RNA associated with murine intracisternal type A particles codes for the main particle protein
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Abstract:
Intracisternal type A particles were isolated from MOPC-104E myeloma grown subcutaneously and from N 4 neuroblastoma cells in culture. Polyadenylated RNA was prepared from the particles and tested in a cell-free translation system derived from rabbit reticulocytes. RNA from the two sources directed the synthesis of multiple polypeptides with similar distributions of electrophoretic mobilities in sodium dodecyl sulfate-containing polyacrylamide gels, including one conponent of the same size as the major A-particle structural protein (73,000 daltons). Analysis of the RNAs by electrophoresis in methyl mercury-containing agarose gels revealed a 35S component common to A-particles from both cell types. This was a major component of the N4 preparations, whereas a 28S species predominated in the case of MOPC-104E. These two RNAs (35S from N4 cells and 28S from MOPC-104E), when isolated on isokinetic sucrose gradients, each directed the synthesis of a 73,000-molecular-weight polypeptide that comigrated on gels with authentic A-particle structural protein. Idnetity of the cell-free product was confirmed by two-dimensional analysis of the [35S]methionine-labeled tryptic peptides. The N4 RNA preparations also contained a major32S component which did not code effectively for the A-particle structural protein.
Insights
Researchers isolated intracisternal type A particles from myeloma and neuroblastoma cells. Their RNA directed the synthesis of a 73,000-dalton protein, confirming its role as the major structural component.
Area of Science:
- Molecular Biology
- Virology
- Cell Biology
Background:
- Intracisternal type A particles (IAPs) are retroviral elements found in various mouse tissues.
- Understanding the genetic material and protein synthesis of IAPs is crucial for studying their role in cellular processes and potential oncogenesis.
Purpose of the Study:
- To characterize the polyadenylated RNA of IAPs isolated from MOPC-104E myeloma and N4 neuroblastoma cells.
- To determine if this RNA directs the synthesis of the major structural protein of IAPs in a cell-free system.
Main Methods:
- Isolation of intracisternal type A particles from MOPC-104E myeloma and N4 neuroblastoma cells.
- Preparation and analysis of polyadenylated RNA using methyl mercury-agarose gel electrophoresis and isokinetic sucrose gradients.
- Cell-free translation of isolated RNAs using a rabbit reticulocyte system.
- Analysis of synthesized polypeptides by SDS-polyacrylamide gel electrophoresis and two-dimensional peptide mapping.
Main Results:
- RNA from both IAP sources directed the synthesis of multiple polypeptides, including one of 73,000 daltons, the size of the major structural protein.
- A common 35S RNA component was identified in both N4 and MOPC-104E IAPs, although a 28S species predominated in MOPC-104E.
- Isolated 35S (from N4) and 28S (from MOPC-104E) RNAs specifically directed the synthesis of the 73,000-dalton polypeptide, confirmed by peptide analysis.
Conclusions:
- The polyadenylated RNA from intracisternal type A particles contains the coding information for the major structural protein.
- Variations in RNA species (35S vs. 28S) may exist between different IAP sources but can both produce the structural protein.