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

9724032 Weiss对于表现出严格的组织特异性表达或仅在诱导下表达的基因，负调控元件的贡献尤为重要。本研究的目的是确定与白细胞介素-4 （IL-4）基因负调控元件NEG-1相互作用的核因子。IL-4仅由肥大细胞和某些T细胞亚群产生，然后仅在诱导下产生。IL-4的信使RNA水平在诱导后迅速上升，一旦诱导刺激被移除，信使RNA水平就会下降。这一观察结果与其他数据相结合，表明IL-4的产生受转录控制。先前的研究表明，在该基因的5'侧区有几个DNA调控元件，并且特定的核因子与这些序列相互作用以调节IL-4的诱导转录。最近确定的DNA元件NEG-1（相对于转录起始位点-326至-302）有助于该基因的下调。初步结果表明，NEG-1的两个亚序列（A和B）相互作用，NEG-1 A与肥大细胞核因子(s)形成特异性复合物，在未受刺激和受刺激的细胞中都存在。本研究确定了与NEG-1的A区直接接触的核因子，并将利用肝素-sepharose和DNA亲和层析纯化足够的因子以获得序列信息。构建cDNA文库，筛选与序列信息相对应的简并寡核苷酸。阳性克隆将进行亚克隆、测序和表达；它们的NEG-1 A结合能力将被EMSA证实。对于表现出严格的组织特异性表达或仅在诱导下表达的基因，负调控元件的贡献尤为重要。白细胞介素-4 （IL-4）仅由肥大细胞和某些T细胞亚群产生，然后仅在诱导下产生。本研究的目的是确定与IL-4基因负调控元件NEG-1相互作用的核因子。＊＊＊

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. ***