Properties of protein kinase and adenylate cyclase-deficient variants of a macrophage-like cell line
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Abstract:
Stable variants of the macrophage-like cell line J774.2, defective in adenylate cyclase and protein kinase activities, were selected by cloning cells resistant to the growth-inhibitory effect of cholera toxin and 8-bromo-adenosine 3':5' cyclic monophosphoric acid (8 Br-cAMP), respectively. These variants were analyzed for their ability to respond to cyclic AMP-mediated enhancement of phagocytosis and cyclic AMP-mediated inhibition of plasminogen activator secretion and growtn. The adenylate cyclase variants were unaffected by cholera toxin but were sensitive to 8 Br-cAMP-mediated inhibition of plasminogen activator secretion and growth. One of these variants exhibited a defect in phagocytosis that could be corrected by 8 Br-cAMP. The protein kinase variants exhibited normal basal phagocytosis that could not be stimulated by either 8 Br-cAMP or cholera toxin; they were also insensitive to cyclic AMP-mediated inhibition of plasminogen activator secretion and growth. The studies demonstrate that the three effects of cyclic AMP in J774.2--inhibition of growth and plasminogen activator secretion, and enhancement of basal Fc-mediated phagocytosis--are mediated by a cyclic AMP-dependent portein kinase. The results support the usefulness of variants in cyclic nucleotide metabolism in understanding the regulation of differentiated cell function by cyclic AMP.
Insights
Researchers created cell variants to study cyclic AMP (adenosine monophosphate) functions. These variants confirmed that cyclic AMP regulates cell growth, plasminogen activator secretion, and phagocytosis via a specific protein kinase, highlighting its role in cell function.
Area of Science:
- Cell Biology
- Molecular Biology
- Immunology
Background:
- The macrophage-like cell line J774.2 exhibits responses modulated by cyclic AMP (adenosine monophosphate).
- Understanding the precise mechanisms of cyclic AMP signaling is crucial for deciphering cellular regulation.
Purpose of the Study:
- To investigate the role of cyclic AMP-dependent protein kinase in mediating cellular functions in J774.2 cells.
- To elucidate the specific pathways through which cyclic AMP influences phagocytosis, cell growth, and plasminogen activator secretion.
Main Methods:
- Selection and characterization of stable J774.2 cell variants deficient in adenylate cyclase or protein kinase activities.
- Analysis of variant responses to cholera toxin and 8-bromo-adenosine 3':5' cyclic monophosphoric acid (8 Br-cAMP).
- Assessment of phagocytosis, plasminogen activator secretion, and growth modulation by cyclic AMP.
Main Results:
- Adenylate cyclase variants were insensitive to cholera toxin but responsive to 8 Br-cAMP for some functions.
- Protein kinase variants showed defects in cyclic AMP-mediated enhancement of phagocytosis and inhibition of secretion and growth.
- Specific defects in phagocytosis were corrected by 8 Br-cAMP in one adenylate cyclase variant.
Conclusions:
- The findings demonstrate that cyclic AMP-dependent protein kinase mediates the effects of cyclic AMP on J774.2 cell growth, plasminogen activator secretion, and Fc-mediated phagocytosis.
- Cellular variants defective in cyclic nucleotide metabolism are valuable tools for studying the regulation of differentiated cell functions by cyclic AMP.