A Na+/Cl- -coupled GABA transporter, GAT-1, from Caenorhabditis elegans: structural and functional features, specific expression in GABA-ergic neurons, and involvement in muscle function.
GABA functions as an inhibitory neurotransmitter in body muscles and as an excitatory neurotransmitter in enteric muscles in Caenorhabditis elegans. Whereas many of the components of the GABA-ergic neurotransmission in this organism have been identified at the molecular and functional levels, no transporter specific for this neurotransmitter has been identified to date. Here we report on the cloning and functional characterization of a GABA transporter from C. elegans (ceGAT-1) and on the functional relevance of the transporter to the biology of body muscles and enteric muscles. ceGAT-1 is coded by snf-11 gene, a member of the sodium-dependent neurotransmitter symporter gene family in C. elegans. The cloned ceGAT-1 functions as a Na(+)/Cl(-)-coupled high-affinity transporter selective for GABA with a K(t) of approximately 15 microm. The Na(+):Cl(-):GABA stoichiometry for ceGAT-1-mediated transport process is 2:1:1. The transport process is electrogenic as evidenced from GABA-induced inward currents in Xenopus laevis oocytes that express ceGAT-1 heterologously. The transporter is expressed exclusively in GABA-ergic neurons and in two other additional neurons. We also investigated the functional relevance of ceGAT-1 to the biology of body muscles and enteric muscles by ceGAT-1-specific RNA interference (RNAi) in rrf-3 mutant, a strain of C. elegans in which neurons are not refractory to RNAi as in the wild type strain. The down-regulation of ceGAT-1 by RNAi leads to an interesting phenotype associated with altered function of body muscles (as evident from changes in thrashing frequency) and enteric muscles (as evident from the rates of defecation failure) and also with altered sensitivity to aldicarb-induced paralysis. These findings provide unequivocal evidence for a modulatory role of GABA and ceGAT-1 in the biology of cholinergic neurons and in the function of body muscles and enteric muscles in this organism.
Jiang
Guoliang
G
Department of Biochemistry and Molecular Biology and Institute of Molecular Medicine and Genetics, Medical College of Georgia, Augusta, Georgia 30912, USA.
Zhuang
Lina
L
Miyauchi
Seiji
S
Miyake
Katsuya
K
Fei
You-Jun
YJ
Ganapathy
Vadivel
V
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017173:Caenorhabditis elegans / Q000378:metabolism*; D029742:Caenorhabditis elegans Proteins; D003001:Cloning, Molecular; D017931:DNA Primers; D050485:GABA Plasma Membrane Transport Proteins; D007700:Kinetics; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009132:Muscles / Q000502:physiology*; D009474:Neurons / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D050495:Vesicular Inhibitory Amino Acid Transport Proteins; D014982:Xenopus laevis; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
D029742:Caenorhabditis elegans Proteins; D017931:DNA Primers; D050485:GABA Plasma Membrane Transport Proteins; D026901:Membrane Transport Proteins; D050495:Vesicular Inhibitory Amino Acid Transport Proteins; C493420:unc-47 protein, C elegans; D005680:gamma-Aminobutyric Acid