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Defective induction of Jun and Fos-related proteins in phorbol ester-resistant EL4 mouse thymoma cells
D E Jensen1, R C Frankis, J J Sando
1Department of Pharmacology, University of Virginia, Charlottesville 22908.
Abstract:
Treatment of sensitive EL4 mouse thymoma cells with phorbol esters causes growth inhibition, adherence to substrate and production of several lymphokines including Interleukin 2. Resistant cells lack all of these responses. Since production of Interleukin 2 mRNA is dependent on protein synthesis, and the Interleukin 2 gene has a phorbol ester responsive element, we examined both cell lines for expression of the various Jun and Fos species which bind to this element. Phorbol ester induced c-fos, jun-B, and jun-D RNAs within 20 min in both cell lines. Fos-B was similarly induced in sensitive cells but induction was delayed and greatly enhanced in resistant cells. C-jun RNA induction was detected only in sensitive cells. Western analysis confirmed the induction of c-Jun and a Fos-related protein in sensitive cells only. Southern analysis indicated that both cell lines contain c-jun and fra-1 genes. These results suggest that defective induction of c-Jun and/or Fos-related proteins may contribute to the absence of phorbol ester-induced lymphokine production in resistant EL4 cells.
Insights
Sensitive EL4 cells produce Interleukin 2 (IL-2) when treated with phorbol esters, unlike resistant cells. This difference is linked to defective induction of c-Jun and Fos-related proteins in resistant cells.
Area of Science:
- Molecular Biology
- Immunology
- Cell Biology
Background:
- Phorbol esters induce growth inhibition, adherence, and lymphokine production, including Interleukin 2 (IL-2), in sensitive EL4 mouse thymoma cells.
- Resistant EL4 cells do not exhibit these responses to phorbol ester treatment.
- Interleukin 2 mRNA production is protein synthesis-dependent and regulated by a phorbol ester-responsive element in the IL-2 gene.
Purpose of the Study:
- To investigate the role of Jun and Fos protein species in the differential response of sensitive and resistant EL4 cells to phorbol esters.
- To determine if defects in the induction of specific Jun/Fos proteins contribute to the lack of IL-2 production in resistant cells.
Main Methods:
- Analysis of RNA expression (c-fos, jun-B, jun-D, Fos-B, c-jun) following phorbol ester treatment using Northern blot.
- Western blot analysis to confirm protein induction of c-Jun and Fos-related proteins.
- Southern blot analysis to assess the presence of c-jun and fra-1 genes.
Main Results:
- Phorbol esters induced c-fos, jun-B, and jun-D RNAs rapidly in both sensitive and resistant EL4 cells.
- Fos-B induction was similar in sensitive cells but delayed and enhanced in resistant cells.
- c-Jun RNA and protein induction, along with a Fos-related protein, were detected only in sensitive cells.
- Both cell lines possessed the c-jun and fra-1 genes.
Conclusions:
- Defective induction of c-Jun and/or Fos-related proteins in resistant EL4 cells likely contributes to their inability to produce Interleukin 2 in response to phorbol esters.
- This study highlights the critical role of specific transcription factor induction in mediating cellular responses to external stimuli.