Diversity of guanylate cyclase-activating proteins (GCAPs) in teleost fish: characterization of three novel GCAPs (GCAP4, GCAP5, GCAP7) from zebrafish (Danio rerio) and prediction of eight GCAPs (GCAP1-8) in pufferfish (Fugu rubripes).
The guanylate cyclase-activating proteins (GCAPs) are Ca(2+)-binding proteins of the calmodulin (CaM) gene superfamily that function in the regulation of photoreceptor guanylate cyclases (GCs). In the mammalian retina, two GCAPs (GCAP 1-2) and two transmembrane GCs have been identified as part of a complex regulatory system responsive to fluctuating levels of free Ca(2+). A third GCAP, GCAP3, is expressed in human and zebrafish (Danio rerio) retinas, and a guanylate cyclase-inhibitory protein (GCIP) has been shown to be present in frog cones. To explore the diversity of GCAPs in more detail, we searched the pufferfish (Fugu rubripes) and zebrafish (Danio rerio) genomes for GCAP-related gene sequences (fuGCAPs and zGCAPs, respectively) and found that at least five additional GCAPs (GCAP4-8) are predicted to be present in these species. We identified genomic contigs encoding fuGCAPl-8, fuGCIP, zGCAPl-5, zGCAP7 and zGCIP. We describe cloning, expression and localization of three novel GCAPs present in the zebrafish retina (zGCAP4, zGCAP5, and zGCAP7). The results show that recombinant zGCAP4 stimulated bovine rod outer segment GC in a Ca(2+)-dependent manner. RT-PCR with zGCAP specific primers showed specific expression of zGCAPs and zGCIP in the retina, while zGCAPl mRNA is also present in the brain. In situ hybridization with anti-sense zGCAP4, zGCAP5 and zGCAP7 RNA showed exclusive expression in zebrafish cone photoreceptors. The presence of at least eight GCAP genes suggests an unexpected diversity within this subfamily of Ca(2+)-binding proteins in the teleost retina, and suggests additional functions for GCAPs apart from stimulation of GC. Based on genome searches and EST analyses, the mouse and human genomes do not harbor GCAP4-8 or GCIP genes.
Department of Ophthalmology, University of Washington, Seattle, WA 98195, USA.
Yoshikazu
Y
Imanishi
Department of Biology, University of Utah, Salt Lake City, UT 84112, USA.
Lili
L
Yang
Department of Ophthalmology, University of Washington, Seattle, WA 98195, USA.
Izabela
I
Sokal
International Institute of Molecular and Cell Biology, Warsaw, PL 02109, Poland.
Slawomir
S
Filipek
Department of Ophthalmology, University of Washington, Seattle, WA 98195, USA.|Department of Pharmacology, University of Washington, Seattle, WA 98195, USA.|Department of Chemistry, University of Washington, Seattle, WA 98195, USA.
Krzysztof
K
Palczewski
Department of Biology, University of Utah, Salt Lake City, UT 84112, USA.|Department of Ophthalmology, University of Utah, Salt Lake City, UT 84112, USA.|Department of Neurobiology and Anatomy, University of Utah, Salt Lake City, UT 84112, USA.
Wolfgang
W
Baehr
D000595:Amino Acid Sequence; D000818:Animals; D001482:Base Composition; D001483:Base Sequence; D002135:Calcium-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D016000:Cluster Analysis; D017931:DNA Primers; D030541:Databases, Genetic; D015870:Gene Expression; D014644:Genetic Variation; D051604:Guanylate Cyclase-Activating Proteins; D017403:In Situ Hybridization; D008957:Models, Genetic; D008969:Molecular Sequence Data; D010802:Phylogeny; D011487:Protein Conformation; D012160:Retina / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D023741:Takifugu / Q000235:genetics*; D015027:Zebrafish / Q000235:genetics*; D029961:Zebrafish Proteins / Q000235:genetics* / Q000378:metabolism
D002135:Calcium-Binding Proteins; D017931:DNA Primers; D051604:Guanylate Cyclase-Activating Proteins; D029961:Zebrafish Proteins; C499851:guca1d protein, zebrafish; C499852:guca1e protein, zebrafish; C499853:guca1g protein, zebrafish