RUI: Negative Transcriptional Regulation of the Inducible Interleukin-4 Gene

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9724032 Weiss For genes which exhibit strict tissue specific expression or are expressed only upon induction, the contribution by negative regulatory elements is particularly important. The objective of this research is to define the nuclear factors which interact with the negative regulatory element NEG-1 of the Interleukin-4 (IL-4) gene. IL-4 is produced only by mast cells and certain T cell subsets, and then only upon induction. Messenger RNA levels for IL-4 increase rapidly after induction and then decrease once the inducing stimulus is removed. This observation coupled with other data indicates that IL-4 production is under transcriptional control. Previous studies have demonstrated that there are several DNA regulatory elements within the 5' flanking region of this gene and that specific nuclear factors interact with these sequences to regulate the inducible transcription of IL-4. The recently defined DNA element NEG-1, (-326 to -302 relative to the transcription initiation start site) contributes to the down regulation of this gene. Preliminary results indicate that two sub-sequences (A and B) within NEG-1 interact co-operatively and that NEG-1 A forms a specific complex with mast cell nuclear factor(s) found in both unstimulated and stimulated cells. This research identify the nuclear factor(s) in direct contact with the A region of NEG-1, and will use heparin-sepharose and DNA affinity chromatography to purify enough factor to obtain sequence information. A cDNA library will be constructed and screened with degenerate oligonucleotides corresponding to the sequence information obtained. Positive clones will be subcloned, sequenced, and expressed; their NEG-1 A binding ability will be confirmed by EMSA. %%% For genes which exhibit strict tissue specific expression or are expressed only upon induction, the contribution by negative regulatory elements is particularly important. Interleukin-4 (IL-4) is produced only by mast cells and certain T cell subsets, and then only upon induction. The objective of this research is to define the nuclear factors which interact with the negative regulatory element NEG-1 of the IL-4 gene. ***