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Synthesis of apoferritin in mouse peritoneal macrophages. Characterization of 20 S particles
Abstract:
Apoferritin particles were found in mouse peritoneal macrophages cultured in vitro. They were found as 20S particles in the "ribosomal fraction" of macrophages labeled with L-[14C]glutamic acid. Possibilities that they were breakdown products of ribosomes or of other well-known contaminants of the ribosomal fraction were excluded because they did not incorporate [5-3H]uridine. They were resistant to RNase and were relatively resistant to detergent. The antibody against horse spleen apoferritin precipitated about 70% of the particles in the 20S region, judging by measurement of radioactivity. On in vitro incubation with Fe2+ and suitable oxidizing agents the sedimentation coefficient of 80% of the 20S particles changed to about 60S, which corresponds to that of ferritin. SDS-polyacrylamide gel electrophoresis revealed the presence of subunit structures with the same molecular size as that of mouse liver apoferritin. Under the electron microscope, the particles appeared spherical with a relatively uniform diameter of about 130 A.
Insights
Mouse peritoneal macrophages contain apoferritin particles. These 20S particles transform into 60S ferritin upon iron addition, confirming their identity and function in iron storage.
Area of Science:
- Cell Biology
- Biochemistry
Background:
- Macrophages play a crucial role in iron homeostasis.
- Apoferritin is the protein shell of ferritin, involved in iron storage.
Purpose of the Study:
- To identify and characterize apoferritin particles in mouse peritoneal macrophages.
- To investigate the transformation of apoferritin to ferritin in vitro.
Main Methods:
- Isolation of 20S particles from macrophage ribosomal fractions labeled with L-[14C]glutamic acid.
- RNase resistance, detergent stability, antibody precipitation, and iron-induced sedimentation assays.
- SDS-polyacrylamide gel electrophoresis and electron microscopy for structural analysis.
Main Results:
- 20S apoferritin particles were identified in macrophages, distinct from ribosomes and other contaminants.
- Antibody precipitation confirmed the particles' identity as apoferritin.
- In vitro incubation with Fe2+ converted 80% of 20S particles to 60S ferritin.
- Electrophoresis and electron microscopy revealed subunit structure and spherical morphology consistent with apoferritin.
Conclusions:
- Mouse peritoneal macrophages synthesize and contain apoferritin particles.
- These particles can be converted to ferritin, indicating their role in cellular iron storage.