There are five major families of cyclic nucleotide phosphodiesterases (PDEs) with distinct physicochemical, structural and kinetic properties. One class, the so-called Type III or CGMP-inhibited low Km CAMP PDE (CGI PDE), is characterized by its high affinity for CAMP and CGMP and specific inhibition of its CAMP hydrolysis by CGMP and positive inotropic and vasodilatory agents (amrinone, milrinone, cilostamide...). The CGI PDEs are important in the regulation of myocardial contractility, vascular tonus, platelet aggregation and antilipolytic action of insulin. At the present time little is known about the relationships and structural/functional similarities between CGI PDEs from different tissues. Using partial information from platelet CGI PDE protein, oligonucleotides were synthesized and used for screening of a human heart CDNA library and for PCR amplification and screening a rat adipose tissue CDNA library. Two positive CDNA clones (RcGIP1 and RcGIP2) were isolated from rat adipose tissue CDNA libraries and sequenced (Taira, M. et. al, submitted). Northern analysis indicated different tissue distributions for the two CGI PDE. RcGIP2 CDNA hybridized more with rat heart than rat adipose tissue RNA and not with adipocyte RNA. Using RcGIP2 CDNA probes, we screened a rat adipose tissue CDNA library. One of several positive clones (2-12-1; 7,1 kb) was isolated and sequenced. Its nucleotide sequence exhibited more than 90% identity to human heart CGI PDE nucleotide sequence, especially in the conserved domain and putative phosphorylation sites. This clone most likely represents the rat homolog of human heart (HCAR). Northern blot analysis with 2-12-1 CDNA showed strong hybridization with rat heart, aorta and lung RNA, and only weakly with brain RNA. Whether these CGI PDEs are specific for cardiac or cardiovascular tissues is not certain, since PCR amplification of human aortic CDNA and human liver CDNA with non-degenerate primers revealed the presence of partial human cardiac CGI PDE sequence. Diversity within the CGI PDE gene family will be approached by PCR amplification with degenerate primers of aortic and smooth muscle CDNA.
