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Prosomes and their multicatalytic proteinase activity
1Institut Jacques Monod, CNRS, Université Paris 7, France.
Abstract:
Prosomes were first described as being mRNA-associated RNP (ribonucleoprotein) particles and subcomponents of repressed mRNPs (messenger ribonucleoprotein). We show here that prosomes isolated from translationally inactive mRNP have a protease activity identical to that described by others for the multicatalytic proteinase complex (MCP, 'proteasome'). By RNase or non-ionic detergent treatment, the MCP activity associated with repressed non-globin mRNP from avian erythroblasts, sedimenting at 35 S, could be quantitatively shifted on sucrose gradients to the 19-S sedimentation zone characteristic of prosomes, which were identified by monoclonal antibodies. The presence of small RNA in the enzymatic complex was shown by immunoprecipitation of the protease activity out of dissociated mRNP using a mixture of anti-prosome monoclonal antibodies; a set of small RNAs 80-120 nucleotides long was isolated from the immunoprecipitate. Furthermore, on CsCl gradients, colocalisation of the MCP activity with prosomal proteins and prosomal RNA was found, and no difference in the prosomal RNA pattern was observed whether the particles were fixed or not prior to centrifugation. These data indicate that the MCP activity is a property of prosomes, shown to be in part RNP and subcomplexes of in vivo untranslated mRNP. A hypothesis for the role of the prosome-MCP particles in maintaining homeostasis of specific protein levels is proposed.
Insights
Prosomes, identified as messenger ribonucleoprotein (mRNP) complexes, possess protease activity, functioning as multicatalytic proteinase complexes (MCPs). These prosome-MCP particles are implicated in regulating protein levels within cells.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Prosomes were initially characterized as messenger ribonucleoprotein (mRNA-associated RNP) particles.
- They are recognized as subcomponents of repressed messenger ribonucleoprotein (mRNP) complexes.
Purpose of the Study:
- To investigate the protease activity of prosomes isolated from translationally inactive mRNP.
- To determine if this protease activity is identical to the multicatalytic proteinase complex (MCP, or proteasome).
Main Methods:
- Isolation of prosomes from avian erythroblast repressed non-globin mRNP.
- Sucrose gradient sedimentation analysis following RNase or detergent treatment.
- Identification of prosomes using monoclonal antibodies.
- Immunoprecipitation of protease activity and isolation of associated small RNAs.
- Cesium chloride (CsCl) gradient centrifugation to assess colocalization.
Main Results:
- Prosomes isolated from translationally inactive mRNP exhibit protease activity identical to MCP.
- MCP activity associated with repressed mRNP can be shifted to the 19-S sedimentation zone characteristic of prosomes.
- Small RNAs (80-120 nucleotides) are associated with the prosome-MCP complex.
- MCP activity, prosomal proteins, and prosomal RNA colocalize on CsCl gradients.
Conclusions:
- The multicatalytic proteinase complex (MCP) activity is an intrinsic property of prosomes.
- Prosomes are ribonucleoprotein (RNP) particles and subcomplexes of in vivo untranslated mRNP.
- Prosome-MCP particles may play a role in maintaining cellular protein homeostasis.