A novel H(+)-coupled oligopeptide transporter (OPT3) from Caenorhabditis elegans with a predominant function as a H(+) channel and an exclusive expression in neurons.
We have cloned and functionally characterized a novel, neuron-specific, H(+)-coupled oligopeptide transporter (OPT3) from Caenorhabditis elegans that functions predominantly as a H(+) channel. The opt3 gene is approximately 4.4 kilobases long and consists of 13 exons. The cDNA codes for a protein of 701 amino acids with 11 putative transmembrane domains. When expressed in mammalian cells and in Xenopus laevis oocytes, OPT3 cDNA induces H(+)-coupled transport of the dipeptide glycylsarcosine. Electrophysiological studies of the transport function of OPT3 in Xenopus oocytes show that this transporter, although capable of mediating H(+)-coupled peptide transport, functions predominantly as a H(+) channel. The H(+) channel activity of OPT3 is approximately 3-4-fold greater than the H(+)/peptide cotransport activity as determined by measurements of H(+) gradient-induced inward currents in the absence and presence of the dipeptide using the two-microelectrode voltage clamp technique. A downhill influx of H(+) was accompanied by a large intracellular acidification as evidenced from the changes in intracellular pH using an ion-selective microelectrode. The H(+) channel activity exhibits a K(0.5)(H) of 1.0 microM at a membrane potential of -50 mV. At the level of primary structure, OPT3 has moderate homology with OPT1 and OPT2, two other H(+)-coupled oligopeptide transporters previously cloned from C. elegans. Expression studies using the opt3::gfp fusion constructs in transgenic C. elegans demonstrate that opt3 gene is exclusively expressed in neurons. OPT3 may play an important physiological role as a pH balancer in the maintenance of H(+) homeostasis in C. elegans.
Fei
Y J
YJ
Department of Biochemistry and Molecular Biology, Medical College of Georgia, Augusta, Georgia 30912, USA. yjfei@mail.mcg.edu
Romero
M F
MF
Krause
M
M
Liu
J C
JC
Huang
W
W
Ganapathy
V
V
Leibach
F H
FH
D000143:Acids; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017173:Caenorhabditis elegans / Q000378:metabolism*; D029742:Caenorhabditis elegans Proteins; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D017931:DNA Primers; D005786:Gene Expression Regulation; D006863:Hydrogen-Ion Concentration; D007473:Ion Channels / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism*; D010455:Peptides / Q000378:metabolism; D011522:Protons; D027981:Symporters; D014982:Xenopus laevis
D000143:Acids; D029742:Caenorhabditis elegans Proteins; D002352:Carrier Proteins; D017931:DNA Primers; D007473:Ion Channels; D009419:Nerve Tissue Proteins; D010455:Peptides; D011522:Protons; D027981:Symporters; C407085:pept-3 protein, C elegans