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Lipopolysaccharide-mediated induction of RNA polymerase I activity and amount in murine B lymphocytes
Abstract:
The effect of lipopolysaccharide on RNA polymerase I activity in primary cultures of murine B lymphocytes has been examined. In cells treated with mitogen for 48 h, the activity of RNA polymerase I was approximately 15 times greater than in control cells. In situ localization of RNA polymerase I using indirect immunofluorescence indicated that there was at least a 10-fold increase in the amount of this enzyme associated with nucleoli of 48 h mitogen-treated cells relative to control cells. Immunoblotting experiments demonstrated a similar increase in the concentration of the 190-kDa subunit bound to DNA; the concentrations of the other polymerase I-associated polypeptides did not correlate with rRNA synthesis. Assuming 1 mol of the 190-kDa polypeptide/mol of polymerase I, it was estimated that 2,300 and 30,000 molecules of enzyme were associated with rDNA in the unstimulated and stimulated B cell, respectively. Thus, an increased cellular concentration of the 190-kDa subunit of RNA polymerase I and its association with ribosomal DNA may be a crucial step in rRNA synthesis.
Insights
Lipopolysaccharide stimulation significantly boosts RNA polymerase I activity in B lymphocytes, increasing enzyme association with ribosomal DNA. This suggests a key role for the 190-kDa subunit in ribosomal RNA synthesis.
Area of Science:
- Immunology
- Molecular Biology
- Biochemistry
Background:
- Lipopolysaccharide (LPS) is a potent immune stimulant.
- RNA polymerase I (Pol I) is responsible for ribosomal RNA (rRNA) synthesis.
- B lymphocyte activation involves significant changes in gene expression.
Purpose of the Study:
- To investigate the effect of lipopolysaccharide on RNA polymerase I activity in murine B lymphocytes.
- To determine the changes in Pol I localization and subunit concentration during B cell activation.
Main Methods:
- Primary cultures of murine B lymphocytes were treated with mitogen (lipopolysaccharide).
- RNA polymerase I activity was measured.
- In situ immunofluorescence was used to localize Pol I.
- Immunoblotting was performed to quantify Pol I subunits.
Main Results:
- Mitogen treatment for 48 hours increased RNA polymerase I activity approximately 15-fold.
- A 10-fold increase in Pol I associated with nucleoli was observed in stimulated cells.
- Immunoblotting revealed a corresponding increase in the 190-kDa subunit concentration bound to DNA.
Conclusions:
- Increased cellular concentration of the 190-kDa subunit of RNA polymerase I is crucial for enhanced rRNA synthesis.
- The association of Pol I with ribosomal DNA is a key regulatory step in stimulated B cells.