Mutations away from splice site recognition sequences might cis-modulate alternative splicing of goat alpha s1-casein transcripts. Structural organization of the relevant gene.

The human alpha-tropomyosin gene hTMnm has two mutually exclusive versions of exon 5 (NM and SK), one of which is expressed specifically in skeletal muscle (exon SK). A minigene construct expresses only the nonmuscle (NM) isoform when transfected into COS-1 cells and both forms when transfected into myoblasts. Twenty-four mutants were produced to determine why the SK exon is not expressed in COS cells. The results showed that exons NM and SK are not in competition for splicing to the flanking exons and that there is no intrinsic barrier to splicing between the exons. Instead, exon SK is skipped whenever there are flanking introns. Splicing of exon SK was induced when the branch site sequence 70 nucleotides upstream of the exon was mutated to resemble the consensus and when the extremities of the exon itself were changed to the corresponding NM sequence. Precise swaps of the NM and SK exon sequences showed that the exon sequence effect was dominant to that of intron sequences. The mechanism of regulation appears to be unlike that of other tropomyosin genes. We propose that exclusion of exon SK arises because its 3' splicing signals are weak and are prevented by an exon-specific repressor from competing for splice site recognition.

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Alternative splicing of a human alpha-tropomyosin muscle-specific exon: identification of determining sequences

Molecular and Cellular Biology ◽

10.1128/mcb.12.9.3872-3882.1992 ◽

1992 ◽

Vol 12 (9) ◽

pp. 3872-3882

Author(s):

I R Graham ◽

M Hamshere ◽

I C Eperon

Keyword(s):

Skeletal Muscle ◽

Alternative Splicing ◽

Splice Site ◽

Sequence Effect ◽

Cos Cells ◽

Branch Site ◽

Exon Sequence ◽

Site Recognition

The human alpha-tropomyosin gene hTMnm has two mutually exclusive versions of exon 5 (NM and SK), one of which is expressed specifically in skeletal muscle (exon SK). A minigene construct expresses only the nonmuscle (NM) isoform when transfected into COS-1 cells and both forms when transfected into myoblasts. Twenty-four mutants were produced to determine why the SK exon is not expressed in COS cells. The results showed that exons NM and SK are not in competition for splicing to the flanking exons and that there is no intrinsic barrier to splicing between the exons. Instead, exon SK is skipped whenever there are flanking introns. Splicing of exon SK was induced when the branch site sequence 70 nucleotides upstream of the exon was mutated to resemble the consensus and when the extremities of the exon itself were changed to the corresponding NM sequence. Precise swaps of the NM and SK exon sequences showed that the exon sequence effect was dominant to that of intron sequences. The mechanism of regulation appears to be unlike that of other tropomyosin genes. We propose that exclusion of exon SK arises because its 3' splicing signals are weak and are prevented by an exon-specific repressor from competing for splice site recognition.

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Spliceosome Assembly Pathways for Different Types of Alternative Splicing Converge during Commitment to Splice Site Pairing in the A Complex

Molecular and Cellular Biology ◽

10.1128/mcb.01071-08 ◽

2008 ◽

Vol 29 (4) ◽

pp. 1072-1082 ◽

Cited By ~ 15

Author(s):

Matthew V. Kotlajich ◽

Tara L. Crabb ◽

Klemens J. Hertel

Keyword(s):

Alternative Splicing ◽

Branch Point ◽

Splice Site ◽

Atp Hydrolysis ◽

Splice Sites ◽

Mrna Isoforms ◽

Exon Definition ◽

U2 Snrnp ◽

Kinetic Trap ◽

Site Recognition

ABSTRACT Differential splice site pairing establishes alternative splicing patterns resulting in the generation of multiple mRNA isoforms. This process is carried out by the spliceosome, which is activated by a series of sequential structural rearrangements of its five core snRNPs. To determine when splice sites become functionally paired, we carried out a series of kinetic trap experiments using pre-mRNAs that undergo alternative 5′ splice site selection or alternative exon inclusion. We show that commitment to splice site pairing in both cases occurs in the A complex, which is characterized by the ATP-dependent association of the U2 snRNP with the branch point. Interestingly, the timing of splice site pairing is independent of the intron or exon definition modes of splice site recognition. Using the ATP analog ATPγS, we showed that ATP hydrolysis is required for splice site pairing independent from U2 snRNP binding to the pre-mRNA. These results identify the A complex as the spliceosomal assembly step dedicated to splice site pairing and suggest that ATP hydrolysis locks splice sites into a splicing pattern after stable U2 snRNP association to the branch point.

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NDNF variants are rare in patients with congenital hypogonadotropic hypogonadism

Human Genome Variation ◽

10.1038/s41439-021-00137-x ◽

2021 ◽

Vol 8 (1) ◽

Author(s):

Satoshi Tamaoka ◽

Erina Suzuki ◽

Atsushi Hattori ◽

Tsutomu Ogata ◽

Maki Fukami ◽

...

Keyword(s):

Splice Site ◽

Genetic Heterogeneity ◽

Hypogonadotropic Hypogonadism ◽

Causative Gene ◽

Synonymous Substitutions ◽

Congenital Hypogonadotropic Hypogonadism ◽

Site Recognition

AbstractAlthough NDNF was recently reported as a novel causative gene for congenital hypogonadotropic hypogonadism (CHH), this conclusion has yet to be validated. In this study, we sequenced NDNF in 61 Japanese CHH patients. No variants, except for nine synonymous substitutions that appear to have no effect on splice-site recognition, were identified in NDNF coding exons or flanking intronic sequences. These results indicate the rarity of NDNF variants in CHH patients and highlight the genetic heterogeneity of CHH.

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Functional properties of p54, a novel SR protein active in constitutive and alternative splicing.

Molecular and Cellular Biology ◽

10.1128/mcb.16.10.5400 ◽

1996 ◽

Vol 16 (10) ◽

pp. 5400-5408 ◽

Cited By ~ 53

Author(s):

W J Zhang ◽

J Y Wu

Keyword(s):

Alternative Splicing ◽

Splice Site ◽

Sr Proteins ◽

Dependent Manner ◽

Sr Protein ◽

Protein Protein Interaction ◽

Small Nuclear Ribonucleoprotein ◽

S100 Extract ◽

U2 Auxiliary Factor ◽

Auxiliary Factor

The p54 protein was previously identified by its reactivity with an autoantiserum. We report here that p54 is a new member of the SR family of splicing factors, as judged from its structural, antigenic, and functional characteristics. Consistent with its identification as an SR protein, p54 can function as a constitutive splicing factor in complementing splicing-deficient HeLa cell S100 extract. However, p54 also shows properties distinct from those of other SR family members, p54 can directly interact with the 65-kDa subunit of U2 auxiliary factor (U2AF65), a protein associated with the 3' splice site. In addition, p54 interacts with other SR proteins but does not interact with the U1 small nuclear ribonucleoprotein U1-70K or the 35-kDa subunit of U2 auxiliary factor (U2AF35). This protein-protein interaction profile is different from those of prototypical SR proteins SC35 and ASF/SF2, both of which interact with U1-70K and U2AF35 but not with U2AF65. p54 promotes the use of the distal 5' splice site in E1A pre-mRNA alternative splicing, while the same site is suppressed by ASF/SF2 and SC35. These findings and the differential tissue distribution of p54 suggest that this novel SR protein may participate in regulation of alternative splicing in a tissue- and substrate-dependent manner.

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