scholarly journals A novel pathway for alternative splicing: identification of an RNA intermediate that generates an alternative 5' splice donor site not present in the primary transcript of AMPD1.

1990 ◽  
Vol 10 (10) ◽  
pp. 5271-5278 ◽  
Author(s):  
I Mineo ◽  
P R Clarke ◽  
R L Sabina ◽  
E W Holmes
Keyword(s):  
Alternative Splicing ◽  
Donor Site ◽  
Specific Pattern ◽  
Splice Donor Site ◽  
Splice Donor ◽  
Primary Transcript ◽  
Exon 2 ◽  
Tissue Specific ◽  
Exon 1 ◽  
Alternatively Spliced

AMP deaminase (AMPD) is a central enzyme in eucaryotic energy metabolism, and tissue-specific as well as stage-specific isoforms are found in many vertebrates. This study demonstrates the AMPD1 gene product in rat is alternatively spliced. The second exon, a 12-base miniexon, was found to be excluded or included in a tissue-specific and stage-specific pattern. This example of cassette splicing utilizes a unique pathway through an RNA intermediate that generates an alternative 5' splice donor site at the point where exon 2 is ligated to exon 1. In the analogous intermediate of human AMPD1, the potential 5' splice donor site created at the boundary of exon 1 and exon 2 was a poor substrate for splicing because of differences in exon 2 sequences, and human AMPD1 was not alternatively spliced. These results demonstrate that in some cases alternative splicing may proceed through an RNA intermediate that generates an alternative splice donor site not present in the primary transcript. Discrimination between alternative 5' splice donor sites in the RNA intermediate of AMPD1 is apparently controlled by tissue-specific and stage-specific signals.

A novel pathway for alternative splicing: identification of an RNA intermediate that generates an alternative 5' splice donor site not present in the primary transcript of AMPD1

1990 ◽  
Vol 10 (10) ◽  
pp. 5271-5278
Author(s):  
I Mineo ◽  
P R Clarke ◽  
R L Sabina ◽  
E W Holmes
Keyword(s):  
Alternative Splicing ◽  
Donor Site ◽  
Specific Pattern ◽  
Splice Donor Site ◽  
Splice Donor ◽  
Primary Transcript ◽  
Exon 2 ◽  
Tissue Specific ◽  
Exon 1 ◽  
Alternatively Spliced

AMP deaminase (AMPD) is a central enzyme in eucaryotic energy metabolism, and tissue-specific as well as stage-specific isoforms are found in many vertebrates. This study demonstrates the AMPD1 gene product in rat is alternatively spliced. The second exon, a 12-base miniexon, was found to be excluded or included in a tissue-specific and stage-specific pattern. This example of cassette splicing utilizes a unique pathway through an RNA intermediate that generates an alternative 5' splice donor site at the point where exon 2 is ligated to exon 1. In the analogous intermediate of human AMPD1, the potential 5' splice donor site created at the boundary of exon 1 and exon 2 was a poor substrate for splicing because of differences in exon 2 sequences, and human AMPD1 was not alternatively spliced. These results demonstrate that in some cases alternative splicing may proceed through an RNA intermediate that generates an alternative splice donor site not present in the primary transcript. Discrimination between alternative 5' splice donor sites in the RNA intermediate of AMPD1 is apparently controlled by tissue-specific and stage-specific signals.

Alternative promoter usage and alternative splicing contribute to mRNA heterogeneity of mouse monocarboxylate transporter 2

2007 ◽  
Vol 32 (1) ◽  
pp. 95-104 ◽  
Author(s):  
Shelley X. L. Zhang ◽  
Tina R. Searcy ◽  
Yiman Wu ◽  
David Gozal ◽  
Yang Wang
Keyword(s):  
Transcription Factors ◽  
Alternative Splicing ◽  
Alternative Promoter ◽  
Monocarboxylate Transporter ◽  
Specific Expression ◽  
Exon 2 ◽  
Tissue Specific ◽  
Tissue Specific Expression ◽  
Exon 1 ◽  
Promoter Usage

Expression patterns of monocarboxylate transporter 2 (MCT2) display mRNA diversity in a tissue-specific fashion. We cloned and characterized multiple mct2 5′-cDNA ends from the mouse and determined the structural organization of the mct2 gene. We found that transcription of this gene was initiated from five independent genomic regions that spanned >80 kb on chromosome 10, resulting in five unique exon 1 variants (exons 1a, 1b, 1c, 1d, and 1e) that were then spliced to the common exon 2. Alternative splicing of four internal exons (exons AS1, AS2, AS3, and exon 3) greatly increased the complexity of mRNA diversity. While exon 1c was relatively commonly used for transcription initiation in various tissues, other exon 1 variants were used in a tissue-specific fashion, especially exons 1b and 1d that were used exclusively for testis-specific expression. Sequence analysis of 5′-flanking regions upstream of exons 1a, 1b, and 1c revealed the presence of numerous potential binding sites for ubiquitous transcription factors in all three regions and for transcription factors implicated in testis-specific or hypoxia-induced gene expression in the 1b region. Transient transfection assays demonstrated that each of the three regions contained a functional promoter and that the in vitro, cell type-specific activities of these promoters were consistent with the tissue-specific expression pattern of the mct2 gene in vivo. These results indicate that tissue-specific expression of the mct2 gene is controlled by multiple alternative promoters and that both alternative promoter usage and alternative splicing contribute to the remarkable mRNA diversity of the gene.

scholarly journals Myosin light-chain 1/3 gene alternative splicing: cis regulation is based upon a hierarchical compatibility between splice sites.

1990 ◽  
Vol 10 (5) ◽  
pp. 2133-2144 ◽  
Author(s):  
M E Gallego ◽  
B Nadal-Ginard
Keyword(s):  
Alternative Splicing ◽  
Light Chain ◽  
Myosin Light Chain ◽  
Cooperative Interaction ◽  
Splice Sites ◽  
Exon 2 ◽  
Exon 1 ◽  
Alternatively Spliced ◽  
Flanking Sequences ◽  
Exon 4

The mechanisms involved in the selective joining of appropriate 5' and 3' splice sites are still poorly understood in both constitutive and alternatively spliced genes. With two promoters associated with different exons, the myosin light-chain 1/3 gene generates two pre-mRNAs that also differ by the use of a pair of internal exons, 3 and 4, that are spliced in a mutually exclusive fashion. When the promoter upstream from exon 1 is used, only exon 4 is included. If the promoter upstream from exon 2 is used, only exon 3 is included. In an attempt to understand the molecular basis for the mutually exclusive behavior of these two exons and the basis of their specific selection, a number of minigene constructs containing exons 3 and 4 were tested in a variety of homologous or heterologous cis and trans environments. The results demonstrate that the mutually exclusive behavior of myosin light-chain exons 3 and 4 and selection between the two exons are cis regulated and are affected by the nature of the flanking sequences. Both exons competed for the common flanking 5' and 3' splice sites. Flanking exons were found that favored inclusion into mature mRNA of exon 3, exon 4, both, or neither, suggesting a specific cooperative interaction between certain 5' and 3' splice sites. Thus, alternative splicing of myosin light-chain 1/3 pre-mRNAs is regulated in cis by a hierarchy of compatibilities between pairs of 5' and 3' splice sites.

Exon truncation by alternative splicing of murine ICAM-1

2002 ◽  
Vol 12 (1) ◽  
pp. 47-51 ◽  
Author(s):  
Joseph P. Mizgerd ◽  
Matt R. Spieker ◽  
Michal M. Lupa
Keyword(s):  
Alternative Splicing ◽  
Splice Variant ◽  
Donor Site ◽  
Exon Skipping ◽  
Intercellular Adhesion Molecule ◽  
Splice Donor Site ◽  
Intercellular Adhesion Molecule 1 ◽  
Membrane Expression ◽  
Alternatively Spliced ◽  
Ig Domain

The murine gene for intercellular adhesion molecule-1 (ICAM-1) encodes multiple products, arising from alternative splicing. Full-length ICAM-1 contains five extracellular Ig domains, each encoded by a separate exon. Alternatively spliced forms have Ig domains 2, 3, and/or 4 excised as a result of exon skipping. We report here a novel splice variant of murine ICAM-1, resulting from exon truncation rather than exon skipping and affecting Ig domain 5. A 5′ splice donor site within exon 6 generates transcripts missing 69 nucleic acids from the 3′ terminus of the exon. This in-frame exon truncation is predicted to replace 24 amino acids within Ig domain 5 with a single aspartic acid residue, yielding a structure other than an Ig domain immediately external to the membrane. Expression of this alternatively spliced form is induced in mouse lungs, spleen, and kidneys during LPS-induced pulmonary inflammation. Since the affected region is critical for ICAM-1 presentation, dimerization, and solubilization, this alternative splice variant may have unique physiological functions.

Alternative 5' splice site selection induced by heat shock

1994 ◽  
Vol 14 (1) ◽  
pp. 567-575
Author(s):  
H Takechi ◽  
N Hosokawa ◽  
K Hirayoshi ◽  
K Nagata
Keyword(s):  
Alternative Splicing ◽  
Heat Shock ◽  
Site Selection ◽  
Donor Site ◽  
Splice Donor Site ◽  
In Vivo Model ◽  
Recognition Mechanism ◽  
Amino Acid Analog ◽  
Alternatively Spliced

The mouse HSP47 gene consists of six exons separated by five introns. Three HSP47 cDNAs differing only in their 5' noncoding regions have been reported. One of these alternatively spliced mRNAs was detected only after heat shock, which caused an alternative 5' splice donor site selection. Other stress inducers, including an amino acid analog and sodium arsenite, had no effect on the alternative splicing. The alternatively spliced mRNA, which was 169 nucleotides longer in the 5' noncoding region compared to mRNA transcribed in non-heat shock conditions, was efficiently translated under heat shock conditions. This novel finding that alternative splicing is caused by artificial treatment like heat shock will provide a useful in vivo model for understanding the exon-intron recognition mechanism as well as heat shock-induced alterations in gene expression.

scholarly journals Alternative 5' splice site selection induced by heat shock.

1994 ◽  
Vol 14 (1) ◽  
pp. 567-575 ◽  
Author(s):  
H Takechi ◽  
N Hosokawa ◽  
K Hirayoshi ◽  
K Nagata
Keyword(s):  
Alternative Splicing ◽  
Heat Shock ◽  
Site Selection ◽  
Donor Site ◽  
Splice Donor Site ◽  
In Vivo Model ◽  
Recognition Mechanism ◽  
Amino Acid Analog ◽  
Alternatively Spliced

The mouse HSP47 gene consists of six exons separated by five introns. Three HSP47 cDNAs differing only in their 5' noncoding regions have been reported. One of these alternatively spliced mRNAs was detected only after heat shock, which caused an alternative 5' splice donor site selection. Other stress inducers, including an amino acid analog and sodium arsenite, had no effect on the alternative splicing. The alternatively spliced mRNA, which was 169 nucleotides longer in the 5' noncoding region compared to mRNA transcribed in non-heat shock conditions, was efficiently translated under heat shock conditions. This novel finding that alternative splicing is caused by artificial treatment like heat shock will provide a useful in vivo model for understanding the exon-intron recognition mechanism as well as heat shock-induced alterations in gene expression.

Abstract 3412: Molecular and Cellular Characterization of Splice Site Mutations in MYBPC3 -encoded Myosin Binding Protein C

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Jeanne L Theis ◽  
J M Bos ◽  
Steve R Ommen ◽  
Michael J Ackerman
Keyword(s):  
Splice Site ◽  
Protein C ◽  
Binding Protein ◽  
Donor Site ◽  
Splice Donor Site ◽  
Splice Donor ◽  
Myosin Binding Protein ◽  
Myosin Binding Protein C ◽  
Alternatively Spliced ◽  
Myosin Binding

Background: Mutations in MYBPC3 -encoded myosin binding protein C underlie the most common genotype in hypertrophic cardiomyopathy (HCM). Compared to most HCM-associated mutations which are primarily missense mutations, MYBPC3-HCM is often caused by insertions, deletions, nonsense mutations or mutations involving the canonical splice site. However, the effects of most mutations that prematurely truncate MYBPC3 are unknown. Access to cardiac specific mRNA and tissue has permitted a molecular and cellular elucidation of the consequences of two splice site mutations. Methods: Mutational analysis of MYBPC3 revealed 2 HCM-susceptibility mutations involving the splice donor sites of exons 7 and 30. Reverse transcription was performed on cDNA generated from RNA extracted from cardiac tissue obtained following surgical myectomy. Western blot analysis and immunohistochemistry (IHC) of the myofilaments were performed to further assess the impact of the splice-site mutations. Results: Both splice donor mutations produced abnormal RNA splicing of MYBPC3. The c.821+1 g>a mutation in the splice donor site of exon 7 generated two alternatively-spliced mutant transcripts resulting in two frame-shifted, premature truncations (H257 fs/37 and H257 fs/15). The c.3330+2 t>g mutation in exon 30’s splice donor site produced a single alternatively-spliced mutant transcript that translated into a frame-shifted premature truncation (V1063 fs/37). Expression levels of wild type myosin binding protein C were decreased in the myectomy specimen from the patient with the exon 7 splice donor site mutation. By IHC, the spatial organization of myosin binding protein C was disrupted severely in both patients. Conclusions: This study provides the first molecular and cellular characterization of HCM-causing mutations involving canonical splice-site motifs within the intron. Loss of function of the splice site resulted in exon skipping and generation of frame-shifted and prematurely truncated myosin binding protein C and disruption of the distribution and organization of myosin binding protein C in the heart tissue. This research has received full or partial funding support from the American Heart Association, AHA Midwest Affiliate (Illinois, Indiana, Iowa, Kansas, Michigan, Minnesota, Missouri, Nebraska, North Dakota, South Dakota & Wisconsin).

Exon recognition and nucleocytoplasmic partitioning determine AMPD1 alternative transcript production

1991 ◽  
Vol 11 (10) ◽  
pp. 5356-5363
Author(s):  
I Mineo ◽  
E W Holmes
Keyword(s):  
Rna Processing ◽  
Donor Site ◽  
Alternative Transcript ◽  
Primary Transcript ◽  
Exon 2 ◽  
Mature Mrna ◽  
Cell Biol ◽  
Ampd1 Gene ◽  
Exon 1 ◽  
Intron 2

Two mature transcripts are produced from the rat AMP deaminase 1 (AMPD1) gene, one that retains exon 2 and one from which exon 2 has been removed. The ratio of these two transcripts is controlled by stage-specific and tissue-specific signals (I. Mineo, P. R. H. Clarke, R. L. Sabina, and E. W. Holmes, Mol. Cell. Biol. 10:5271-5278, 1990; R. L. Sabina, N. Ogasawara, and E. W. Holmes, Mol. Cell. Biol. 9:2244-2246, 1989). By using transfection studies with native, mutant, and chimeric minigene constructs, two steps in RNA processing that determine the ratio of these two transcripts have been identified. The first step is recognition of this exon in the primary transcript. The primary transcript is subject to alternative splicing in which exon 2 is either recognized and thereby included in the mature mRNA or is ignored and retained in a composite intron containing intron 1-exon 2-intron 2. The following properties of the primary transcript influence exon recognition. (i) Exon 2 is intrinsically difficult to recognize, possibly because of its small size (only 12 bases) and/or a suboptimal 5' donor site at the exon 2-intron 2 boundary. (ii) Intron 2 plays a permissive role in recognition of exon 2 because it is removed at a relatively slow rate, presumably because of the suboptimal polypyrimidine tract in the putative 3' branch site. The second step in RNA processing that influences the ratio of mature transcripts produced from the AMPD1 gene occurs subsequent to the ligation of exon 2 to exon 1. An RNA intermediate, composed of exon 1-exon 2-intron 2-exon 3, is produced in the first processing step, but it is variably retained in the nucleus. Retention of this intermediate in the nucleus is associated with accumulation of the mature mRNA containing exon 2, while cytoplasmic escape of this intermediate is reactions, exon recognition and nucleocytoplasmic partitioning, determine the relative abundance of alternative mRNAs derived from the AMPD1 gene.

scholarly journals Exon recognition and nucleocytoplasmic partitioning determine AMPD1 alternative transcript production.

1991 ◽  
Vol 11 (10) ◽  
pp. 5356-5363 ◽  
Author(s):  
I Mineo ◽  
E W Holmes
Keyword(s):  
Rna Processing ◽  
Donor Site ◽  
Alternative Transcript ◽  
Primary Transcript ◽  
Exon 2 ◽  
Mature Mrna ◽  
Cell Biol ◽  
Ampd1 Gene ◽  
Exon 1 ◽  
Intron 2

Two mature transcripts are produced from the rat AMP deaminase 1 (AMPD1) gene, one that retains exon 2 and one from which exon 2 has been removed. The ratio of these two transcripts is controlled by stage-specific and tissue-specific signals (I. Mineo, P. R. H. Clarke, R. L. Sabina, and E. W. Holmes, Mol. Cell. Biol. 10:5271-5278, 1990; R. L. Sabina, N. Ogasawara, and E. W. Holmes, Mol. Cell. Biol. 9:2244-2246, 1989). By using transfection studies with native, mutant, and chimeric minigene constructs, two steps in RNA processing that determine the ratio of these two transcripts have been identified. The first step is recognition of this exon in the primary transcript. The primary transcript is subject to alternative splicing in which exon 2 is either recognized and thereby included in the mature mRNA or is ignored and retained in a composite intron containing intron 1-exon 2-intron 2. The following properties of the primary transcript influence exon recognition. (i) Exon 2 is intrinsically difficult to recognize, possibly because of its small size (only 12 bases) and/or a suboptimal 5' donor site at the exon 2-intron 2 boundary. (ii) Intron 2 plays a permissive role in recognition of exon 2 because it is removed at a relatively slow rate, presumably because of the suboptimal polypyrimidine tract in the putative 3' branch site. The second step in RNA processing that influences the ratio of mature transcripts produced from the AMPD1 gene occurs subsequent to the ligation of exon 2 to exon 1. An RNA intermediate, composed of exon 1-exon 2-intron 2-exon 3, is produced in the first processing step, but it is variably retained in the nucleus. Retention of this intermediate in the nucleus is associated with accumulation of the mature mRNA containing exon 2, while cytoplasmic escape of this intermediate is reactions, exon recognition and nucleocytoplasmic partitioning, determine the relative abundance of alternative mRNAs derived from the AMPD1 gene.

Sign in / Sign up

Export Citation Format

Share Document