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Activation of translationally inactive lipoxygenase mRNP particles from rabbit reticulocytes
1Institute of Biochemistry, Humboldt University, Berlin, GDR.
Abstract:
The free cytoplasmic mRNP fraction of rabbit reticulocytes consists mainly of globin mRNP particles but contains substantial amounts of lipoxygenase mRNPs. mRNPs were isolated by affinity chromatography on oligo(dT)cellulose. The lipoxygenase specific mRNPs are in size between 30 and 80 S in sucrose density gradients as judged by Northern blot analysis with a specific lipoxygenase-plasmid probe and are translationally inactive in cell-free protein synthesis. Inhibition of lipoxygenase mRNP translation can be abolished by treatment of the particles with 0.65-0.8 M KCI which leads to a dissociation of masking proteins. Lipoxygenase mRNPs prepared from young reticulocytes can be activated in vitro by incubation with cytoplasm of more mature reticulocytes but not with cytoplasm of erythrocytes. Preliminary experiments indicate that Ca2+ ions may be involved in lipoxygenase mRNP activation.
Insights
Rabbit reticulocyte mRNPs contain lipoxygenase mRNA, which is translationally inactive. This inactivity can be reversed by removing masking proteins or through cytoplasmic factors, possibly involving calcium ions.
Area of Science:
- Molecular Biology
- Gene Expression Regulation
Background:
- Reticulocytes contain messenger ribonucleoprotein (mRNP) particles.
- The cytoplasmic mRNP fraction includes globin and lipoxygenase mRNPs.
Purpose of the Study:
- To investigate the translational status and regulation of lipoxygenase mRNPs in rabbit reticulocytes.
Main Methods:
- Isolation of mRNPs using affinity chromatography on oligo(dT)cellulose.
- Size determination via sucrose density gradients and Northern blot analysis.
- Assessment of translational activity in cell-free protein synthesis systems.
Main Results:
- Lipoxygenase mRNPs are translationally inactive and range from 30-80S.
- Treatment with high salt concentrations (0.65-0.8 M KCl) dissociates masking proteins, abolishing translational inhibition.
- mRNPs from young reticulocytes can be activated in vitro by mature reticulocyte cytoplasm.
Conclusions:
- Lipoxygenase mRNP translation is regulated by masking proteins.
- Cytoplasmic factors, potentially including calcium ions, play a role in activating these mRNPs.