Phares, W., Franza, B. R., Herr, W. (December 1992) The Kb Enhancer Motifs in Human-Immunodeficiency-Virus Type-1 and Simian Virus-40 Recognize Different Binding Activities in Human Jurkat and H9 T-Cells - Evidence for Nf-Kb-Independent Activation of the Kb Motif. Journal of Virology, 66 (12). pp. 7490-7498. ISSN 0022-538X
Abstract
The kappaB transcriptional enhancer motif, present in many viruses, is broadly active in many cell types. It is recognized by c-Rel/HIVEN86A in DNA affinity precipitation (DNAP) assays and by the Rel-related p50 and p65 subunits of the nuclear factor NF-kappaB in electrophoretic mobility shift assays (EMSA). We have analyzed activities that bind the human immunodeficiency virus type 1 and simian virus 40 kappaB motifs in two human leukemia cell lines, Jurkat and H9. In both DNAP and EMSA analyses of Jurkat cell extracts, we detected multiple kappaB motif-binding activities in addition to c-Rel/HIVEN86A and p50-p65 NF-kappaB. In Jurkat cell nuclear extracts, EMSA analysis revealed at least six specific DNA-protein complexes, of which one comigrated with the p50-p65 NF-kappaB complex. Formation of all six complexes was enhanced by stimulation of the cells with phorbol 12-myristate-13-acetate and phytohemagglutinin but was differentially affected by the salt concentration in the binding reaction and by the conditions of Jurkat cell growth. Nuclear extracts from both unstimulated and stimulated H9 cells revealed similar levels of five kappaB motif-specific complexes, all of which displayed mobilities distinct from those of the Jurkat cell complexes. Indeed, a complex corresponding to p50-p65 NF-kappaB was not detectable in nuclear extracts from unstimulated H9 cells although such a complex was apparent in nuclear extracts from stimulated H9 cells. In contrast to the inducibility of a p50-p65 NF-kappaB-like complex, transcriptional enhancers composed of multimerized kappaB motifs displayed similar high levels of activity in both the unstimulated and stimulated H9 cells. Thus, the activity of the kappaB motif in H9 cells corresponded to the abundance of the H9 cell-specific kappaB motif complexes and not to the levels of p50-p65 NF-kappaB complex. These results suggest that the broad activity of the kappaB enhancer element is not only due to the broadly distributed NF-kappaB activator but also to cell type-specific kappaB motif-binding activities.
| Item Type: | Paper |
|---|---|
| Subjects: | bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > transcription diseases & disorders > viral diseases > HIV diseases & disorders > viral diseases |
| CSHL Authors: | |
| Communities: | CSHL labs > Herr lab |
| Depositing User: | Matt Covey |
| Date: | December 1992 |
| Date Deposited: | 28 Sep 2015 20:00 |
| Last Modified: | 28 Sep 2015 20:00 |
| PMCID: | PMC240457 |
| Related URLs: | |
| URI: | https://repository.cshl.edu/id/eprint/31814 |
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