Molecular comparison of rat cyclic nucleotide phosphodiesterase 8 family: unique expression of PDE8B in rat brain☆
Introduction
Cyclic nucleotides, cAMP and cGMP, which are produced in response to extracellular stimulation through activation of adenylyl and guanylyl cyclases, work as second messengers regulating many cellular functions in various tissues. Cyclic nucleotide phosphodiesterases (PDEs) play a role in elimination of this signaling through hydrolysis of cyclic nucleotides. In mammalians, PDEs have been classified into 11 families (PDEs 1–11) based on their amino acid sequence homology, enzymatic characteristics and inhibitor sensitivity profiles Beavo, 1995, Conti and Jin, 1999, Fujishige et al., 1999, Hayashi et al., 1998, Houslay, 2001, Soderling et al., 1998a, Soderling et al., 1998b, Soderling and Beavo, 2000, Yuasa et al., 2000. The phylogenic tree analysis indicated that PDEs are classified into two groups; GAF-PDE families containing two GAF (for cGMP binding and stimulated phosphodiesterases, Anabaena adenyl cyclases and Escherichia coli FhlA) domains (Aravind and Ponting, 1997) in their N-terminal regions and non-GAF-PDE families lacking the domain (Yuasa et al., 2001). The former families include PDE2, PDE5, PDE6, PDE10 and PDE11. Half of PDE families such as PDE1, PDE3, PDE4, PDE6, PDE7 and PDE8 are composed of subfamily genes encoding highly similar but distinct products. In addition, mostly each PDE gene produces several splice variants. They show unique tissue expression, subcellular localization, alternative enzymatic regulation, or unique protein–protein interaction, and some of them are subject to transcriptional regulation (e.g., Bolger et al., 1997, Han et al., 1997, O'Connell et al., 1996, Sette et al., 1994, Swinnen et al., 1991, Yuasa et al., 2000). In some cases, difference in their expression levels among animal species has also been reported (e.g., Engels et al., 1994, Yuasa et al., 2001). Thus, the PDE superfamily is composed of more than 60 kinds of transcripts produced from 21 genes. PDEs 8–11 are identified using an approach with bioinformatics. Compared with classical PDEs such as PDEs 1–6, information of these recently discovered PDEs is poor.
In humans, PDE8 family is composed of two isogenes, PDE8A and PDE8B. Thus far, PDE8A cDNAs have been isolated from humans and mice Fisher et al., 1998, Soderling et al., 1998a, Wang et al., 2001. A full-length cDNA sequence and gene organization of human PDE8B have been reported by Hayashi et al. (2002) and Gamanuma et al. (2003). Very recently, we have documented comparison of enzymatic characteristics of human PDE8A1 and PDE8B1, which are a major full-length form (Gamanuma et al., 2003). Human PDE8A1 and PDE8B1 are 68% identical overall, and 80% identical in the PDE catalytic domain. PDE8A1 and PDE8B1 are cAMP-specific PDEs with Km values of 40 and 101 nM, respectively, and are insensitive to 3-isobutyl-1-methylxanthine and rolipram. Human PDE8A1 transcripts are observed in testis, spleen, colon, small intestine, ovary, placenta and kidney in decreasing order (Wang et al., 2001). The mouse PDE8A1, which is 79% identical to human PDE8A1 at the amino acid level, is expressed in several tissues as testis>eye>skeletal muscle>heart>embryo 7 days>kidney>ovary>brain (Soderling et al., 1998a). Expression levels of PDE8A transcripts were different in several tissues such as spleen, skeletal muscle, heart and liver between humans and mice. Human PDE8B exhibits quite unique tissue-specific expression pattern, which is confined to the thyroid gland. It is significant to explore tissue expression pattern of PDE8B transcripts in other mammalians from the view of understanding a physiological role of the enzyme. However, any other mammalian orthologues of PDE8B than humans have not been identified yet.
Here we demonstrate cDNA sequences encoding rat PDE8A and PDE8B. Analysis of tissue expression patterns of these transcripts in rats reveals unique tissue expression patterns of PDE8 orthologues. Detailed expression of rat PDE8B transcripts in the brain is investigated by Northern blot and in situ hybridization analyses. These findings contribute to our better understanding of physiological functions of PDE8 proteins.
Section snippets
Materials
E. coli DH5α (Sambrook et al., 1989) was used as a host for plasmid construction. Restriction endonucleases, DNA-modifying enzymes, LA Taq and Ex Taq were obtained from Takara Bio (Kyoto, Japan). Marathon-Ready cDNA (rat testis and brain), Advantage 2 Polymerase Mix, Rat Multiple Tissue Northern blot, Mouse RNA Master Blot and ExpressHyb were purchased from Clontech (Palo Alto, CA). The cloning vectors pGEM-T Easy and pBluescriptII KS(+) were from Promega (Madison, WI) and Stratagene (La Jolla,
cDNA cloning of rodent PDE8
Fig. 1A shows cDNA fragments coding for the rat PDE8A1 and PDE8B1, which were obtained by a combination of PCR, 3′-RACE and database search as described in Materials and methods. An N-terminal sequence of rat PDE8A1, which was similar to the corresponding region of human PDE8A1, was found in rat EST databases using the 5′-sequence carried by pPDER8A2 (accession no. BE110619). A full-length cDNA of rat PDE8A1 of 2.6 kb was generated by PCR using primer sets based on the 5′- and 3′-sequences of
Discussion
Characteristic structural features of PDE8 proteins are the presence of the PAS and REC domains in their N-terminal parts. The PAS domain is observed in the N-terminal part of many proteins involved in the signal transduction. The REC domain is implicated in receiving the signal from sensor proteins in bacterial two-component systems, and sensor kinase-catalyzed aspartyl phosphorylation of the domain leads to regulation of the effector domain, which is covalently linked with the REC domain Pao
Acknowledgements
The authors thank Dr. Fujimura and Ms. Kurabe for their technical assistance and kind advice for in situ hybridization analysis.
References (36)
- et al.
Basic local alignment search tool
J. Mol. Biol.
(1990) - et al.
The GAF domain: an evolutionary link between diverse phototransducing proteins
Trends Biochem. Sci.
(1997) - et al.
The molecular biology of cyclic nucleotide phosphodiesterases
Prog. Nucleic Acid Res. Mol. Biol.
(1999) - et al.
Expression and regulation of human and rat phosphodiesterase type IV isogenes
FEBS Lett.
(1994) - et al.
Isolation and characterization of PDE8A, a novel human cAMP-specific phosphodiesterase
Biochem. Biophys. Res. Commun.
(1998) - et al.
Cloning and characterization of a novel human phosphodiesterase that hydrolyzes both cAMP and cGMP (PDE10A)
J. Biol. Chem.
(1999) - et al.
Localization of 63-kDa calmodulin-stimulated phosphodiesterase mRNA in the rat brain by in situ hybridization histochemistry
Brain Res., Mol. Brain Res.
(1994) - et al.
Comparison of enzymatic characterization and gene organization of cyclic nucleotide phosphodiesterase 8 (PDE8) family in humans
Cell. Signal.
(2003) - et al.
Alternative splicing of the high affinity cAMP-specific phosphodiesterase (PDE7A) mRNA in human skeletal muscle and heart
J. Biol. Chem.
(1997) - et al.
Molecular cloning and characterization of human PDE8B, a novel thyroid-specific isozyme of 3′,5′-cyclic nucleotide phosphodiesterase
Biochem. Biophys. Res. Commun.
(1998)
Genomic organization, chromosomal localization, and alternative splicing of the human phosphodiesterase 8B gene
Biochem. Biophys. Res. Commun.
PDE4 cAMP-specific phosphodiesterases
Prog. Nucleic Acid Res. Mol. Biol.
A cyclic GMP-stimulated cyclic nucleotide phosphodiesterase gene is highly expressed in the limbic system of the rat brain
Neuroscience
The ratPDE3/IVd phosphodiesterase gene codes for multiple proteins differentially activated by cAMP-dependent protein kinase
J. Biol. Chem.
Regulation of cAMP and cGMP signaling: new phosphodiesterases and new functions
Curr. Opin. Cell Biol.
Identification and characterization of a novel family of cyclic nucleotide phosphodiesterases
J. Biol. Chem.
Properties and hormonal regulation of two structurally related cAMP phosphodiesterases from the rat Sertoli cell
J. Biol. Chem.
The RdeA–RegA system, a eukaryotic phospho-relay controlling cAMP breakdown
J. Biol. Chem.
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The nucleotide and deduced amino acid sequences reported in this paper have been deposited with the DDBJ database under accession nos. AB092696 and AB092697.
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Both authors contributed equally to this work.