Congenital end-plate acetylcholinesterase deficiency caused by a nonsense mutation and an A→G splice-donor-site mutation at position +3 of the collagenlike-tail-subunit gene (COLQ)

How does G at position +3 result in aberrant splicing?

Kinji Ohno, Joan M. Brengman, Kevin J. Felice, David R. Cornblath, Andrew G Engel

Research output: Contribution to journalArticle

71 Citations (Scopus)

Abstract

Congenital end-plate acetylcholinesterase (AChE) deficiency (CEAD), the cause of a disabling myasthenic syndrome, arises from defects in the COLQ gene, which encodes the AChE triple-helical collagenlike-tail subunit that anchors catalytic subunits of AChE to the synaptic basal lamina. Here we describe a patient with CEAD with a nonsense mutation (R315X) and a splice- donor-site mutation at position +3 of intron 16 (IVS16+3A→G) of COLQ. Because both A and G are consensus nucleotides at the +3 position of splice- donor sites, we constructed a minigene that spans exons 15-17 and harbors IVS16+3A→G for expression in COS cells. We found that the mutation causes skipping of exon 16. The mutant splice-donor site of intron 16 harbors five discordant nucleotides (at -3, -2, +3, +4, and +6) that do not base-pair with U1 small-nuclear RNA (snRNA), the molecule responsible for splice-donor-site recognition. Versions of the minigene harboring, at either +4 or +6, nucleotides complementary to U1 snRNA restore normal splicing. Analysis of 1,801 native splice-donor sites reveals that presence of a G nucleotide at +3 is associated with preferential usage, at positions +4 to +6, of nucleotides concordant to U1 snRNA. Analysis of 11 disease-associated IVS+3A→G mutations indicates that, on average, two of three nucleotides at positions +4 to +6 fail to base-pair, and that the nucleotide at +4 never base-pairs, with U1 snRNA. We conclude that, with G at +3, normal splicing generally depends on the concordance that residues at +4 to +6 have with U1 snRNA, but other cis- acting elements may also be important in assuring the fidelity of splicing.

Original languageEnglish (US)
Pages (from-to)635-644
Number of pages10
JournalAmerican Journal of Human Genetics
Volume65
Issue number3
DOIs
StatePublished - 1999

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RNA Splice Sites
Nonsense Codon
Acetylcholinesterase
Nucleotides
Mutation
Genes
Base Pairing
Introns
Exons
COS Cells
Muscle Weakness
Basement Membrane
Catalytic Domain
U1 small nuclear RNA

ASJC Scopus subject areas

  • Genetics

Cite this

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title = "Congenital end-plate acetylcholinesterase deficiency caused by a nonsense mutation and an A→G splice-donor-site mutation at position +3 of the collagenlike-tail-subunit gene (COLQ): How does G at position +3 result in aberrant splicing?",
abstract = "Congenital end-plate acetylcholinesterase (AChE) deficiency (CEAD), the cause of a disabling myasthenic syndrome, arises from defects in the COLQ gene, which encodes the AChE triple-helical collagenlike-tail subunit that anchors catalytic subunits of AChE to the synaptic basal lamina. Here we describe a patient with CEAD with a nonsense mutation (R315X) and a splice- donor-site mutation at position +3 of intron 16 (IVS16+3A→G) of COLQ. Because both A and G are consensus nucleotides at the +3 position of splice- donor sites, we constructed a minigene that spans exons 15-17 and harbors IVS16+3A→G for expression in COS cells. We found that the mutation causes skipping of exon 16. The mutant splice-donor site of intron 16 harbors five discordant nucleotides (at -3, -2, +3, +4, and +6) that do not base-pair with U1 small-nuclear RNA (snRNA), the molecule responsible for splice-donor-site recognition. Versions of the minigene harboring, at either +4 or +6, nucleotides complementary to U1 snRNA restore normal splicing. Analysis of 1,801 native splice-donor sites reveals that presence of a G nucleotide at +3 is associated with preferential usage, at positions +4 to +6, of nucleotides concordant to U1 snRNA. Analysis of 11 disease-associated IVS+3A→G mutations indicates that, on average, two of three nucleotides at positions +4 to +6 fail to base-pair, and that the nucleotide at +4 never base-pairs, with U1 snRNA. We conclude that, with G at +3, normal splicing generally depends on the concordance that residues at +4 to +6 have with U1 snRNA, but other cis- acting elements may also be important in assuring the fidelity of splicing.",
author = "Kinji Ohno and Brengman, {Joan M.} and Felice, {Kevin J.} and Cornblath, {David R.} and Engel, {Andrew G}",
year = "1999",
doi = "10.1086/302551",
language = "English (US)",
volume = "65",
pages = "635--644",
journal = "American Journal of Human Genetics",
issn = "0002-9297",
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T1 - Congenital end-plate acetylcholinesterase deficiency caused by a nonsense mutation and an A→G splice-donor-site mutation at position +3 of the collagenlike-tail-subunit gene (COLQ)

T2 - How does G at position +3 result in aberrant splicing?

AU - Ohno, Kinji

AU - Brengman, Joan M.

AU - Felice, Kevin J.

AU - Cornblath, David R.

AU - Engel, Andrew G

PY - 1999

Y1 - 1999

N2 - Congenital end-plate acetylcholinesterase (AChE) deficiency (CEAD), the cause of a disabling myasthenic syndrome, arises from defects in the COLQ gene, which encodes the AChE triple-helical collagenlike-tail subunit that anchors catalytic subunits of AChE to the synaptic basal lamina. Here we describe a patient with CEAD with a nonsense mutation (R315X) and a splice- donor-site mutation at position +3 of intron 16 (IVS16+3A→G) of COLQ. Because both A and G are consensus nucleotides at the +3 position of splice- donor sites, we constructed a minigene that spans exons 15-17 and harbors IVS16+3A→G for expression in COS cells. We found that the mutation causes skipping of exon 16. The mutant splice-donor site of intron 16 harbors five discordant nucleotides (at -3, -2, +3, +4, and +6) that do not base-pair with U1 small-nuclear RNA (snRNA), the molecule responsible for splice-donor-site recognition. Versions of the minigene harboring, at either +4 or +6, nucleotides complementary to U1 snRNA restore normal splicing. Analysis of 1,801 native splice-donor sites reveals that presence of a G nucleotide at +3 is associated with preferential usage, at positions +4 to +6, of nucleotides concordant to U1 snRNA. Analysis of 11 disease-associated IVS+3A→G mutations indicates that, on average, two of three nucleotides at positions +4 to +6 fail to base-pair, and that the nucleotide at +4 never base-pairs, with U1 snRNA. We conclude that, with G at +3, normal splicing generally depends on the concordance that residues at +4 to +6 have with U1 snRNA, but other cis- acting elements may also be important in assuring the fidelity of splicing.

AB - Congenital end-plate acetylcholinesterase (AChE) deficiency (CEAD), the cause of a disabling myasthenic syndrome, arises from defects in the COLQ gene, which encodes the AChE triple-helical collagenlike-tail subunit that anchors catalytic subunits of AChE to the synaptic basal lamina. Here we describe a patient with CEAD with a nonsense mutation (R315X) and a splice- donor-site mutation at position +3 of intron 16 (IVS16+3A→G) of COLQ. Because both A and G are consensus nucleotides at the +3 position of splice- donor sites, we constructed a minigene that spans exons 15-17 and harbors IVS16+3A→G for expression in COS cells. We found that the mutation causes skipping of exon 16. The mutant splice-donor site of intron 16 harbors five discordant nucleotides (at -3, -2, +3, +4, and +6) that do not base-pair with U1 small-nuclear RNA (snRNA), the molecule responsible for splice-donor-site recognition. Versions of the minigene harboring, at either +4 or +6, nucleotides complementary to U1 snRNA restore normal splicing. Analysis of 1,801 native splice-donor sites reveals that presence of a G nucleotide at +3 is associated with preferential usage, at positions +4 to +6, of nucleotides concordant to U1 snRNA. Analysis of 11 disease-associated IVS+3A→G mutations indicates that, on average, two of three nucleotides at positions +4 to +6 fail to base-pair, and that the nucleotide at +4 never base-pairs, with U1 snRNA. We conclude that, with G at +3, normal splicing generally depends on the concordance that residues at +4 to +6 have with U1 snRNA, but other cis- acting elements may also be important in assuring the fidelity of splicing.

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