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Rat liver nuclerar protein kinases
Canadian Journal of Biochemistry
|March 1, 1975
Summary
Cyclic AMP-dependent histone kinase is primarily a cytoplasmic enzyme, not nuclear. Nuclear casein kinases NI and NII exhibit distinct chromatin affinities and behaviors, with NII readily dissociating and NI requiring higher salt concentrations for extraction.
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- Nuclear enzyme activity is crucial for cellular regulation.
- Distinguishing between nuclear and cytoplasmic enzyme localization is essential for understanding cellular processes.
- Cyclic AMP-dependent protein kinases play significant roles in signal transduction pathways.
Purpose of the Study:
- To investigate the localization and properties of cyclic AMP-dependent histone kinase in isolated nuclei.
- To characterize the distinct behaviors and chromatin affinities of nuclear casein kinases NI and NII.
Main Methods:
- Isolation of liver nuclei using two different methods.
- Assay of cyclic AMP-dependent histone kinase and binding activity.
- Enzyme activity measurements for lactate dehydrogenase, glutamate dehydrogenase, and glucose-6-phosphatase.
- Sucrose density gradient centrifugation of nuclear extracts in varying salt concentrations.
- Differential salt extraction of nuclear casein kinases from chromatin.
Main Results:
- Washing isolated nuclei removed most cyclic AMP-dependent histone kinase and binding activity, indicating cytoplasmic localization.
- Nuclear cyclic AMP-dependent histone kinase activity represented a very small fraction of total cytoplasmic activity.
- Casein kinases NI and NII were separated by sucrose density gradients, showing distinct sedimentation coefficients (3.0 S and 593 S, respectively) at 0.5 M NaCl.
- Casein kinase NII dissociated easily in hypotonic solutions, while NI required sequential extractions with increasing salt concentrations (0.14 M, 0.5 M, 1.0 M NaCl) for complete removal from chromatin.
Conclusions:
- Cyclic AMP-dependent histone kinase is predominantly a cytoplasmic enzyme, with minimal association with isolated nuclei.
- Nuclear casein kinases NI and NII are distinct entities with differing affinities for chromatin, influencing their extraction patterns.
- The reversible aggregation-disaggregation of casein kinase NII impacts its behavior in nuclear extracts.