scholarly journals Characterization of SRp46, a Novel Human SR Splicing Factor Encoded by a PR264/SC35 Retropseudogene

1998 ◽  
Vol 18 (8) ◽  
pp. 4924-4934 ◽  
Author(s):  
Johann Soret ◽  
Renata Gattoni ◽  
Cécile Guyon ◽  
Alain Sureau ◽  
Michel Popielarz ◽  
...  
Keyword(s):  
Splicing Factor ◽  
Sr Protein ◽  
Adenovirus E1a ◽  
Cdna Sequences ◽  
Normal Tissues ◽  
Protein Functions ◽  
S100 Extract ◽  
In Vitro Splicing ◽  
First Time

ABSTRACT The highly conserved SR family contains a growing number of phosphoproteins acting as both essential and alternative splicing factors. In this study, we have cloned human genomic and cDNA sequences encoding a novel SR protein designated SRp46. Nucleotide sequence analyses have revealed that the SRp46 gene corresponds to an expressed PR264/SC35 retropseudogene. As a result of mutations and amplifications, the SRp46 protein significantly differs from the PR264/SC35 factor, mainly at the level of its RS domain. Northern and Western blot analyses have established that SRp46 sequences are expressed at different levels in several human cell lines and normal tissues, as well as in simian cells. In contrast, sequences homologous to SRp46 are not present in mice. In vitro splicing studies indicate that the human SRp46 recombinant protein functions as an essential splicing factor in complementing a HeLa cell S100 extract deficient in SR proteins. In addition, complementation analyses performed with β-globin or adenovirus E1A transcripts and different splicing-deficient extracts have revealed that SRp46 does not display the same activity as PR264/SC35. These results demonstrate, for the first time, that an SR splicing factor, which represents a novel member of the SR family, is encoded by a functional retropseudogene.

Splicing efficiency of minor introns in a mouse model of SMA predominantly depends on their branchpoint sequence and can involve the contribution of major spliceosome components.

RNA ◽  
2021 ◽  
pp. rna.078329.120
Author(s):  
Valentin Jacquier ◽  
Manon Prevot ◽  
Thierry Gostan ◽  
Remy Bordonne ◽  
Sofia Benkhelifa-Ziyyat ◽  
...  
Keyword(s):  
Muscular Atrophy ◽  
Differential Splicing ◽  
Molecular Determinants ◽  
In Vitro Splicing ◽  
Splicing Efficiency ◽  
Smn Protein ◽  
Survival Of Motor Neuron ◽  
Branchpoint Sequence ◽  
First Time

Spinal Muscular Atrophy (SMA) is a devastating neurodegenerative disease caused by reduced amounts of the ubiquitously expressed Survival of Motor Neuron (SMN) protein. In agreement with its crucial role in the biogenesis of spliceosomal snRNPs, SMN-deficiency is correlated to numerous splicing alterations in patient cells and various tissues of SMA mouse models. Among the snRNPs whose assembly is impacted by SMN-deficiency, those involved in the minor spliceosome are particularly affected. Importantly, splicing of several, but not all U12-dependent introns has been shown to be affected in different SMA models. Here, we have investigated the molecular determinants of this differential splicing in spinal cords from SMA mice. We show that the branchpoint sequence (BPS) is a key element controlling splicing efficiency of minor introns. Unexpectedly, splicing of several minor introns with suboptimal BPS is not affected in SMA mice. Using in vitro splicing experiments and oligonucleotides targeting minor or major snRNAs, we show for the first time that splicing of these introns involves both the minor and major machineries. Our results strongly suggest that splicing of a subset of minor introns is not affected in SMA mice because components of the major spliceosome compensate for the loss of minor splicing activity.

The zinc finger protein GLI transforms primary cells in cooperation with adenovirus E1A

1991 ◽  
Vol 11 (3) ◽  
pp. 1724-1728 ◽  
Author(s):  
J M Ruppert ◽  
B Vogelstein ◽  
K W Kinzler
Keyword(s):  
Zinc Finger ◽  
Nude Mice ◽  
Zinc Finger Protein ◽  
Primary Cells ◽  
Adenovirus E1a ◽  
In Vitro Transformation ◽  
Finger Protein ◽  
Protein Functions

The GLI gene was previously isolated by virtue of its amplification in human glioblastomas. We have now found that GLI expression can result in the in vitro transformation of both primary and secondary rodent cells. When coexpressed with adenovirus E1A, the GLI protein functions analogously to RAS, resulting in the formation of dense foci of cells which are tumorigenic in nude mice.

scholarly journals Roles for SR Proteins and hnRNP A1 in the Regulation of c-src Exon N1

2003 ◽  
Vol 23 (6) ◽  
pp. 1874-1884 ◽  
Author(s):  
Nanette Rooke ◽  
Vadim Markovtsov ◽  
Esra Cagavi ◽  
Douglas L. Black
Keyword(s):  
Regulatory Element ◽  
Sr Proteins ◽  
Hnrnp A1 ◽  
Sr Protein ◽  
Enhancer Element ◽  
Splicing Regulators ◽  
In Vitro Splicing ◽  
Hnrnp F

ABSTRACT The splicing of the c-src exon N1 is controlled by an intricate combination of positive and negative RNA elements. Most previous work on these sequences focused on intronic elements found upstream and downstream of exon N1. However, it was demonstrated that the 5′ half of the N1 exon itself acts as a splicing enhancer in vivo. Here we examine the function of this regulatory element in vitro. We show that a mutation in this sequence decreases splicing of the N1 exon in vitro. Proteins binding to this element were identified as hnRNP A1, hnRNP H, hnRNP F, and SF2/ASF by site-specific cross-linking and immunoprecipitation. The binding of these proteins to the RNA was eliminated by a mutation in the exonic element. The activities of hnRNP A1 and SF2/ASF on N1 splicing were examined by adding purified protein to in vitro splicing reactions. SF2/ASF and another SR protein, SC35, are both able to stimulate splicing of c-src pre-mRNA. However, splicing activation by SF2/ASF is dependent on the N1 exon enhancer element whereas activation by SC35 is not. In contrast to SF2/ASF and in agreement with other systems, hnRNP A1 repressed c-src splicing in vitro. The negative activity of hnRNP A1 on splicing was compared with that of PTB, a protein previously demonstrated to repress splicing in this system. Both proteins repress exon N1 splicing, and both counteract the enhancing activity of the SR proteins. Removal of the PTB binding sites upstream of N1 prevents PTB-mediated repression but does not affect A1-mediated repression. Thus, hnRNP A1 and PTB use different mechanisms to repress c-src splicing. Our results link the activity of these well-known exonic splicing regulators, SF2/ASF and hnRNP A1, to the splicing of an exon primarily controlled by intronic factors.

scholarly journals Downregulation of long non-coding RNA ZXF1 restricts cell survival by targeting miR-634-GRB2 in lung adenocarcinoma

2020 ◽  
Author(s):  
Xubin Ren ◽  
Nie Xu ◽  
Yunting Zhang ◽  
Tao Wang
Keyword(s):  
Lung Adenocarcinoma ◽  
Luciferase Reporter ◽  
Normal Tissues ◽  
Non Coding Rna ◽  
Enhanced Expression ◽  
Rna Immunoprecipitation ◽  
Underlying Mechanisms ◽  
First Time ◽  
Long Non Coding Rna

Increasing evidence demonstrates that long non-coding RNAs (lncRNAs) play important regulatory roles in mediating initiation and progression of lung adenocarcinoma (LA), which is one of the most lethal in humans. A previous study reported that lncRNAZXF1 was dysregulated in LA and enhanced expression of ZXF1 promoted the invasion and metastasis in LA. However, the effect of ZXF1 on LA progression and its underlying mechanisms were not thoroughly investigated. In our in vitro experiments, qRT-PCR revealed that the expression level of ZXF1 in LA tissues and tumor cells were significantly higher than that in adjacent normal tissues and normal cells. Furthermore, bioinformatics analysis, luciferase reporter assay, western blot and RNA immunoprecipitation (RIP) assay showed that ZXF1 could directly interact with miR-634, which targets GRB2. Therefore, we propose that ZXF1 could function as an oncogene partly by sponging miR-634 and therefore regulating GRB2 expression in LA. Overall, this study demonstrated, for the first time, that the lncRNA ZXF1/miR-634/GRB2 axis plays crucial roles in modulating LA progression. Moreover, lncRNA ZXF1 might potentially improve LA prognosis and serve as a therapeutic target for the treatment of LA.

scholarly journals The 216-nucleotide intron of the E1A pre-mRNA contains a hairpin structure that permits utilization of unusually distant branch acceptors.

1989 ◽  
Vol 9 (11) ◽  
pp. 4852-4861 ◽  
Author(s):  
K Chebli ◽  
R Gattoni ◽  
P Schmitt ◽  
G Hildwein ◽  
J Stevenin
Keyword(s):  
Splice Site ◽  
Site Directed Mutagenesis ◽  
Hairpin Structure ◽  
Cis Acting ◽  
Adenovirus E1a ◽  
Sequence Elements ◽  
In Vitro Splicing ◽  
Hairpin Formation

A recently characterized 216-nucleotide intron-splicing reaction occurs within the adenovirus E1A pre-mRNA through the use of three branch acceptor sites, located at 59, 55, and 51 nucleotides from the 3' splice site. To investigate the role of the cis-acting sequence elements in the selection of such unusually distant branch sites, transcripts differing in sequence downstream of the branch sites were analyzed for in vitro splicing. Initial results suggested that secondary structure could be involved in the use of distant branch sites. The involvement of a hairpin structure, including a nine-G C-base-pair stem, was supported by the results of site-directed mutagenesis analyses. Mutations that destroyed or weakened this hairpin resulted in an inefficient splicing reaction. In contrast, complementary mutation or deletion of two bulges, which involved a restoration or reinforcement of the hairpin, resulted in a reactivation or improvement of the splicing efficiency, respectively. Therefore, we conclude that the hairpin structure shortens the operational distance between the 3' splice site and the branch acceptors and brings the branch sites into the branch-permissive window, 18 to 40 nucleotides upstream of the 3' splice site. Our results confirm the importance of the constraint of distance for the splicing reaction and show that this constraint may be overcome by means of a stable hairpin formation.

ZNF280A Promotes Lung Adenocarcinoma Development through Regulating the Expression of EIF3C

2020 ◽  
Author(s):  
Hongsheng Liu ◽  
Yingzhi Qin ◽  
Na Zhou ◽  
Dongjie Ma ◽  
Yingyi Wang
Keyword(s):  
Lung Cancer ◽  
Lung Adenocarcinoma ◽  
Small Cell Lung ◽  
Normal Tissues ◽  
Tumor Promotor ◽  
And Migration ◽  
First Time ◽  
The Relationship

Abstract Background: Lung cancer is the most commonly diagnosed malignant tumor worldwide. Lung adenocarcinoma (LUAD) is the most common histological subtype in non-small cell lung cancer (NSCLC). The relationship between ZNF280A and LUAD has not been demonstrated and remains unclear. Methods: In this study, it was demonstrated that ZNF280A was upregulated in LUAD tissues compared with the normal tissues. Further investigations indicated that the overexpression/knockdown of ZNF280A could promote/inhibit proliferation, colony formation and migration of LUAD cells, while inhibiting/promoting cell apoptosis. Moreover, knockdown of ZNF280A could also suppress tumorigenicity of LUAD cells in vivo. RNA-sequencing followed by Ingenuity pathway analysis (IPA) was performed for exploring downstream of ZNF280A and identified EIF3C as the potential target. Results: Furthermore, our study revealed that knockdown of EIF3C could inhibit development of LUAD in vitro, and alleviate the ZNF280A overexpression induced promotion of LUAD. Conclusions: In conclusion, our study showed, as the first time, ZNF280A as a tumor promotor for LUAD, whose function was carried out probably through the regulation of EIF3C.

scholarly journals Pre-mRNA Splicing by the EssentialDrosophila Protein B52: Tissue and Target Specificity

2000 ◽  
Vol 20 (1) ◽  
pp. 181-186 ◽  
Author(s):  
Bryan E. Hoffman ◽  
John T. Lis
Keyword(s):  
Mrna Splicing ◽  
Larval Brain ◽  
Sr Protein ◽  
In Vitro System ◽  
Nuclear Extracts ◽  
S100 Extract ◽  
Z Ring ◽  
Splicing Defects ◽  
The Brain

ABSTRACT B52, an essential SR protein of Drosophila melanogaster, stimulates pre-mRNA splicing in splicing-deficient mammalian S100 extracts. Surprisingly, mutant larvae depleted of B52 were found to be capable of splicing at least several pre-mRNAs tested (H. Z. Ring and J. T. Lis, Mol. Cell. Biol. 14:7499–7506, 1994). In a homologous in vitro system, we demonstrated that B52 complements a Drosophila S100 extract to allow splicing of a Drosophila fushi tarazu(ftz) mini-pre-mRNA. Moreover, Kc cell nuclear extracts that were immunodepleted of B52 lost their ability to splice thisftz pre-mRNA. In contrast, splicing of this sameftz pre-mRNA occurred in whole larvae homozygous for theB52 deletion. Other SR protein family members isolated from these larvae could substitute for B52 splicing activity in vitro. We also observed that SR proteins are expressed variably in different larval tissues. B52 is the predominant SR protein in specific tissues, including the brain. Tissues in which B52 is normally the major SR protein, such as larval brain tissue, failed to produce ftzmRNA in the B52 deletion line. These observations support a model in which the lethality of the B52 deletion strain is a consequence of splicing defects in tissues in which B52 is normally the major SR protein.

scholarly journals IL-17 Affects the Progression, Metastasis, and Recurrence of Laryngeal Cancer via the Inhibition of Apoptosis through Activation of the PI3K/AKT/FAS/FASL Pathways

2020 ◽  
Vol 2020 ◽  
pp. 1-14
Author(s):  
Yang Song ◽  
Ming Yang ◽  
Hongjian Zhang ◽  
Yan Sun ◽  
Ye Tao ◽  
...  
Keyword(s):  
Laryngeal Cancer ◽  
Time Course ◽  
Clinical Stage ◽  
Immune Surveillance ◽  
Tumor Promoter ◽  
Interleukin 17 ◽  
Normal Tissues ◽  
First Time

Background. Cytokines play important roles in the development and prognosis of laryngeal cancer (LC). Interleukin-17 (IL-17) from a distinct subset of CD4+ T cells may significantly induce cancer-elicited inflammation to prevent tumor immune surveillance. Methods. The expression levels of IL-17 were examined among 60 patients with LC. Immunofluorescence colocalization experiments were performed to verify the localization of IL-17 and FAS/FASL in Hep-2 and Tu212 cells. The role of IL-17 was determined using siRNA techniques in the LC cell line. Results. In the LC patients, cytokines were dysregulated in LC tissues compared with normal tissues. It was found that IL-17 was overexpressed in a cohort of 60 LC tumors paired with nontumor tissues. Moreover, high IL-17 expression was significantly associated with the advanced T category, the late clinical stage, differentiation, lymph node metastasis, and recurrence. In addition, the time course expression of FAS and FASL was observed after stimulation and treatment with the IL-17 stimulator. Finally, in vitro experiments demonstrated that IL-17 functioned as an oncogene by inhibiting the apoptosis of LC cells via the PI3K/AKT/FAS/FASL pathways. Conclusions. In summary, these findings demonstrated for the first time the role of IL-17 as a tumor promoter and a prometastatic factor in LC and indicated that IL-17 may have an oncogenic role and serve as a potential prognostic biomarker and therapeutic target in LC.

scholarly journals Tumor Proteins D52 and D54 Have Opposite Effects on the Terminal Differentiation of Chondrocytes

2017 ◽  
Vol 2017 ◽  
pp. 1-11
Author(s):  
Chihiro Ito ◽  
Yoshiki Mukudai ◽  
Masakatsu Itose ◽  
Kosuke Kato ◽  
Hiromi Motohashi ◽  
...  
Keyword(s):  
Terminal Differentiation ◽  
Dependent Manner ◽  
Epiphyseal Growth Plate ◽  
Knock Down ◽  
Normal Tissues ◽  
Proliferation And Differentiation ◽  
First Time ◽  
Tumor Proteins

The tumor protein D (TPD) family consists of four members, TPD52, TPD53, TPD54, and TPD55. The physiological roles of these genes in normal tissues, including epidermal and mesenchymal tissues, have rarely been reported. Herein, we examined the expression of TPD52 and TPD54 genes in cartilage in vivo and in vitro and investigated their involvement in the proliferation and differentiation of chondrocytes in vitro. TPD52 and TPD54 were uniformly expressed in articular cartilage and trabecular bone and were scarcely expressed in the epiphyseal growth plate. In MC3T3E-1 cells, the expressions of TPD52 and TPD54 were increased in a differentiation-dependent manner. In contrast, their expressions were decreased in ATDC5 cells. In ATDC5 cells, overexpression of TPD52 decreased alkaline phosphatase (ALPase) activity, while knock-down of TPD52 showed little effect. In contrast, overexpression of TPD54 enhanced ALPase activity, Ca2+ deposition, and the expressions of type X collagen and ALPase genes, while knock-down of TPD54 reduced them. The results revealed that TPD52 inhibits and that TPD54 promotes the terminal differentiation of a chondrocyte cell line. As such, we report for the first time the important roles of TPD52 and TPD54, which work oppositely, in the terminal differentiation of chondrocytes during endochondral ossification.

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