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At least 523 records · Page 29Linked to original sources
PubMed1997

Tissue-specific alternative RNA splicing of rat vesicle-associated membrane protein-1 (VAMP-1).

The vesicle-associated membrane protein (VAMP) family is essential to vesicle-mediated protein transport. Three mammalian isoforms, VAMP-1, VAMP-2, and cellubrevin, play a role in protein transport to the plasma membrane. In this study, we describe a new rat VAMP-1 isoform produced by alternative pre-mRNA splicing. Only one VAMP-1 isoform dominates in each tissue. Analysis of the nucleotide sequence for the newly discovered isoform, VAMP-1b, reveals that its expression is determined by whether an intron is retained or removed. The predicted amino acid sequences for the VAMP-1 isoforms differ at the carboxy-terminal end of the protein. A similar process has been described for VAMPs in Drosophila melanogaster and suggests a conserved function for the carboxy-terminal domain that can be modulated.

Alternative Splicing↗
PubMed2001

Human angiotensin II type 1 receptor isoforms encoded by messenger RNA splice variants are functionally distinct.

Human tissues that express the angiotensin II (Ang II) type 1 receptor (hAT(1)R) can synthesize four distinct alternatively spliced hAT(1)R mRNA transcripts. In this study, we show that the relative abundance of these mRNA transcripts varies widely in human tissues, suggesting that each splice variant is functionally distinct. Here we demonstrate, for the first time, that the hAT(1)R-B mRNA splice variant encodes a novel long hAT(1)R isoform in vivo that has significantly diminished affinity for Ang II (i.e. >3-fold) when compared with the short hAT(1)R isoform (encoded by hAT(1)R-A mRNA splice variant). This reduced agonist affinity caused a significant shift to the right in the dose-response curve for Ang II-induced inositol trisphosphate production and Ca(2+) mobilization of the long hAT(1)R when compared with that of the short hAT(1)R. The functional differences between these isoforms allows Ang II responsiveness to be fine-tuned by regulating the relative abundance of the long and short hAT(1)R isoform expressed in a given human tissue.

Adrenal Cortex Neoplasms↗
PubMed1994

The calcitonin exon and its flanking intronic sequences are sufficient for the regulation of human calcitonin/calcitonin gene-related peptide alternative RNA splicing.

The primary transcript of the calcitonin (CT)/calcitonin gene-related peptide (CGRP) is alternatively spliced in a cell-specific fashion to produce CT in thyroid C cells and CGRP in neuronal cells. The key step in this regulatory process is the recognition and inclusion of exon 4 to produce CT mRNA or nonrecognition and exclusion of exon 4 to produce CGRP mRNA. To determine whether inclusion/exclusion of CT exon is regulated independently of its position, we created a series of minigene constructs containing decreasing amounts of CT gene sequence. A human glioblastoma cell line, T98G, was tested and used as a CT exon exclusion cell line, while HeLa cells were used as a CT exon inclusion cell line. CT exon inclusion/exclusion was regulated when either the relative position of exon 4 within the CT gene was changed or when the exon with flanking sequence was inserted into a completely heterologous gene. Our results demonstrate that CT exon functions as a unit in a position-independent fashion in regulating its own inclusion/exclusion. We believe that the heterologous fusion gene containing only exon 4 and part of its flanking intron sequences will be useful for further defining the sequence elements involved in the regulation of CT/CGRP splicing.

Alternative Splicing↗
PubMed1995

Alternative RNA splicing of the hyaluronic acid receptor CD44 in the normal human brain and in brain tumors.

The cell-surface receptor for hyaluronic acid, CD44, is expressed by both normal and malignant cells. Numerous CD44 isoforms have recently been identified that are derived by alternative ribonucleic acid splicing. The expression of some CD44 isoforms has been shown to be involved in tumor progression and metastatic spread in a rat carcinoma model and in human carcinomas. In the present study, CD44 isoform expression was evaluated by reverse transcriptase-polymerase chain reaction (PCR) analysis in frozen sections derived from three samples of normal brain tissue and from 40 brain tumors, including samples of glioblastoma multiforme, anaplastic astrocytoma, low-grade astrocytoma, cerebral primitive neuroectodermal tumor, medulloblastoma, metastatic colon carcinoma, and metastatic melanoma. Normal brain tissue adjacent to the tumors was also examined in 14 of 18 glioblastomas. In all normal brain and tumor samples, with the exception of metastases from colon carcinoma, PCR analysis demonstrated one prominent product that corresponded to the CD44H hematopoietic form of CD44. Metastases from colon carcinoma demonstrated two prominent PCR amplification products corresponding to CD44H and CD44R1. These results suggest that CD44H is the predominant isoform of this protein in normal human brain tissue and in human neuroectodermal tumors of varying degrees of malignancy. The ability of CD44H to mediate tumor cell motility and invasiveness (in contrast to CD44R1) suggests that the CD44 alternative splicing pattern of neuroectoderm-derived tumors may enhance their local biological aggressiveness and intracerebral spread. The lack of expression of larger molecular weight CD44 variants by primary brain tumors may also partially explain why these tumors rarely metastasize to distant sites.

Alternative Splicing↗
PubMed1994

Alternate RNA splicing of murine nfkb1 generates a nuclear isoform of the p50 precursor NF-kappa B1 that can function as a transactivator of NF-kappa B-regulated transcription.

The NF-kappa B1 subunit of the transcription factor NF-kappa B is derived by proteolytic cleavage from the N terminus of a 105-kDa precursor protein. The C terminus of p105NF-kappa B1, like those of I kappa B proteins, contains ankyrin-related repeats that inhibit DNA binding and nuclear localization of the precursor and confer I kappa B-like properties upon p105NF-kappa B1. Here we report the characterization of two novel NF-kappa B1 precursor isoforms, p84NF-kappa B1 and p98NF-kappa B1, that arise by alternate splicing within the C-terminal coding region of murine nfkb1. p98NF-kappa B1, which lacks the 111 C-terminal amino acids (aa) of p105NF-kappa B1, has a novel 35-aa C terminus encoded by an alternate reading frame of the gene. p84NF-kappa B1 lacks the C-terminal 190 aa of p105NF-kappa B1, including part of ankyrin repeat 7. RNA and protein analyses indicated that the expression of p84NF-kappa B1 and p98NF-kappa B1 is restricted to certain tissues and that the phorbol myristate acetate-mediated induction of p84NF-kappa B1 and p105NF-kappa B1 differs in a cell-type-specific manner. Both p84NF-kappa B1 and p98NF-kappa B1 are found in the nuclei of transfected cells. Transient transfection analysis revealed that p98NF-kappa B1, but not p105NF-kappa B1 or p84NF-kappa B1, acts as a transactivator of NF-kappa B-regulated gene expression and that this is dependent on sequences in the Rel homology domain required for DNA binding and on the novel 35 C-terminal aa of this isoform. In contrast to previous findings, which indicated that p105NF-kappa B1 does not bind DNA, all of the NF-kappa B1 precursors were found to specifically bind with low affinity to a highly restricted set of NF-kappa B sites in vitro, thereby raising the possibility that certain of the NF-kappa B1 precursor isoforms may directly modulate gene expression.

Alternative Splicing↗
PubMed1996

restless, an active Ac-like transposon from the fungus Tolypocladium inflatum: structure, expression, and alternative RNA splicing.

Elements of the hAT transposon family, such as the maize activator (Ac), have been discovered in a large number of eukaryotic species. This type of class II transposon, present in both plants and animals, has not been previously detected in any fungal organism. However, using a differential screening method to detect repetitive DNA, we have identified a hAT transposon in the hyphomycete Tolypocladium inflatum. The transposon, which we named restless, is 4,097 bp long, carries 20-bp inverted repeats and an 8-bp target site duplication, and encodes a long open reading frame which is interrupted by a single intronic sequence. The derived mRNA exhibits alternative splicing, resulting in the formation of two transcripts that may be translated into polypeptides of 157 or 803 amino acids. The predicted amino acid sequence of the larger polypeptide demonstrates significant homology with transposases from the hAT transposon family. A chromosomal analysis using pulsed-field gel electrophoresis showed that all seven chromosomal bands carry copies of the 4.1-kb transposon. This was confirmed in hybridization experiments with rare-cutting restriction endonucleases which indicate that about 15 copies are present in T. inflatum. The genomic distribution of restless and its transcriptional expression, alternative mRNA splicing, and genomic mobility all imply a potential role for this element in developing a transposon tagging system for use in filamentous fungi.

Alternative Splicing↗
PubMed1994

Modification of alternative messenger RNA splicing of fibroblast growth factor receptors in human cardiac allografts during rejection.

Accelerated coronary atherosclerosis in cardiac transplants (cardiac allograft vasculopathy, CAV) is characterized by coronary intimal hyperplasia. Acidic fibroblast growth factor (aFGF) is a potent mitogen for vascular smooth muscle cells and endothelial cells, and its expression is increased in cardiac allografts, suggesting it may play a role in the pathogenesis of CAV. The activity of aFGF is dependent on binding to transmembrane receptors. To investigate whether receptors for aFGF are also induced after transplantation, polymerase chain reaction, in situ hybridization, and immunohistochemistry were used to analyze expression of four receptors for aFGF (FGFR1-FGFR4). Expression of mRNA encoding extracellular immunoglobulin-like domains of FGFR1 was increased 35-fold in cardiac allografts compared with normal hearts and was predominantly present in cardiac myocytes and vascular structures. Alternatively spliced mRNA that encodes transmembrane forms of FGFR1, which contain the signal-transducing tyrosine kinase domains, was induced in allografts during rejection, in infiltrating cells, vascular structures, and myocytes. In vitro experiments showed that differential expression of FGF receptor isoforms was induced by aFGF, and also by IL-6 and TGF-beta, which are expressed in cardiac allografts during rejection. The results show that expression of both aFGF and its receptors is altered in cardiac allografts and suggest that these events are important in the pathogenesis of CAV.

Adolescent↗
PubMed1997

Structural analysis of the human BIN1 gene. Evidence for tissue-specific transcriptional regulation and alternate RNA splicing.

BIN1 is a putative tumor suppressor that was identified through its interaction with the MYC oncoprotein. To begin to identify elements of BIN1 whose alteration may contribute to malignancy, we cloned and characterized the human BIN1 gene and promoter. Nineteen exons were identified in a region of >54 kilobases, six of which were alternately spliced in a cell type-specific manner. One alternately spliced exon encodes part of the MYC-binding domain, suggesting that splicing controls the MYC-binding capacity of BIN1 polypeptides. Four other alternately spliced exons encode amphiphysin-related sequences that were included in brain-specific BIN1 species, also termed amphiphysin isoforms or amphiphysin II. The 5'-flanking region of BIN1 is GC-rich and lacks a TATA box but directs transcriptional initiation from a single site. A approximately 0. 9-kilobase fragment from this region was sufficient for basal transcription and transactivation by MyoD, which may account for the high levels of BIN1 observed in skeletal muscle. This study lays the foundation for genetic and epigenetic investigations into the role of BIN1 in normal and neoplastic cell regulation.

Adaptor Proteins, Signal Transducing↗
PubMed1997

Comparisons of CYP2D messenger RNA splice variant profiles in human lung tumors and normal tissues.

Allelic variants of the CYP2D6 gene, a member of the cytochrome P450 gene superfamily, have been implicated in susceptibility to lung carcinogenesis. Human breast CYP2D6 and CYP2D7P (from a pseudogene) mRNAs were previously reported to be expressed as a series of splice variants. In this study, the expression of full-length and splice variants of these mRNAs in human lung tissue and tumors are reported for the first time and are compared in order to probe the potential for differential CYP2D6 regulation in lung normal tissue and tumors. The splice variant profiles differed within the same individual, but no consistent differences were detected.

Adult↗
PubMed1993

IL-2 regulates the expression of the NK-TR gene via an alternate RNA splicing mechanism.

We have recently isolated and characterized human and mouse genes of a putative natural killer (NK) cell tumour-recognition protein (NK-TR) that is specifically expressed in NK cells. This gene codes for a 150 kD protein with a cyclophilin-related amino terminus followed by several positively charged domains. We report here the discovery of two sites of alternate splicing in the 5' region of the NK-TR mRNA. One of these events caused a frameshift in the open reading frame by splicing in a 28 bp exon within the cyclophilin coding region, resulting in the premature termination of the NK-TR protein. The second alternate splice stemmed from the use of an internal splice acceptor within an exon, producing a deletion of 25 amino acids in the NK-TR protein. The activation of NK cells by IL-2 produced a change in the splicing pattern that resulted in increased production of mRNAs capable of producing the complete NK-TR protein.

Alternative Splicing↗
PubMed1993

Genomic structure of the locus encoding protein 4.1. Structural basis for complex combinational patterns of tissue-specific alternative RNA splicing.

Protein 4.1 (P4.1) is a multifunctional protein with heterogeneity in molecular weight, intracellular localization, tissue- and development-specific expression patterns. We have analyzed the genomic structure of the locus encoding mouse P4.1 and have systematically analyzed diverse P4.1 mRNA isoforms expressed in erythroid and nonerythroid tissues. Our results indicate that the mouse protein 4.1 gene, over 90 kilobases long, comprises at least 23 exons (13 constitutive exons, 10 alternative exons) interrupted by 22 introns. The donor and acceptor splice site sequences match the consensus sequences for the exon-intron boundaries of most eukaryotic genes. No significant sequence difference was observed between splice junctions of alternative and constitutive exons. Apparently, most alternative exon-encoded peptides are located within particular functional domains of the P4.1 protein: two peptides encoded by alternative exons 4 and 5 are located near or within the glycophorin/calmodulin binding domain, whereas three other alternative exon-encoded peptides (19-amino acid encoded by exon 14, 14-amino acid encoded by exon 15, and 21-amino acid encoded by exon 16) are located near or within the spectrin-actin binding domain. Selective use of exon 2', which carries an upstream translation initiation codon (AUG), may produce an elongated P4.1 isoform (135 kDa) that is predominantly expressed in nonerythroid tissues. Combinatorial splicing of these exons may generate different isoforms that exhibit complicated tissue-specific expression patterns.

3T3 Cells↗
PubMed1997

RNA splicing ligase activity in the archaeon Haloferax volcanii.

At least two separate enzymes, an endonuclease and a ligase, appear to be involved in tRNA splicing in halophilic archaea. We have identified and partially characterized a splicing ligase activity in cell extracts of Haloferax volcanii that can ligate deproteinized exon products generated in a separate endonuclease reaction. As in vitro transcribed partial intron-deleted derivative of H. volcanii elongator tRNA(Met) is used as substrate for the endonuclease. The ligase can also join the two exons that are independently eluted from the gels. This ligase activity is observed at a range (50 mM to 2.8 M) of monovalent cations in the assays, but is abolished when the enzyme preparations are depleted of the monovalent cations. In contrast, H. volcanii splicing endonuclease has been reported to require divalent cations and is inhibited by monovalent cations. Our endonuclease assays confirm these reports, and also show that the endonuclease is not permanently inactivated even in high monovalent cation containing extracts. The ligase activity in the extracts does not appear to require any divalent cation or exogenously added source of energy or phosphate.

Archaea↗
PubMed1994

Alternative splicing of RNA transcripts encoded by the murine p105 NF-kappa B gene generates I kappa B gamma isoforms with different inhibitory activities.

The gene encoding the 105-kDa protein (p105) precursor of the p50 subunit of transcription factor NF-kappa B also encodes a p70 I kappa B protein, I kappa B gamma, which is identical to the C-terminal 607 amino acids of p105. Here we show that alternative RNA splicing generates I kappa B gamma isoforms with properties different from those of p70. One 63-kDa isoform, termed I kappa B gamma-1, which lacks 59 amino acids C-terminal to ankyrin repeat 7, has a novel 35-amino acid C terminus encoded by an alternative reading frame of the p105 gene. A 55-kDa isoform, I kappa B gamma-2, lacks the 190 C-terminal amino acids of p70I kappa B gamma. In contrast to p70I kappa B gamma, which is a cytoplasmic protein, I kappa B gamma-1 is found in both the cytoplasm and nucleus, whereas I kappa B gamma-2 is predominantly nuclear. The I kappa B gamma isoforms also display differences in specificity and affinity for Rel/NF-kappa B proteins. While p70I kappa B gamma inhibits p50-, p65-, and c-Rel-mediated transactivation and/or DNA binding, both I kappa B gamma-1 and I kappa B gamma-2 are specific for p50 and have different affinities for this subunit. The absence in I kappa B gamma-1 and I kappa B gamma-2 of a protein kinase A site whose phosphorylation modulates p70I kappa B gamma inhibitory activity suggests that alternative RNA splicing may be used to generate I kappa B gamma isoforms that respond differently to intracellular signals.

Alternative Splicing↗
PubMed1999

A series of mutations in the dihydropteridine reductase gene resulting in either abnormal RNA splicing or DHPR protein defects. Mutations in brief no. 244. Online.

Five novel mutations are described which result in the rare hyperphenylalaninemia DHPR-deficiency. Three of these are located at different intron/exon boundaries within the DHPR gene, and disrupt the maturation of the DHPR transcript such that little full-length mRNA can be detected by RT-PCR. Each mutation alters a conserved nucleotide within the splice site consensus sequence, and results in the skipping of an exon and, in one case, the activation of an inappropriate splicing signal. Two further mutations are missense mutations resulting in a non-conservative amino acid change within the DHPR protein (L14P and G17V) and are associated with a severe phenotype.

Alternative Splicing↗
PubMed2005

Intron 2 [IVS2, T-C +4] HFE gene mutation associated with S65C causes alternative RNA splicing and is responsible for iron overload.

UNLABELLED: A patient with congenital liver fibrosis revealed a high transferrin saturation index and iron overload on liver biopsy. He did not carry the most frequent HFE mutations: C282Y or H63D. Heterozygosity was detected for S65C. Unknown HFE mutations were also sought using a combined denaturing high performance liquid chromatography (DHPLC)/direct sequence approach and another point mutation, a transition T-C (nt 4910), at the fourth base of the donor splice site of intron 2 [HFE, intervening sequence (IVS) 2, T-C +4] was found. Family screening revealed that a daughter carried both S65C and [IVS2, T-C +4]. CONCLUSION: : The existence in our proband of a partly-altered HFE protein in the region encoded by exon 2 might be responsible for the histologically-demonstrated iron overload.

Journal Article↗