Search PubMed⌕ Search

Biomedical subjects

R J Ho

Publications and source records attributed to R J Ho.

At least 73 records · Page 4Linked to original sources
PubMed1978

Epinephrine-induced elevation of guanosine 3':5'-cyclic monophosphate in isolated fat cells of rat.

The effects of epinephrine (as low as 0.1 muM) on guanosine 3':5'-cyclic monophosphate (cGMP) and adenosine 3':5'-cyclic monophosphate (cAMP) in isolated fat cells were examined. Epinephrine increased both cGMP and cAMP levels, with the elevation of cAMP preceding the rise of cGMP. Maximal elevation was obtained with 1 muM epinephrine for each nucleotide. The increase in content of cGMP and cAMP due to epinephrine was completely blocked by a beta-adrenergic antagonist (5 muM propranolol). Phentolamine (10-100 muM), an alpha-adrenergic antagonist, enhanced the response to epinephrine resulting in elevation of cAMP levels, whereas a high concentration (100 muM) of phentolamine suppressed the elevation of cGMP. The ability of epinephrine to increase cGMP and cAMP levels was markedly diminished by "feedback regulator" partially purified from the incubation mixtures of isolated fat cells exposed to epinephrine [Ho, R.J. & Sutherland, E. W. (1971) J. Biol. Chem. 246, 6822-6827], whereas an increase in cGMP, but not cAMP, levels was observed in isolated fat cells incubated with "feedback regulator" alone (without epinephrine). These observations suggest the possibility that the epinephrine-induced elevation of cGMP levels in isolated fat cells might be mediated by an increase in formation of intracellular "feedback regulator" due to an elevation of cAMP by epinephrine.

Adipose Tissue↗
PubMed1977

Modulation of protein phosphorylation by a factor purified from adipocytes.

1. A factor which modulates the activity of cyclic AMP-dependent protein kinase copurifies from rat adipocytes with an inhibitor of adenylate cyclase. Purification and stability studies suggest that both effects reside in a single factor previously referred to as a feedback regulator. 2. The magnitude and direction of the feedback regulator effect on cyclic AMP-dependent protein kinase activity was dependent on the concentration of feedback regulator and the concentration and type of protein substrate. Using histone type IIA as substrate, feedback regulator was inhibitory at low histone concentrations and stimulatory at high concentrations. Preincubation of protein kinase with feedback regulator resulted in inhibition at all histone concentrations. With some protein substrates, e.g. histone f2b and casein, inhibition was observed at all histone concentrations. 3. The stimulation of histone type IIA phosphorylation resulted from an increased V with no effect on either the apparent Ka for cyclic AMP or the Km for ATP. Time course studies suggest that feedback regulator increased the rate of phosphorylation without increasing the total number of phosphorylation sites. Increased histone phosphorylation was observed regardless of whether the cyclic AMP-dependent protein kinase was peak I or peak II (off Deae-cellulose), isolated from bovine or rabbit skeletal muscle or rat heart. A small stimulation was observed using cyclic GMP-dependent protein kinase. 4. These results indicate that feedback regulator can inhibit or stimulate protein kinase, an effect which is probably substrate directed, and depends on the reaction conditions. Whether feedback regulator modulated protein phosphorylation in vivo in addition to its inhibition of adenylate cyclase is unknown. However, stimulation of protein kinase activity in the presence of cyclic AMP is a valuable and rapid assay for monitoring feedback regulator fractions during purification procedures.

Adipose Tissue↗
PubMed1975

Action of feedback regulator on adenylate cyclase.

A factor [the feedback regulator (FR)] formed by adipocytes after the stimulation of a cAMP raising hormone has been found to be a potent inhibitor of membrane-bound adenylate cyclase [EC 4.6.1.1.; ATP pyrophosphate-lyase (cyclizing)]. In a standard assay system using rat adipocyte plasma membrane as the source of adenylate cyclase, the FR inhibited adenylate cyclase by lowering the Vmax without affecting the apparent Km for ATP (0.3-0.6 mM). The apparent Ka for epinephrine (5-6 muM) was also not affected by FR. The inhibitory action of FR was partially countered by Mg2+ ions. An increase in phosphorylation of plasma membrane was observed when FR was present in the incubation system. The concentration required for a 50% inhibition was four times higher when adenosine 5-(beta,gamma-imino) triphosphate [AMP-P(NH)P] replaced ATP as the substrate for adenylate cyclase, implying that adenylate cyclase was inactivated by phosphorylation caused by FR. Increase in FR inhibition obtained by adding low concentrations of adenosine 5-(alpha,beta-methylene) triphosphate or ATP to AMP-(NH)P as the substrate supports this view. The inhibitory action was reversible. These results are consistent with the previously reported phenomena that (1) the undue to the formation of FR, and (2) the recovery of responsiveness of the stimulated cells by washing the cells with regular buffer medium is a result of the removal of FR. The hormone-initiated biphasic curve of cAMP levels in adipocytes is believed to be due to the negative feedback action of FR on adenylate cyclase. The mechanism of action of FR on inhibition of adenylate cyclase appears to be related to the phosphorylation of certain membrane components.

Adenosine Triphosphate↗
PubMed1975

Inhibition of cyclic nucleotide phosphodiesterase activity by an endogenous factor.

Cyclic nucleotide phosphodiesterase activity of several tissues of rat is inhibited by an endogenous factor isolated from rat adipocytes following exposure of these cells to agents that raise intracellular cyclic AMP levels. The inhibitory action was demonstrated with varying cAMP concentrations from 0.1-400 muM. Enzyme from 10,000 X g supernatant of epididymal adipose tissue was inhibited approximately 2-3 fold more than the plasma membrane of adipocytes by a given concentration of the feedback regulator. Kinetic analysis of cAMP phosphodiesterase of plasma membrane showed that feedback regulator (8.8 U/ml) inhibited the Vmax 48%. The maximum inhibition of phosphodiesterase by feedback regulator (20 U/ml) was about 80%. The apparent Km for cAMP was increased. The ability of phosphodiesterase from several tissues of rat (10,000 X g supernatant) to hydrolyze cAMP and cGMP was tested. Feedback regulator inhibited cGMP hydrolysis in cardiac muscle and 5 other tissues 23-92% more than it inhibited the hydrolysis of cAMP. The physiological significance of this inhibitory effect can begin to be clarified when the feedback regulator is purified to homogeneity and characterized.

3',5'-Cyclic-AMP Phosphodiesterases↗