have remained elusive principally because genetic studies have not been highly revealing. We have developed a biochemical assay in which processing of a DNA substrate containing a site-specific psoralen ICL can be monitored in vitro. Using this assay we have shown previously that the mismatch repair factor MutSbeta, the nucleotide excision repair heterodimer Ercc1-Xpf, and the replication proteins RPA and PCNA are involved in an early stage of psoralen ICL processing. Here, we report the identification of two additional factors required in the ICL repair process, a previously characterized pre-mRNA splicing complex composed of Pso4/Prp19, Cdc5L, Plrg1, and Spf27 (Pso4 complex), and WRN the protein deficient in Werner syndrome. Analysis of the WRN protein indicates that its DNA helicase function, but not its exonuclease activity, is required for ICL processing in vitro. In addition, we show that WRN and the Pso4 complex interact through a direct physical association between WRN and Cdc5L. A putative model for uncoupling of ICLs in mammalian cells is presented.

Carrier Proteins↗

PubMed1994

Conservation of regulated alternative splicing and identification of functional domains in vertebrate homologs to the Drosophila splicing regulator, suppressor-of-white-apricot.

Although several splicing regulatory proteins have been identified in Drosophila through characterization of various genetic mutations, including sex-lethal, transformer, transformer-2, suppressor-of-white-apricot (su(wa)), and possibly suppressor-of-sable, none of these have been identified in vertebrates. We describe the cloning and characterization of human (HsSWAP) and mouse (MmSWAP) homologs of the su(wa) gene. Comparison of the Drosophila and mammalian proteins reveals five highly homologous regions, including an arginine/serine-rich domain and two repeated modules that are homologous to regions in the constitutive splicing factor, SPP91/PRP21. These modules thus define a new motif likely important in the regulatory and constitutive splicing functions of these proteins. The Drosophila su(wa) gene autoregulates its expression by control of splicing of its first two introns. Comparison of mammalian and Drosophila SWAP mRNAs revealed that the splice junctions of these regulated introns are precisely conserved, showing definitively that these genes are ancestrally related. Moreover, mammalian SWAP mRNAs are also alternatively spliced at the same splice sites, showing that mammalian SWAP expression is regulated (presumably autogenously) by control of splicing of these two introns. These several structural features therefore strongly suggest that the mammalian SWAP gene functions as a vertebrate alternative splicing regulator.

Alternative Splicing↗

PubMed2000

Functional analyses of interacting factors involved in both pre-mRNA splicing and cell cycle progression in Saccharomyces cerevisiae.

Through a genetic screen to search for factors that interact with Prp17/Cdc40p, a protein involved in both cell cycle progression and pre-mRNA splicing, we identify three novel factors, which we call Syf1p, Syf2p, and Syf3 (SYnthetic lethal with cdc Forty). Here we present evidence that all three proteins are spliceosome associated, that they associate weakly or transiently with U6 and U5 snRNAs, and that Syf1p and Syf3p (also known as Clf1p) are required for pre-mRNA splicing. In addition we show that depletion of Syf1p or Syf3p results in cell cycle arrest at the G2/M transition. Thus, like Prp17/Cdc40p, Syf1p and Syf3p are involved in two distinct cellular processes. We discuss the likelihood that Syf1p, Syf2p, and Syf3p are components of a protein complex that assembles into spliceosomes and also regulates cell cycle progression.

Base Sequence↗

PubMed2005

Human splicing factor SF3a, but not SF1, is essential for pre-mRNA splicing in vivo.

The three subunits of human splicing factor SF3a are essential for the formation of the functional 17S U2 snRNP and prespliceosome assembly in vitro. RNAi-mediated depletion indicates that each subunit is essential for viability of human cells. Knockdown of single subunits results in a general block in splicing strongly suggesting that SF3a is a constitutive splicing factor in vivo. In contrast, splicing of several endogenous and reporter pre-mRNAs is not affected after knockdown of SF1, which functions at the onset of spliceosome assembly in vitro and is essential for cell viability. Thus, SF1 may only be required for the splicing of a subset of pre-mRNAs. We also observe a reorganization of U2 snRNP components in SF3a-depleted cells, where U2 snRNA and U2-B'' are significantly reduced in nuclear speckles and the nucleoplasm, but still present in Cajal bodies. Together with the observation that the 17S U2 snRNP cannot be detected in extracts from SF3a-depleted cells, our results provide further evidence for a function of Cajal bodies in U2 snRNP biogenesis.

Blotting, Northern↗

PubMed1995

Prp16p, Slu7p, and Prp8p interact with the 3' splice site in two distinct stages during the second catalytic step of pre-mRNA splicing.

For the second catalytic step of pre-mRNA splicing to occur, a 3' splice site must be selected and juxtaposed with the 5' exon. Four proteins, Prp16p, Slu7p, Prp17p, Prp18p, and an integral spliceosomal protein, Prp8p, are known to be required for the second catalytic step. prp8-101, an allele of PRP8 defective in 3' splice site recognition, exhibits specific genetic interactions with mutant alleles of the other second step splicing factors. The prp8-101 mutation also results in decreased crosslinking of Prp8p to the 3' splice site. To determine the role of the step-two-specific proteins in 3' splice site recognition and in binding of Prp8p to the 3' splice site, we performed crosslinking studies in mutant and immunodepleted extracts. Our results suggest an ordered pathway in which, after the first catalytic step, Prp16p crosslinks strongly to the 3' splice site and Prp8p and Slu7p crosslink weakly. ATP hydrolysis by Prp16p affects a conformational change that reduces the crosslinking of Prp16p with the 3' splice site and allows stronger crosslinking of Prp8p and Slu7p. Thus, the 3' splice site appears to be recognized in two stages during the second step of splicing. Strong 3' splice site crosslinking of Prp8p and Slu7p also requires the functions of Prp17p and Prp18p. Therefore, Prp8p and Slu7p interact with the 3' splice site at the latest stage of splicing prior to the second catalytic step that can currently be defined, and may be at the active site.

Adenosine Triphosphatases↗

PubMed2002

Mutations in genes of Saccharomyces cerevisiae encoding pre-mRNA splicing factors cause cell cycle arrest through activation of the spindle checkpoint.

Previous work has identified a group of genes whose products play important roles in two seemingly unrelated processes: cell cycle progression and splicing. The products of these genes show a network of physical and genetic interactions suggestive of the existence of a protein complex, the cell cycle and splicing complex (CSC). Here we analyze the genetic interactions between ISY1, SYF2 and NTC20, three non-essential components of the CSC. We show that mutations in ISY1 cause lethality in the absence of Ntc20p, and that the double mutant isy1Delta syf2Delta shows a temperature-dependent cell cycle arrest. This arrest is due to lower levels of alpha-tubulin, a protein encoded by TUB1 and TUB3, two intron-containing genes. We show that the low levels of alpha-tubulin in isy1Delta syf2Delta trigger activation of the spindle checkpoint, causing cell cycle arrest. Thus, our results have uncovered an unexpected role for pre-mRNA splicing in the maintenance of the fidelity of chromosome transmission during cell division.

Biological Transport↗

PubMed2004

Exonic splicing enhancers in fission yeast: functional conservation demonstrates an early evolutionary origin.

Discrete sequence elements known as exonic splicing enhancers (ESEs) have been shown to influence both the efficiency of splicing and the profile of mature mRNAs in multicellular eukaryotes. While the existence of ESEs has not been demonstrated previously in unicellular eukaryotes, the factors known to recognize these elements and mediate their communication with the core splicing machinery are conserved and essential in the fission yeast Schizosaccharomyces pombe. Here, we provide evidence that ESE function is conserved through evolution by demonstrating that three exonic splicing enhancers derived from vertebrates (chicken ASLV, mouse IgM, and human cTNT) promote splicing of two distinct S. pombe pre-messenger RNAs (pre-mRNAs). Second, as in extracts from mammalian cells, ESE function in S. pombe is compromised by mutations and increased distance from the 3'-splice site. Third, three-hybrid analyses indicate that the essential SR (serine/arginine-rich) protein Srp2p, but not the dispensable Srp1p, binds specifically to both native and heterologous purine-rich elements; thus, Srp2p is the likely mediator of ESE function in fission yeast. Finally, we have identified five natural purine-rich elements from S. pombe that promote splicing of our reporter pre-mRNAs. Taken together, these results provide strong evidence that the genesis of ESE-mediated splicing occurred early in eukaryotic evolution.

Animals↗

PubMed2005

Nucleocytoplasmic shuttling of the splicing factor SIPP1.

SIPP1 (splicing factor that interacts with PQBP1 and PP1) is a widely expressed protein of 70 kDa that has been implicated in pre-mRNA splicing. It interacts with protein Ser/Thr phosphatase-1 (PP1) and with the polyglutamine-tract-binding protein 1 (PQBP1), which contributes to the pathogenesis of X-linked mental retardation and neurodegenerative diseases caused by polyglutamine tract expansions. We show here that SIPP1 is a nucleocytoplasmic shuttling protein. Under basal circumstances SIPP1 was largely nuclear, but it accumulated in the cytoplasm following UV- or X-radiation. Nuclear import was mediated by two nuclear localization signals. In addition, SIPP1 could be piggy-back transported to the nucleus with its ligand PQBP1. In the nucleus SIPP1 and PQBP1 formed inclusion bodies similar to those detected in polyglutamine diseases. SIPP1 did not function as a nuclear targeting subunit of PP1 but re-localized nuclear PP1 to storage sites for splicing factors. The C-terminal residues of SIPP1, which do not conform to a classic nuclear export signal, were required for its nuclear export via the CMR-1 pathway. Finally, SIPP1 activated pre-mRNA splicing in intact cells, and the extent of splicing activation correlated with the nuclear concentration of SIPP1. We conclude that SIPP1 is a positive regulator of pre-mRNA splicing that is regulated by nucleocytoplasmic shuttling. These findings also have potential implications for a better understanding of the pathogenesis of X-linked mental retardation and polyglutamine-linked neurodegenerative disorders.

Active Transport, Cell Nucleus↗