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?

Am J Hum Genet. 1999 Sep;65(3):635-44. doi: 10.1086/302551.


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.

Publication types

  • Case Reports
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Acetylcholinesterase / deficiency*
  • Acetylcholinesterase / genetics*
  • Acetylcholinesterase / metabolism
  • Alternative Splicing / genetics*
  • Animals
  • Base Pairing
  • Base Sequence
  • COS Cells
  • Collagen*
  • DNA Mutational Analysis
  • Exons / genetics
  • Female
  • Gene Expression
  • Humans
  • Introns / genetics
  • Male
  • Middle Aged
  • Motor Endplate / enzymology*
  • Motor Endplate / physiopathology
  • Muscle Proteins*
  • Mutation*
  • Pedigree
  • RNA, Messenger / analysis
  • RNA, Messenger / genetics
  • RNA, Small Nuclear / genetics
  • Reverse Transcriptase Polymerase Chain Reaction
  • Transfection


  • Muscle Proteins
  • RNA, Messenger
  • RNA, Small Nuclear
  • Collagen
  • Acetylcholinesterase
  • COLQ protein, human