Differential distribution of the G protein gamma3 subunit in the developing zebrafish nervous system.
G proteins play an essential role in the transduction and propagation of extracellular signals across the plasma membrane. It was once thought that the G protein alpha subunit was the sole regulator of intracellular molecules. The G protein betagamma complex is now recognized as participating in many signaling events. While screening a zebrafish cDNA library to identify members of the protein 4.1 superfamily (Kelly, G.M., Reversade, B., Biochem. Cell Biol. 75 (1997), 623), we fortuitously identified a clone that encodes a zebrafish G protein gamma subunit. The 666 nucleotides of the zebrafish G protein gamma subunit cDNA encodes a polypeptide of 75 amino acids with high degree of homology to human, bovine, rat and mouse gamma subunits. BLAST search analysis of GenBank revealed that the zebrafish gamma subunit is 93% identical and 97% similar to the mammalian gamma3 subunit. The gamma3 gene was mapped to the zebrafish linkage group 21, approximately 10.76 cRays from bf, a gene with sequence homology to the human properdin factor gene. RT-PCR and in situ hybridization analyses first detected gamma3 mRNA during late somitogenesis, where it was expressed preferentially in the Vth cranial nerve, the forebrain and in ventrolateral regions of the mid- and hindbrain including the spinal cord. The ability of the zebrafish gamma3 subunit to form a signaling heterodimeric complex with a beta subunit was tested using a human beta2 subunit. The gamma3 formed a heterodimer with beta2 and the complex was capable of binding calmodulin in a calcium-dependent manner. Overexpression of the beta2gamma3 complex in zebrafish embryos lead to the loss of dorsoanterior structures and heart defects, possibly owing to an up-regulation of mitogen-activated protein kinase activity and/or decline in protein kinase A signaling. Together, these data imply that a betagamma heterodimer plays a role in signal transduction events involving G protein coupled receptors and that these events occur in specific regions in the nervous system of the developing zebrafish.
Kelly
G M
GM
Department of Zoology, Molecular Genetics Unit, University of Western Ontario, ON, N6A 5B7, London, Canada. gkelly@julian.uwo.ca
Vanderbeld
B
B
Krawetz
R
R
Mangos
S
S
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001921:Brain / Q000196:embryology; D001923:Brain Chemistry; D002147:Calmodulin / Q000032:analysis; D018076:DNA, Complementary / Q000235:genetics; D019281:Dimerization; D044387:GTP-Binding Protein beta Subunits; D044388:GTP-Binding Protein gamma Subunits; D018507:Gene Expression Regulation, Developmental; D015723:Gene Library; D020962:Heterotrimeric GTP-Binding Proteins / Q000032:analysis* / Q000096:biosynthesis / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000032:analysis* / Q000096:biosynthesis / Q000235:genetics; D009420:Nervous System / Q000737:chemistry* / Q000196:embryology; D016372:RNA, Antisense / Q000032:analysis; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D015027:Zebrafish / Q000196:embryology*
D002147:Calmodulin; D018076:DNA, Complementary; C118981:G-protein Beta gamma; D044387:GTP-Binding Protein beta Subunits; D044388:GTP-Binding Protein gamma Subunits; D009419:Nerve Tissue Proteins; D016372:RNA, Antisense; D012333:RNA, Messenger; D020962:Heterotrimeric GTP-Binding Proteins