An abnormal ketamine response in mutants defective in the ryanodine receptor gene ryr-1 (unc-68) of Caenorhabditis elegans.
To characterize excitation-contraction coupling in Caenorhabditis elegans, we applied two approaches. First, we isolated a mutant having abnormal responses to ketamine, an anesthetic in vertebrates. The novel mutation unc-68(kh30) (isolated as kra-1(kh30)), exhibited strict ketamine-dependent convulsions followed by paralysis. Second, we cloned the C. elegans ryanodine receptor gene ryr-1 that is located near the center of chromosome V. ryr-1 consists of 46 exons, which encode a predicted protein of 5071 amino acid residues that is homologous to Drosophila and vertebrate ryanodine receptors. ryr-1 promoter/lacZ plasmids were expressed in body-wall and pharyngeal muscles. Non-muscle cell expression may be seen with a truncated promoter. In addition, we show that the unc-68/kra-1(kh30) mutation is a Ser1444 Asn substitution at a putative protein kinase C phosphorylation site in ryr-1, and that unc-68(e540) contains a splice acceptor mutation that creates a premature stop codon in the ryr-1 gene. We confirmed that unc-68(e540) is a mutation in ryr-1 by injecting the complete ryr-1 gene into unc-68(e540) animals and recovering wild-type progeny. Results presented here will be useful in studying the structure and function of ryanodine receptors in excitation-contraction coupling and in understanding the evolution of ryanodine receptor tissue specificity.
Sakube
Y
Y
Department of Biology, Faculty of Science, Okayama University, Japan.
Ando
H
H
Kagawa
H
H
D000595:Amino Acid Sequence; D000778:Anesthetics, Dissociative / Q000494:pharmacology*; D000818:Animals; D030801:Animals, Genetically Modified; D001224:Aspartic Acid / Q000494:pharmacology; D017173:Caenorhabditis elegans / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D015220:Calcium Channels / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D015870:Gene Expression; D017238:Genes, Helminth / Q000235:genetics*; D007649:Ketamine / Q000494:pharmacology*; D008969:Molecular Sequence Data; D009119:Muscle Contraction / Q000187:drug effects; D009124:Muscle Proteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D016202:N-Methylaspartate / Q000494:pharmacology; D009928:Organ Specificity; D010766:Phosphorylation; D016194:Receptors, N-Methyl-D-Aspartate / Q000819:agonists / Q000037:antagonists & inhibitors; D011993:Recombinant Fusion Proteins / Q000032:analysis; D019837:Ryanodine Receptor Calcium Release Channel; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D000778:Anesthetics, Dissociative; D015220:Calcium Channels; D009124:Muscle Proteins; D016194:Receptors, N-Methyl-D-Aspartate; D011993:Recombinant Fusion Proteins; D019837:Ryanodine Receptor Calcium Release Channel; D001224:Aspartic Acid; D016202:N-Methylaspartate; D007649:Ketamine