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At least 37 records · Page 2Linked to original sources
PubMed1994

Diesel exhaust particles induce local IgE production in vivo and alter the pattern of IgE messenger RNA isoforms.

Diesel exhaust particles (DEP) have been implicated in the increased incidence of allergic airway disorders. We investigated the effects of DEP on localized immunoglobulin production by performing nasal challenges with varying doses of DEP and analyzing the local immune response in nasal lavages obtained before and after. A significant rise in nasal IgE but not IgG, IgA, IgM, or albumin was observed in subjects 4 d after challenge with 0.30 mg DEP, equivalent to exposure on an average Los Angeles day. Direct evidence for DEP-enhanced local production of IgE was that challenge increased the number of IgE-secreting cells in lavage fluid from < 1 in 2,000,000 to > 1 in 100,000 but did not alter the number of IgA-secreting cells. There was a concomitant increase in epsilon mRNA production in the lavage cells. Additionally, DEP altered the relative amounts of five different epsilon mRNAs generated by alternative splicing, mRNAs that code for different IgE proteins. These results show that DEP exposure in vivo causes both quantitative and qualitative changes in local IgE production. The implication is that natural exposure to DEP may result in increased expression of respiratory allergic disease.

Adult↗
PubMed1994

Structure of the gene for the testis-specific proprotein convertase 4 and of its alternate messenger RNA isoforms.

Proprotein convertase 4 (PC4) is a mammalian secretory serine endoproteinase similar to the yeast KEX2 gene product and specifically expressed in testicular germs cells. PC4 mRNA isoforms that vary in size and 3' coding sequence have been reported (N. G. Seitah, R. Day, J. Hamelin, A. Gaspar, M. W. Collard, and M. Chrétien, 1992, Mol. Endocrinol. 6: 1559-1570). To determine the origin of these various forms, the mouse PC4 gene was cloned and its organization determined. The structural gene is approximately 9.5 kb long. It contains 15 exons and 14 introns. The exon-intron organization is very similar to that of the genes for the related convertases furin, PC1, and PC2. The upstream region carries several GGGCGG and three CCAAT but no TATAA motifs. Analysis of the 5' end of PC4 mRNA in the testis has led to the identification of two novel 5' splice variants that might encode a nonsecretory enzyme. The multiple forms of PC4 mRNA can all be explained by alternate splicing of primary transcripts of a single gene.

Alternative Splicing↗
PubMed2000

Two Sox9 messenger RNA isoforms: isolation of cDNAs and their expression during gonadal development in the frog Rana rugosa.

Sox is a family of SRY-related testis-determining genes. We have isolated two different mRNA isoforms of the frog Sox9 gene from adult frog testis cDNAs. One form (Sox9 alpha) encodes a 482 amino acid protein containing the HMG box, whereas the other form (Sox9 beta), which completely lacks the HMG box, is a truncated 265 amino acid protein of Sox9 alpha. Sox9 alpha is 82% similar to mouse, 86% to chicken, and 77% to trout Sox9 at the amino acid level. Sox9 expression was upregulated in embryos after stage 16, and was seen in both developing testes and ovaries. The size of Sox9 transcripts was determined to be 7.8 knt by Northern blot analysis. In addition, Sox9 alpha expression was found prominently in the testis and brain among various tissues of adult frogs examined, and was considerably higher than Sox9 beta. The fact that Sox9 is expressed in both sexes suggests that this gene is involved in gonadal development of male and female frogs. This is dissimilar to the pattern in birds and mammals, in which Sox9 expression is male-specific.

Animals↗
PubMed1995

Multiple APC messenger RNA isoforms encoding exon 15 short open reading frames are expressed in the context of a novel exon 10A-derived sequence.

Intragenic splice mechanisms affecting the coding exons 8 to 15 of the human adenomatous polyposis coli (APC) gene have been analyzed. Three mechanisms within this gene area were found to contribute to mRNA heterogeneity: (i) facultative expression of exon 9-encoded sequences; (ii) in-frame insertion of a 54-nucleotide sequence encoded by a novel exon located 1.6 kb down-stream from exon 10, provisionally designated APC exon 10A; (iii) skipping of exon 14, resulting in a novel exon 13/15 connection. Interestingly the latter event provided the mRNA with a novel open reading frame, which was terminated after 19 codons of exon 15-derived sequences. Combinatorial joining of these segments yielded 7 different transcripts in addition to an mRNA species resulting from an exon 10/15 connection, as determined by cloning and sequence analysis. RT-PCR expression analyses were carried out to demonstrate that this complexity of splice variants is indeed synthesized in cell lines derived from various tissues. Furthermore, in accordance with our findings at the transcript level, we provide Western blot analyses demonstrating that moderate steady-state levels of genuine APC-specific low m.w. polypeptide chains exist. These APC "light chains", however, are not identical with polypeptide chains, which have been reported to accompany apoptosis and necrosis, since the molecules described here are definitively co-expressed with p300apc at the transcript and protein levels.

Amino Acid Sequence↗
PubMed2003

Identification of new human coding steroid receptor RNA activator isoforms.

SRA is a steroid receptor co-activator which acts as a functional RNA and is classified as belonging to the growing family of functional non-coding RNAs. None of the different SRA transcripts described to date encode a detectable SRA protein following in vitro and in vivo translation experiments. We have identified three new SRA-RNA isoforms differing mainly from the originally cloned SRA by an extended 5(') extremity. These long SRA isoforms, able to encode a stable protein in vitro, led to the production in vivo of a nuclear protein when transfected into the MCF-7 human breast cancer cell line. Reverse-transcription polymerase chain reaction and Western blot analysis of RNA and protein extracts from different breast cancer cell lines confirmed the presence of endogenous coding SRA isoforms and their corresponding proteins. Our results demonstrate that full-length SRA-RNAs likely to encode stable proteins are widely expressed in breast cancer cell lines.

5' Flanking Region↗
PubMed1996

A switch in broad-complex zinc-finger isoform expression is regulated posttranscriptionally during the metamorphosis of Drosophila imaginal discs.

The Broad-Complex (BR-C) is a key member of the 20-hydroxyecdysone regulatory hierarchy that coordinates changes in gene expression during Drosophila metamorphosis. The family of transcription factors encoded by the BR-C share a common amino-terminal domain which is fused by alternative splicing to one of four pairs of C2H2 zinc-finger domains (Z1, Z2, Z3, and Z4). In this study, we examine the temporal expression of transcripts encoding each BR-C zinc-finger isoform-including the newly discovered fourth zinc-finger domain-during the metamorphosis of imaginal discs which form the integumental structures of the adult head and thorax. We find that all BR-C zinc-finger RNA isoforms are induced as a primary response to 20-hydroxyecdysone. However, induced BR-C RNA isoforms exhibit two divergent expression profiles. The Z2, Z3, and Z4 RNA isoforms accumulate to high levels at the beginning of the ecdysone response and abruptly disappear after several hours. In contrast, the Z1 RNA isoform continues to accumulate while the others decline, resulting in a switch in relative isoform levels. Using probes specific to different regions of the BR-C, we show that the switch in BR-C RNA isoform expression appears to be posttranscriptionally regulated, presumably by ecdysone-responsive factors. We propose that this switch results from a change in splice acceptor site choice. Finally, we present a model describing how this temporal switch in isoform expression could mediate changes in BR-C function, from transcriptional activation to repression and vice versa, that are critical for coordinate downstream target gene expression.

Amino Acid Sequence↗
PubMed2002

Species-specific and isoform-specific RNA binding of human and mouse fragile X mental retardation proteins.

The loss of the fragile X RNA binding protein, FMRP, causes macroorchidism and mental retardation in man. The discovery of a mouse ortholog led to the development of several FMRP knockout mouse strains that recapitulate some features of the disease. As mouse and human FMRPs differ in several amino acids in their RNA binding domains, we compared the RNA binding profiles of these two orthologs. Five variant FMRPs, whose differences arose from alternative splicing and mutation within the conserved RNA binding domains, were examined. Homoribopolymer binding studies showed that human FMRPs (hFMRP) bound a broader range of single-stranded mimetics than mouse FMRPs (mFMRP) and these interactions were both complex and cooperative. hFMRP and mFMRP also displayed significant preferences toward binding their own mRNA; specifically we found that the mFMRP isoforms bind mFMR1 mRNA much more tightly than their human counterparts. Finally, these data demonstrate that each FMRP variant binds RNAs uniquely, resulting in a set of proteins with differing affinities.

Alternative Splicing↗
PubMed2000

Altered G protein-coupling functions of RNA editing isoform and splicing variant serotonin2C receptors.

Different isoforms of serotonin subtype 2C receptor (5-HT(2C)R) with altered G protein-coupling efficacy are generated by RNA editing, which converts genomically encoded adenosine residues into inosines. In combination, editing of five sites all located within the second intracellular loop region of 5-HT(2C)R mRNA changes the gene-encoded Ile, Asn, and Ile at positions 156, 158, and 160, respectively. We analyzed the G protein-coupling functions of previously unreported editing isoform receptors. An approximately 13-fold reduction in the agonist potency for G protein-coupling stimulation as well as a significantly reduced basal level activity was observed with the thalamus-specific isoform carrying Ile156, Gly158, and Val160 (5-HT(2C)R-IGV). In contrast, the agonist was four- to five-fold less potent with 5-HT(2C)R-MSV and -IDV, detected in the amygdala and choroid plexus, respectively, indicating a dominant role for the amino acid residue at position 158 in receptor functions. We also identified a splicing variant receptor with a truncated C terminus that displayed no ligand binding capacity or G protein-coupling activity. Examination of the alternatively spliced RNA encoding this truncated receptor suggests that editing of this variant RNA occurs after completion of splicing, resulting in complete editing at all five sites.

3T3 Cells↗
PubMed2004

Human DDX3Y, the Y-encoded isoform of RNA helicase DDX3, rescues a hamster temperature-sensitive ET24 mutant cell line with a DDX3X mutation.

We investigated the function of DDX3Y, the Y chromosome AZFa region encoding a putative DEAD-box RNA helicase protein, the loss of which results in oligozoospermia or azoospermia in humans. The human DDX3Y amino acid sequence is similar to that of the X chromosome gene DDX3X (91.7% homology). Here we report that human Y- and X-encoded DEAD box RNA helicase proteins DDX3Y and DDX3X are interchangeable and have an essential function: both proteins rescued a temperature-sensitive mutant hamster cell line (tsET24) that was otherwise incapable of growth at a nonpermissive temperature. Mouse homologues Ddx3y and D1Pas1-PL10 also rescued the mutant cell line at a nonpermissive temperature. In situ hybridization revealed that Ddx3x mRNA was expressed in almost every cell in mouse testis, suggesting that Ddx3x is involved in spermatogenesis. A comparative study of DDX3X and DDX3Y was performed to determine the significance of DDX3Y for cell growth and spermatogenesis. Both DDX3X and DDX3Y promoter DNAs produced a similar degree of transcription in vivo, whereas deletion studies of the promoter DNAs indicated that these genes are differentially regulated. DDX3Y, similar to DDX3X, shuttles between the nucleus and cytoplasm in a crm1-dependent manner.

Active Transport, Cell Nucleus↗
PubMed1998

Genetic characterization of the mRNAs encoding alpha-bungarotoxin: isoforms and RNA editing in Bungarus multicinctus gland cells.

The mRNA encoding alpha-bungarotoxin (alpha-Butx) was prepared from the venom glands of Bungarus multicinctus by Cap-finder cDNA synthesis. The sequences of the 3'- and 5'-flanking regions including a signal peptide of alpha-Butx were almost identical with those of Elapidae and Hydrophiidae toxins, suggesting that they may have the same origin. Sixteen polymorphic mRNA sequences of alpha-Butx were detected from B.multicinctus gland cells. Analysis of the genomic DNA of alpha-Butx indicated that the polymorphic mRNA originated from one DNA sequence. Most of the mutations in alpha-Butx mRNA were silent and the hot-spot variations occurred at 78, 107, 129, 198 and 201 nt in alpha-Butx mRNA. Ten distinct protein sequences of alpha-Butx could be deduced from the polymorphic mRNA and one of the isoforms has already been isolated. Since alpha-Butx DNA is a single copy in the genome, the RNA polymorphism might result from post-transcriptional editing. These results indicate that the authentic alpha-Butx is in fact derived from edited mRNAs. RNA editing may contribute a common mechanism toward the diversity of alpha-neurotoxins in snake glands.

Amino Acid Sequence↗
PubMed2001

The BCL11 gene family: involvement of BCL11A in lymphoid malignancies.

Many malignancies of mature B cells are characterized by chromosomal translocations involving the immunoglobulin heavy chain (IGH) locus on chromosome 14q32.3 and result in deregulated expression of the translocated oncogene. t(2;14)(p13;q32.3) is a rare event in B-cell malignancies. In contrast, gains and amplifications of the same region of chromosome 2p13 have been reported in 20% of extranodal B-cell non-Hodgkin lymphomas (B-NHL), in follicular and mediastinal B-NHL, and in Hodgkin disease (HD). It has been suggested that REL, an NF-kappaB gene family member, mapping within the amplified region, is the pathologic target. However, by molecular cloning of t(2;14)(p13;q32.3) from 3 cases of aggressive B-cell chronic lymphocytic leukemia (CLL)/immunocytoma, this study has shown clustered breakpoints on chromosome 2p13 immediately upstream of a CpG island located about 300 kb telomeric of REL. This CpG island was associated with a Krüppel zinc finger gene (BCL11A), which is normally expressed at high levels only in fetal brain and in germinal center B-cells. There were 3 major RNA isoforms of BCL11A, differing in the number of carboxy-terminal zinc fingers. All 3 RNA isoforms were deregulated as a consequence of t(2;14)(p13;q32.3). BCL11A was highly conserved, being 95% identical to mouse, chicken, and Xenopus homologues. BCL11A was also highly homologous to another gene (BCL11B) on chromosome 14q32.1. BCL11A coamplified with REL in B-NHL cases and HD lymphoma cell lines with gains and amplifications of 2p13, suggesting that BCL11A may be involved in lymphoid malignancies through either chromosomal translocation or amplification.

Amino Acid Sequence↗
PubMed2003

Regulation of estrogen receptor (ER) isoform messenger RNA expression by different ER ligands in female rat pituitary.

Net estrogen sensitivity in target tissues critically depends on the regulated expression of full-length and alternately processed estrogen receptor (ER) isoforms. However, the molecular mechanisms for the control of pituitary responsiveness to estrogen remain partially unknown. In the present communication, we report the ability of different ligands, with distinct agonistic or antagonistic properties at the ER, to modulate the expression of the transcripts encoding ERalpha and ERbeta isoforms, as well as those for the truncated ERalpha product (TERP), and the variant ERbeta2, in pituitaries from ovariectomized rats, i.e., a background devoid of endogenous estrogen. Compared with expression levels at the morning of proestrus, ovariectomy (OVX) resulted in increased pituitary expression of ERbeta and ERbeta2 mRNAs, whereas it decreased TERP-1 and -2 levels without affecting those of ERalpha. Administration of estradiol benzoate (as potent agonist for alpha and beta forms of ER) or the selective ERalpha agonist, propyl pyrazole triol, fully reversed the responses to OVX, while the ERbeta ligand, diarylpropionitrile, failed to induce any significant effect except for a partial stimulation of TERP-1 and -2 mRNA expression levels. To note, the ERbeta agonist was also ineffective in altering pituitary expression of progesterone receptor-B mRNA, i.e., a major estrogen-responsive target. In all parameters tested, tamoxifen, a selective ER modulator with mixed agonist/antagonist activity, behaved as ERalpha agonist, although the magnitude of tamoxifen effects was significantly lower than those of the ERalpha ligand, except for TERP induction. In contrast, the pure antiestrogen RU-58668 did not modify the expression of any of the targets under analysis. Overall, our results indicate that endogenous estrogen differentially regulates pituitary expression of the mRNAs encoding several ER isoforms with distinct functional properties, by a mechanism that is mostly conducted through ERalpha. Differential regulation of ER isoforms may represent a relevant system for the self-tuning of estrogen responsiveness in female pituitary.

Animals↗
PubMed2004

A quantitative and comparative study of the effects of a synthetic ciguatoxin CTX3C on the kinetic properties of voltage-dependent sodium channels.

Ciguatoxins (CTXs) are known to bind to receptor site 5 of the voltage-dependent Na channel, but the toxin's physiological effects are poorly understood. In this study, we investigated the effects of a ciguatoxin congener (CTX3C) on three different Na-channel isoforms, rNa(v)1.2, rNa(v)1.4, and rNa(v)1.5, which were transiently expressed in HEK293 cells. The toxin (1.0 micromol l(-1)) shifted the activation potential (V(1/2) of activation curve) in the negative direction by 4-9 mV and increased the slope factor (k) from 8 mV to between 9 and 12 mV (indicative of decreased steepness of the activation curve), thereby resulting in a hyperpolarizing shift of the threshold potential by 30 mV for all Na channel isoforms. The toxin (1.0 micromol l(-1)) significantly accelerated the time-to-peak current from 0.62 to 0.52 ms in isoform rNa(v)1.2. Higher doses of the toxin (3-10 micromol l(-1)) additionally decreased time-to-peak current in rNa(v)1.4 and rNa(v)1.5. A toxin effect on decay of I(Na) at -20 mV was either absent or marginal even at relatively high doses of CTX3C. The toxin (1 micromol l(-1)) shifted the inactivation potential (V(1/2) of inactivation curve) in the negative direction by 15-18 mV in all isoforms. I(Na) maxima of the I-V curve (at -20 mV) were suppressed by application of 1.0 micromol l(-1) CTX3C to a similar extent (80-85% of the control) in all the three isoforms. Higher doses of CTX3C up to 10 micromol l(-1) further suppressed I(Na) to 61-72% of the control. Recovery from slow inactivation induced by a depolarizing prepulse of intermediate duration (500 ms) was dramatically delayed in the presence of 1.0 micromol l(-1) CTX3C, as time constants describing the monoexponential recovery were increased from 38+/-8 to 588+/-151 ms (n=5), 53+/-6 to 338+/-85 ms (n=4), and 23+/-3 to 232+/-117 ms (n=3) in rNa(v)1.2, rNa(v)1.4, and rNa(v)1.5, respectively. CTX3C exerted multimodal effects on sodium channels, with simultaneous stimulatory and inhibitory aspects, probably due to the large molecular size (3 nm in length) and lipophilicity of this membrane-spanning toxin.

Animals↗
PubMed1993

APC gene messenger RNA: novel isoforms that lack exon 7.

The APC gene at human chromosome 5q21 is responsible for familial adenomatous polyposis coli. Furthermore, sporadic cancers of not only colon but also other digestive organs often contain mutations in the APC gene. A dominant mouse mutation Min that was generated by chemical mutagenesis and causes polyposis in the digestive tract is in the mouse homologue of the human APC gene. The APC mRNA is generated from 15 exons. Two mRNA isoforms were reported which are produced by alternative splicing in the 9th exon. Here, we report novel mRNA isoforms that lack the 7th exon in both mouse and human cells.

Animals↗
PubMed1998

Expression of a new RNA-splice isoform of WT1 in developing kidney-gonadal complexes of the turtle, Trachemys scripta.

WT1 is a tumor suppressor gene encoding a zinc finger DNA-binding protein required for normal vertebrate kidney and gonad development. Although the sequence and function of this gene has been studied mostly in mammals, comparative analysis in other vertebrates may suggest regions of conservation of function as well as evolution of function. We have initiated a study of this gene in the freshwater turtle, Trachemys scripta, a species that demonstrates temperature dependent sex determination. The turtle WT1 amino acid sequence (GenBank Accession No. AF019779) is over 85% identical to that of other species overall, but there are some major differences. The greatest differences are in the N-terminal portion of the peptide which is thought to mediate transcriptional repression by interaction with other proteins. Turtle WT1, like those of the alligator, chicken, and Xenopus lacks the proline- and glycine-rich stretches that are present in mammalian WT1. Exon 5, which is alternatively spliced in mammals, is altogether absent in the non-mammalian vertebrates. In addition, turtle WT1 is alternatively spliced so that exon 4 is either present or absent. These differences suggest that the interaction of reptilian WT1 with other factor required for mediation of activity may be different than the interaction of mammalian WT1. It also suggests that alternative splicing is a conserved regulatory mechanism of vertebrate WT1. Expression of WT1 in turtle embryonic kidney-gonadal complexes begins after the mesonephroi have formed and continues at least until the bipotential gonad begins to differentiate. Although the proportions of the different splice isoforms are relatively constant during these stages of kidney development, the level of steady state expression is increased in embryos incubated at 26 degrees C, the testis-producing temperature.

Alternative Splicing↗
PubMed2003

Differential expression of equine myosin heavy-chain mRNA and protein isoforms in a limb muscle.

The horse is one of the few animals kept and bred for its athletic performance and is therefore an interesting model for human sports performance. The regulation of the development of equine locomotion in the first year of life, and the influence of early training on later performance, are largely unknown. The major structural protein in skeletal muscle, myosin heavy-chain (MyHC), is believed to be primarily transcriptionally controlled. To investigate the expression of the MyHC genes at the transcriptional level, we isolated cDNAs encoding the equine MyHC isoforms type 1 (slow), type 2a (fast oxidative), and type 2d/x (fast glycolytic). cDNAs encoding the 2b gene were not identified. The mRNA expression was compared to the protein expression on a fiber-to-fiber basis using in situ hybridization (non-radioactive) and immunohistochemistry. Marked differences were detected between the expression of MyHC transcripts and MyHC protein isoforms in adult equine gluteus medius muscle. Mismatches were primarily due to the presence of hybrid fibers expressing two fast (2ad) MyHC protein isoforms, but only one fast (mainly 2a) MyHC RNA isoform. This discrepancy was most likely not due to differential mRNA expression of myonuclei.

Amino Acid Sequence↗
PubMed1995

The rat hepatic leukemia factor (HLF) gene encodes two transcriptional activators with distinct circadian rhythms, tissue distributions and target preferences.

Hepatic leukemia factor (HLF) is a member of the PAR family of transcription regulatory proteins. We have characterized the rat HLF gene and studied its expression and activity. The rat HLF gene is transcribed from two alternative promoters, alpha and beta, with different circadian amplitudes and tissue specificities. The alpha RNA isoforms produce a 43 kDa protein, HLF43, abundant in brain, liver and kidney, like the previously described human HLF RNA. The beta RNA HLF isoforms use a CUG codon to initiate translation of a novel 36 kDa protein, HLF36, which is shorter at its N-terminus relative to the 43 kDa form. HLF36 is expressed uniquely in the liver, where it is the most abundant HLF protein. Surprisingly, the two proteins accumulate in the liver with different circadian amplitudes and have distinct liver-specific promoter preferences in transfection experiments. Thus, HLF43 stimulates transcription from the cholesterol 7 alpha-hydroxylase promoter much more efficiently than from the albumin promoter, while the converse is true for HLF36.

Amino Acid Sequence↗
PubMed2003

Differential expression of the two Drosophila fos/kayak transcripts during oogenesis and embryogenesis.

The Dfos/kayak gene encodes a bZIP protein, DFos, required in a large variety of differentiation and morphogenetic processes throughout Drosophila development. The recent availability of an expressed sequence tag (EST) sequence led us to identify a novel kay mRNA encoding a deduced DFos isoform showing a specific NH(2)-terminal region. To gain further insight into the function and the regulation of this gene, we have investigated the expression pattern of the two kay mRNA isoforms, kay-RA and kay-RB, during oogenesis and embryogenesis by whole-mount in situ hybridization. Results show that, although the two kay RNA isoforms display fully distinct patterns of transcription during oogenesis, they show partially overlapping expression profiles in embryos. These data reveal a previously unsuspected level of complexity in the regulation of the expression of the kay gene. In addition, they suggest a possible requirement for this gene in the invagination processes during early gastrula stages.

Amino Acid Sequence↗