Cloning and characterization of the choline acetyltransferase structural gene (cha-1) from C. elegans

Abstract

We have cloned the cha-1 gene from Caenorhabditis elegans using the method of transposon tagging, cha-1 is the structural gene for ChAT, the enzyme that synthesizes ACh. Sequence analysis of cDNAs predicts a protein of 71.5 kDa; comparison of the deduced amino acid sequence with ChAT sequences from other species confirms that cha-1 encodes ChAT. Comparison of cDNA and genomic sequences reveals that transcription is from right to left on the genetic map, and that some of the transcripts may result from trans-splicing of the 22-base spliced leader SL 1. The cha-1 gene is organized into 11 exons. The first exon contains only untranslated sequences, and is followed by an extremely long intron. The coding sequence of the cha-1 transcript is disrupted by mutations in the cha-1 gene. We have determined the sites of four transposon insertions and the end-points of two deletions that lead to the cha-1 mutant phenotype; one of the deletions appears to eliminate gene function completely. Comparison of the Drosophila, rat, and C. elegans genes reveals conserved motifs and conserved intron sites.

Choline Transport and de novo Choline Synthesis Support Acetylcholine Biosynthesis in Caenorhabditis elegans Cholinergic Neurons

“…Animals homozygous for this mutation are viable, and their growth and development appear to be normal, although they are slightly long and thin. Since cholinergic neurotransmission is essential for viability in C. elegans (Alfonso et al 1993(Alfonso et al , 1994, we conclude that the CHO-1 protein is not essential for cholinergic function.…”

“…The relatively mild phenotype of cho-1 mutants was therefore surprising, since cholinergic neurotransmission is essential for viability in C. elegans (Alfonso et al 1993(Alfonso et al , 1994. Clearly, in the absence of cho-1 function, choline for ACh synthesis must be supplied in other ways, either through other transporters, or by de novo synthesis.…”

“…The mild phenotypes of cho-1 null mutants are somewhat surprising since null mutations in other cholinergic genes in C. elegans, such as choline acetyltransferase (cha-1) or , result in early larval lethality (Alfonso et al 1993(Alfonso et al , 1994. Clearly, cho-1 function is not essential for cholinergic signaling in C. elegans.…”

TMC-1 attenuates C. elegans development and sexual behaviour in a chemically defined food environment

“…3d). eat-2 encodes an ionotropic ACh receptor that is expressed in the MC-pharyngeal muscle neuron junction 30 ; thus, an eat-2 null mutation blocks cholinergic transmission from the MC to the pharyngeal muscles 31 ; cha-1 encodes a choline acetyltransferase that synthesizes the neurotransmitter ACh 32 and a cha-1 mutation results in reduced ACh. To test if tmc-1 and eat-2 act in the same pathway, we made an eat-2(ad1113); tmc-1(ok1859) double mutant and found that 39% (n ¼ 102) develop into adults at 4 d.p.h.…”

Genetic Interactions Between UNC-17/VAChT and a Novel Transmembrane Protein in Caenorhabditis elegans

“…Following transmitter release, cholinergic signaling is terminated primarily through the action of acetylcholinesterase (AChE), which hydrolyses the released ACh into choline and acetyl-CoA. In the nematode Caenorhabditis elegans, the ChAT and VAChT proteins are encoded by the cha-1 and unc-17 genes, respectively (Alfonso et al , 1994b. Mutations that functionally eliminate either protein are lethal , while reduction-of-function mutations result in animals that are small, slow growing, uncoordinated, and resistant to AChE inhibitors such as aldicarb .…”