TY - JOUR
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.
N1 - Funding Information:
This work was supported by National Institutes of Health grant NS6277 (to A.G.E.) and Muscular Dystrophy Association grants (to A.G.E. and K.O.).
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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U2 - 10.1086/302551
DO - 10.1086/302551
M3 - Article
C2 - 10441569
AN - SCOPUS:0033362102
SN - 0002-9297
VL - 65
SP - 635
EP - 644
JO - American journal of human genetics
JF - American journal of human genetics
IS - 3
ER -