Distinct sequence elements involved in the glucocorticoid regulation of the mouse mammary tumor virus promoter identified by linker scanning mutagenesis.

摘要: In the proviral DNA of mouse mammary tumor virus (MMTV), sequences up to approximately equal 200 base-pairs from RNA start site are required for stimulation transcription by glucocorticoid hormones in cultured cells. A total 26 mutant plasmids with clustered point mutations or small deletions hormone control region MMTV long terminal repeat were constructed, linked coding portion Herpes simplex thymidine kinase gene, and introduced transfection into LTK- Transcription presence absence was quantified S1 nuclease protection assays. Our analysis revealed at least three elements that affect extent hormones: (1) a distal element, between -181 -172 initiation site. Linker scanning mutants this segment have reduction 20-fold response respect wild type. (2) An element around position -120, defined mutation 4 -121 -117, which causes fivefold reduction. (3) -78 -70, also roughly lower stimulation. The first two included areas been shown others interact vitro hormone-receptor complexes; last one overlaps binding nuclear protein factor. lacking all (-193 -70) is completely non-inducible glucocorticoids. Together earlier results obtained 5' deletion mutants, data show largest contribution stimulatory made however does require both more-proximal ones be maximal. one, proximal together produce residual order 5 10% type, while -70 alone ineffective. addition, we functional TATA homology maximum It appears transcriptional regulation achieved concerted action multiple sequence modules, not correspond receptor sites vitro.