Isolation and Regulation of the cGMP-lnhibited cAMP Phosphodiesterase in Human Erythroleukemia Cells

S B Sheth; K J Brennan; R Biradavolu; R W Colman

doi:10.1055/s-0038-1655924

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 1997; 77(01): 155-162
DOI: 10.1055/s-0038-1655924

Platelets

Schattauer GmbH Stuttgart

Isolation and Regulation of the cGMP-lnhibited cAMP Phosphodiesterase in Human Erythroleukemia Cells

S B Sheth

The Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, PA, USA

,

K J Brennan

The Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, PA, USA

,

R Biradavolu

The Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, PA, USA

,

R W Colman

The Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, PA, USA

› Author Affiliations

Abstract

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Summary

The predominant cAMP phosphodiesterase in human platelets is the low K_m cGMP-inhibited phosphodiesterase (PDE 3A). We have isolated native PDE3A from platelets and human erythroleukemia (HEL) cells and studied its kinetics. The platelet and HEL cell enzymes hydrolyze cAMP with K_m = 0.5 μM. Incubation of cell supernatant with cAMP dependent protein kinase resulted in a rapid increase in activity within minutes, which resulted from a 2-fold decrease in K_m with no increase in V_max. HEL cells grown for 24 h in the presence of 50 (iM forskolin, an adenylate cyclase activator, demonstrate further increase in PDE3A of 274% of control (p = 0.03). Cells incubated with forskolin and cycloheximide or actinomycin D demonstrated no increase suggesting that cAMP stimulates PDE3A synthesis by transcriptional regulation. The results indicate that cAMP affects both the short and long-term regulation of PDE3A. The latter effect may play a role in the developing hematopoietic cell and the cardiovascular system to regulate cAMP levels.

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References

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