Receptor-evoked Cl- current in Xenopus oocytes is mediated through a beta-type phospholipase C. Cloning of a new form of the enzyme.
Xenopus oocytes exhibit a receptor-evoked Cl- current that is mediated through the activation of phospholipase C (PLC) and release of intracellular Ca2+. The identity of PLC(s) mediating this effect is unknown. We have cloned cDNAs encoding a new form of PLC-beta from a Xenopus oocyte cDNA library. The Xenopus PLC-beta has substantial (33-64%) homology with mammalian beta 1, beta 2, beta 3, and beta 4 phospholipase C and is closest to PLC-beta 3, with 64% identity and 80% similarity. Injection of antisense oligonucleotides to a specific region of Xenopus PLC-beta results in degradation of its mRNA and significantly reduces Cl- currents evoked by both endogenous angiotensin receptors and expressed mammalian alpha 1b-adrenergic receptors and M1-muscarinic receptors as compared to responses in sense oligonucleotide-injected oocytes. Inhibition of the M1-muscarinic response by antisense oligonucleotides was nonadditive with pertussis toxin inhibition. PLC antisense oligonucleotide-injected oocytes show Cl- current responses to IP3 that are indistinguishable from sense oligonucleotide-injected oocytes. Since the receptor responses are pertussis toxin-sensitive, we conclude that we have isolated a new form of PLC-beta involved in the pertussis toxin-sensitive receptor stimulation of the Ca2+ activated Cl- current in Xenopus oocytes.
Department of Pharmacology, Mount Sinai School of Medicine, City University of New York, New York 10029.
H W
HW
Ma
R D
RD
Blitzer
E C
EC
Healy
R T
RT
Premont
E M
EM
Landau
R
R
Iyengar
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D018118:Chloride Channels; D002712:Chlorides / Q000378:metabolism*; D003001:Cloning, Molecular; D004330:Drosophila / Q000201:enzymology / Q000235:genetics; D005260:Female; D015723:Gene Library; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D008322:Mammals; D008565:Membrane Proteins / Q000187:drug effects / Q000502:physiology*; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D009865:Oocytes / Q000187:drug effects / Q000201:enzymology* / Q000502:physiology; D037342:Pertussis Toxin; D012333:RNA, Messenger / Q000378:metabolism; D011942:Receptors, Adrenergic, alpha / Q000096:biosynthesis / Q000502:physiology*; D011945:Receptors, Angiotensin / Q000502:physiology*; D011976:Receptors, Muscarinic / Q000096:biosynthesis / Q000502:physiology*; D010738:Type C Phospholipases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology; D014981:Xenopus
D018118:Chloride Channels; D002712:Chlorides; D007527:Isoenzymes; D008565:Membrane Proteins; D009838:Oligodeoxyribonucleotides; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D011942:Receptors, Adrenergic, alpha; D011945:Receptors, Angiotensin; D011976:Receptors, Muscarinic; D010566:Virulence Factors, Bordetella; D037342:Pertussis Toxin; D010738:Type C Phospholipases