Joseph A. Beavo

Regulation of Cyclic Nucleotides as Signal Transducers:

Many hormones, drugs and other agents modify physiological processes by causing changes in the steady state levels of cAMP and cGMP. One of the major mechanisms by which the levels of these nucleotide second messengers are controlled in the cell is by alteration in the activity of the specific cyclic nucleotide phosphodiesterases (PDEs) that control their degradation. The signaling pathways regulated by these PDEs include such well known processes as the transduction of photon capture in the outer segmentof a photoreceptor to a changes in neurotransmitter release from the inner segment of this neuron. They also include less well known pathways such as regulation of aldosterone production by atrial natriuretic peptide and regulation of platelet aggregation by endothelial relaxation factor. Drugs that selectively inhibit individual PDE isozymes have a wide variety of different effects on an animals suggesting specific roles for most of the different PDEs.

Current evidence indicates that there are several related but unique gene families that code for different phosphodiesterases and that each of these families contain more than one gene. Furthermore, each gene product is differentially spliced in differenttissues to yield different isozymes. A major focus of this laboratory has been to determine the molecular nature for the normal control of these isoenzymes. Many of the current projects involve approaches to determine which cell types contain what isozyme, and how that particular isozyme contributes to the phenotype of the cell. In addition, the recent isolation of cDNAs for many of the isozymes has allowed series of structure/function studies to be initiated. Finally, several of these isozymesare regulated by phosphorylation/dephosphorylation mechanisms. Studies are underway to determine how and to what extent this type of regulation is important to the physiological functions of the cells in which the phosphodiesterases are expressed.

A wide variety of experimental approaches are employed. They vary from basic enzymology studies with extracts of tissues to refined protein chemistry approaches using pure proteins obtained from overexpressing the individual phosphodiesterase cDNAs. Bothin vivo and in vitro techniques are utilized. In all cases, emphasis is placed on how and why each isozyme functions as it does in a tissue. Several students are also using isolated cell culture models to test for roles of the PDEs in such diverse processes as synaptic plasticity and water and salt reabsorbtion. In these studies, traditional approaches such as use of isozyme specific inhibitors of individual PDEs as well as more recent techniques of using antisense RNA, dominant negative mutants and gene disruption are being utilized.