An Altered Intron Inhibits Synthesis of the Acetylcholine Receptor -Subunit in the Paralyzed Zebrafish Mutant nic1

Corresponding author: Monte Westerfield, Institute of Neuroscience, University of Oregon, Eugene, OR 97403, monte{at}uoneuro.uoregon.edu (E-mail).

Communicating editor: N. A. JENKINS

The acetylcholine receptor (AChR), an oligomeric protein composed of five subunits, is a component of the postsynaptic membrane at the vertebrate neuromuscular junction that plays a central role in synaptic transmission. The zebrafish mutation nic1 blocks the expression of functional and clustered nicotinic muscle AChRs. To understand the mechanisms underlying this lack of AChRs, we characterized the molecular defect in nic1 mutants. Our results suggest that the mutation affects the gene coding for the -subunit of the AChR. Southern blot hybridization and DNA sequence analyses showed that the nic1 AChR -subunit gene lacks part of intron 6 where the splicing branchpoint normally forms. Several lines of evidence suggest that this deletion blocks normal splicing; most nic1 -subunit mRNAs retain intron 6 and are larger and less abundant than wild-type, some nic1 -subunit mRNAs are internally deleted, and wild-type -subunit mRNA rescues nic1 mutant cells. The nic1 mutation reduces the size of an intron, which prevents efficient splicing of the pre-mRNA, thus blocking synthesis of the -subunit and assembly of AChRs. By this route, the nic1 mutation leads to paralysis.

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