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Splicing Factor (splicing + factor)
Selected AbstractsMyosin Va phosphorylated on Ser1650 is found in nuclear speckles and redistributes to nucleoli upon inhibition of transcriptionCYTOSKELETON, Issue 6 2008Maria Cristina S. Pranchevicius Abstract Nuclear actin and nuclear myosins have been implicated in the regulation of gene expression in vertebrate cells. Myosin V is a class of actin-based motor proteins involved in cytoplasmic vesicle transport and anchorage, spindle-pole alignment and mRNA translocation. In this study, myosin-Va, phosphorylated on a conserved serine in the tail domain (phospho-ser1650 MVa), was localized to subnuclear compartments. A monoclonal antibody, 9E6, raised against a peptide corresponding to phosphoserine1650 and flanking regions of the murine myosin Va sequence, was immunoreactive to myosin Va heavy chain in cellular and nuclear extracts of HeLa cells, PC12 cells and B16-F10 melanocytes. Immunofluorescence microscopy with this antibody revealed discrete irregular spots within the nucleoplasm that colocalized with SC35, a splicing factor that earmarks nuclear speckles. Phospho-ser1650 MVa was not detected in other nuclear compartments, such as condensed chromatin, Cajal bodies, gems and perinucleolar caps. Although nucleoli also were not labeled by 9E6 under normal conditions, inhibition of transcription in HeLa cells by actinomycin D caused the redistribution of phospho-ser1650 MVa to nucleoli, as well as separating a fraction of phospho-ser1650 MVa from SC35 into near-neighboring particles. These observations indicate a novel role for myosin Va in nuclear compartmentalization and offer a new lead towards the understanding of actomyosin-based gene regulation. Cell Motil. Cytoskeleton 2008. © 2008 Wiley-Liss, Inc. [source] The t(1;9)(p34;q34) and t(8;12)(p11;q15) fuse pre-mRNA processing proteins SFPQ (PSF) and CPSF6 to ABL and FGFR1GENES, CHROMOSOMES AND CANCER, Issue 5 2008Claire Hidalgo-Curtis We have investigated two patients with acquired chromosomal rearrangements, a male presenting with a t(1;9)(p34;q34) and B cell progenitor acute lymphoid leukemia and a female presenting with a t(8;12)(p11;q15) and the 8p11 myeloproliferative syndrome. We determined that the t(1;9) fused ABL to SFPQ (also known as PSF), a gene mapping to 1p34 that encodes a polypyrimidine tract-binding protein-associated splicing factor. The t(8;12) fused CPSF6, a cleavage and polyadenylation specificity factor, to FGFR1. The fusions were confirmed by amplification of the genomic breakpoints and RT-PCR. The predicted oncogenic products of these fusions, SFPQ-ABL and CPSF6-FGFR1, are in-frame and encode the N-terminal domain of the partner protein and the entire tyrosine kinase domain and C-terminal sequences of ABL and FGFR1. SFPQ interacts with two FGFR1 fusion partners, ZNF198 and CPSF6, that are functionally related to the recurrent PDGFR, partner FIP1L1. Our findings thus identify a group of proteins that are important for pre-mRNA processing as fusion partners for tyrosine kinases in hematological malignancies. © 2008 Wiley-Liss, Inc. [source] NSSR1 is regulated in testes development and cryptorchidism and promotes the exon 5-included splicing of CREB transcriptsMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 11 2007Ping-Jie Xiao Abstract Neural salient serine/arginine rich protein 1 (NSSR1, alternatively SRp38) is a newly identified splicing factor that is highly expressed in neural and reproductive tissues. We showed that the expression of testicular NSSR1 increased significantly during mouse testes development. NSSR1 was mainly expressed in germ cells, but barely detected in Sertoli cells. Testicular NSSR1 was mostly phosphorylated and cytosolic in germ cells. In comparison, pituitary NSSR1 was mostly dephosphorylated and nuclear. In the cryptorchid testes, the dephosphorylated NSSR1 was significantly increased. RT-PCR analysis demonstrated that the alternative splicing of CREB and CREM genes was altered in the cryptorchid testes. In addition, CREB transcripts were associated with NSSR1 either in testes tissues or cultured GC-1 cells. Moreover, the studies with NSSR1 over-expression or silence demonstrated that NSSR1 promoted the exon 5 inclusion of CREB, indicating that NSSR1 is a new factor that regulates the alternative exon 5 inclusion of CREB transcripts. The findings for the first time provide the evidence indicating the potential importance of NSSR1 in testes development, spermatogenesis and cryptorchidism. Mol. Reprod. Dev. 74: 1363,1372, 2007. © 2007 Wiley-Liss, Inc. [source] An in vitro enzymatic assay coupled to proteomics analysis reveals a new DNA processing activity for Ewing sarcoma and TAF(II)68 proteinsPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 22 2006Olivier Guipaud Abstract Based on structural and functional similarities, translocated in liposarcoma/fusion (TLS/FUS) protein, Ewing sarcoma,(EWS) protein and human TATA binding protein-associated factor (hTAF(II)68) have been grouped in the TLS-EWS-TAF(II)68 (TET) protein family. Translocations involving their genes lead to sarcomas. Polypyrimidine tract-binding protein-associated splicing factor,(PSF), although not grouped in this family, presents structural and functional similarities with TET proteins and is involved in translocation leading to carcinoma. Beside their role in RNA metabolism, the precise cellular functions of these multifunctional proteins are not yet fully elucidated. We previously showed that both TLS/FUS and PSF display activities able to pair homologous DNA on membrane in an in,vitro assay. In the present study, we address the question whether EWS and hTAF(II)68 also display pairing on membrane activities, and to a larger extent whether other proteins also exhibit such activity. We applied the pairing on membrane assay to 2-DE coupled to MS analysis for a global screening of DNA pairing on membrane activities. In addition to TLS/FUS and PSF, this test allowed us to identify EWS and hTAF(II)68, but no other proteins, indicating a feature specific to a protein family whose members share extensive structural similarities. This common activity suggests a role for TET proteins and PSF in genome plasticity control. [source] Expression of Arabidopsis SR-like splicing proteins confers salt tolerance to yeast and transgenic plantsTHE PLANT JOURNAL, Issue 5 2002Javier Forment Summary Searching for novel targets of salt toxicity in eukaryotic cells, we have screened an Arabidopsis thaliana cDNA library to isolate genes conferring increased tolerance to salt stress when expressed in the yeast Saccharomyces cerevisiae. Here we show that expression of the ,alternating arginine-rich' (or RS) domains of two different SR-like, putative splicing proteins from Arabidopsis allows yeast cells to tolerate higher lithium and sodium concentrations. Protection against salt stress appears to require the in vivo phosphorylation of these plant polypeptides, since the yeast SR protein kinase Sky1p, which was able to phosphorylate in vitro at least one of them, also proved to be essential for the observed salt tolerance phenotype. In addition, a clone encoding the U1A protein, a previously characterised Arabidopsis splicing factor, was also isolated in the screening. No significant decrease in the intracellular concentration of lithium was observed in yeast cells incubated in the presence of LiCl upon expression of any of the Arabidopsis proteins, suggesting that their effects are not mediated by the stimulation of ion transport. In support of the general significance of these data, we also show that the expression of the RS domain of one of the SR-like proteins in transgenic Arabidopsis plants increases their tolerance to LiCl and NaCl. These results point to an important role of pre-mRNA splicing and SR-like proteins in the salt tolerance of eukaryotic cells, offering a novel route to improve this important trait in crop plants. [source] Structure of Mycobacterium tuberculosis Rv2714, a representative of a duplicated gene family in ActinobacteriaACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 10 2009Martin Graña The gene Rv2714 from Mycobacterium tuberculosis, which codes for a hypothetical protein of unknown function, is a representative member of a gene family that is largely confined to the order Actinomycetales of Actinobacteria. Sequence analysis indicates the presence of two paralogous genes in most mycobacterial genomes and suggests that gene duplication was an ancient event in bacterial evolution. The crystal structure of Rv2714 has been determined at 2.6,Å resolution, revealing a trimer in which the topology of the protomer core is similar to that observed in a functionally diverse set of enzymes, including purine nucleoside phosphorylases, some carboxypeptidases, bacterial peptidyl-tRNA hydrolases and even the plastidic form of an intron splicing factor. However, some structural elements, such as a ,-hairpin insertion involved in protein oligomerization and a C-terminal ,-helical domain that serves as a lid to the putative substrate-binding (or ligand-binding) site, are only found in Rv2714 bacterial homologues and represent specific signatures of this protein family. [source] PRP-17 and the pre-mRNA splicing pathway are preferentially required for the proliferation versus meiotic development decision and germline sex determination in Caenorhabditis elegansDEVELOPMENTAL DYNAMICS, Issue 5 2010Jessica Amrozowicz Kerins Abstract In C. elegans, the decision between germline stem cell proliferation and entry into meiosis is controlled by GLP-1 Notch signaling, which promotes proliferation through repression of the redundant GLD-1 and GLD-2 pathways that direct meiotic entry. We identify prp-17 as another gene functioning downstream of GLP-1 signaling that promotes meiotic entry, largely by acting on the GLD-1 pathway, and that also functions in female germline sex determination. PRP-17 is orthologous to the yeast and human pre-mRNA splicing factor PRP17/CDC40 and can rescue the temperature-sensitive lethality of yeast PRP17. This link to splicing led to an RNAi screen of predicted C. elegans splicing factors in sensitized genetic backgrounds. We found that many genes throughout the splicing cascade function in the proliferation/meiotic entry decision and germline sex determination indicating that splicing per se, rather than a novel function of a subset of splicing factors, is necessary for these processes. Developmental Dynamics 239:1555,1572, 2010. © 2010 Wiley-Liss, Inc. [source] Combination of Clk family kinase and SRp75 modulates alternative splicing of Adenovirus E1AGENES TO CELLS, Issue 3 2008Jun-ichiro Yomoda SR proteins are non-snRNP splicing factors harbouring a domain rich in Arg-Ser repeats, which are extensively phosphorylated by several kinases. We performed a comparative study of different SR kinases, including SRPK, Clk, PRP4 and DYRK, and found that only Clks efficiently altered 5, splice site selection of Adenovirus E1A. The phosphorylation state of SR proteins was examined using a phospho-SR specific antibody mAb1H4 and a 75 kDa protein was most evidently hyperphosphorylated by Clks. Administration of TG003, a specific inhibitor for the Clk family members, specifically and rapidly induced dephosphorylation of 75 kDa SR protein. Imaging with mRFP-SRp75 in living cells revealed that its nuclear distribution was rapidly altered upon inhibition of the Clk activity by TG003. Co-transfection experiments demonstrated that HA-tagged SRp75 was hyperphosphorylated by Clk family members, but not by other SR kinases. These results indicate that Clks specifically hyperphosphorylate SRp75. Furthermore, SRp75 over-expression promoted the selection of 12S 5, splice site in E1A pre-mRNA, which is stimulated by co-expression of Clks. These results suggest that the specific combination of SR protein and SR kinase plays a distinct role in alternative splicing through dynamic balance of phosphorylation. [source] A T3 allele in the CFTR gene exacerbates exon 9 skipping in vas deferens and epididymal cell lines and is associated with Congenital Bilateral Absence of Vas Deferens (CBAVD),HUMAN MUTATION, Issue 1 2005Antoine Disset Abstract The different alleles at the (TG)m(T)n polymorphic loci at the 3, end of the human CFTR intron 8 determine the efficiency by which exon 9 is spliced. We identified a novel TG12T3 allele in a congenital bilateral absence of vas deferens (CBAVD) patient who carries a [TG11T7; p.Phe508Cys; p.Met470Val] haplotype on the other chromosome. To better understand the complex regulation of exon 9 splicing, we analyzed the levels of correctly spliced CFTR transcripts in six CFTR-expressing epithelial cell lines derived from lung, colon, testis, vas deferens, and epididymis transiently transfected with four CFTR minigenes (pTG11T7, pTG12T7, pTG12T5, and pTG12T3). In this work, we show that a decrease in the Ts at the polymorphic locus in a TG12 background determines a cell-type dependent reduction in exon 9+ transcripts that is not related to the basal splicing efficiency in the cell line. These data emphasize the role of the T5 allele in CBAVD and identify the T3 allele as a severe cystic fibrosis (CF) disease-causing mutation. Finally, UV cross-linking experiments demonstrated that tissue-specific trans -acting splicing factors do not contribute to the different patterns of exon 9 splicing found between the cell lines. However, we observed that lower numbers of Ts can alter the binding of TDP-43 (TDP43 or TARDBP) to its specific target ug12 in a tissue-specific manner. Our results support the idea that the ratio of general splicing factors plays a role in the tissue variability of exon 9 alternative splicing. Hum Mutat 25:72,81, 2005. © 2004 Wiley-Liss, Inc. [source] cAMP-induced differentiation of human neuronal progenitor cells is mediated by nuclear fibroblast growth factor receptor-1 (FGFR1)JOURNAL OF NEUROCHEMISTRY, Issue 6 2003E. K. Stachowiak Abstract Activation of cAMP signaling pathway and its transcriptional factor cyclic AMP response element binding protein (CREB) and coactivator are key determinants of neuronal differentiation and plasticity. We show that nuclear fibroblast growth factor receptor-1 (FGFR1) mediates cAMP-induced neuronal differentiation and regulates CREB and CREB binding protein (CBP) function in ,-internexin-expressing human neuronal progenitor cells (HNPC). In proliferating HNPC, FGFR1 was associated with the cytoplasm and plasma membrane. Treatment with dB-cAMP induced nuclear accumulation of FGFR1 and caused neuronal differentiation, accompanied by outgrowth of neurites expressing MAP2 and neuron-specific neurofilament-L protein and enolase. HNPC transfected with nuclear/cytoplasmic FGFR1 or non-membrane FGFR1(SP-/NLS), engineered to accumulate exclusively in the cell nucleus, underwent neuronal differentiation in the absence of cAMP stimulation. In contrast, FGFR1/R4, with highly hydrophobic transmembrane domain of FGFR4, was membrane associated, did not enter the nucleus and failed to induce neuronal differentiation. Transfection of tyrosine kinase-deleted dominant negative receptor mutants, cytoplasmic/nuclear FGFR1(TK-) or nuclear FGFR1(SP-/NLS)(TK-), prevented cAMP-induced neurite outgrowth. Nuclear FGFR1 localized in speckle-like domains rich in phosphorylated histone 3 and splicing factors, regions known for active RNA transcription and processing, and activated the neurofilament-L gene promoter. FGFR1(SP-/NLS) transactivated CRE, up-regulated phosphorylation and transcriptional activity of CREB and stimulated the activity of CBP several-fold. Thus, cAMP-induced nuclear accumulation of FGFR1 provides a signal that triggers molecular events leading to neuronal differentiation. [source] | |||||||||||||