MYELOPEROXIDASE EXPRESSION IN DISEASE STATES

疾病状态下的髓过氧化物酶表达

• 批准号: 2895801
• 负责人: WANDA F REYNOLDS
• 金额: $ 26.41万
• 依托单位: SIDNEY KIMMEL CANCER CENTER
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-09-01 至 2002-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2895801
• 关键词: DNA binding protein; DNA directed RNA polymerase; Xenopus; alternatives to animals in research; binding proteins; chromatin; conformation; developmental genetics; eukaryote; gene expression; genetic promoter element; genetic transcription; mutant; nucleic acid sequence; protein structure

Objective. The long term goal of the proposed research is to understand the molecular mechanisms by which Class III genes are differently regulated. During the previous grant period, we demonstrated that sequence differences preceeding the somatic and oocyte 5S promoters effect the relative activities of these genes in vitro and in vivo. This modulatory effect is only observed under conditions whereby the somatic 5S gene has a large (100-200 fold) transcriptional advantage over oocyte 5S. We have shown that these sequence differences increase the affinity for a common transcription factor and thereby aid in stable complex formation. We have further demonstrated that of the common factors IIIA, B, and C, the affinity of TFIIIA is not affected by these sequence differences. This finding is significant since it suggests that the differential expression of these genes is mediated not only by the 5S specific factor TFIIIA, but also through preferential association of IIIB with the somatic 5S gene. We now wish to determine which factor(s) preferentially interacts with the somatic 5S gene as a result of these sequence differences. We also wish to determine why these sequence differences effect gene activity only in whole oocyte S150 extracts in vitro (not in nuclear extracts) and at certain developmental stages in vivo. Is this indicative of stage specific forms of the polymerase III factors? In a related finding, we have shown that two other Class III genes, tDNAmet and OAX, are also differentially expressed in vitro in whole oocyte and cocyte nuclear extracts. Most interestingly, these DNAs compete with 5S DNA for factors in the nuclear extracts but not in the whole oocyte extract. We wish to determine if this finding reflects multiple gene specific forms of the polymerase III factors in the whole oocyte S150 extract. The specific aims of this proposal are: 1. to determine which factor in the S150 extract preferentially interacts with the somatic 5S gene as a result of the sequence differences preceding the promoter. 2. to investigate why these sequence differences provide a transcriptional advantage in the oocyte S150 extract but not in nuclear extracts. 3. to more accurately determine which sequence differences are most significant to this effect. 4. to investigate why E190 and tDNAmet genes are differentially expressed in the S150 and nuclear extracts. 5. to investigate why E190 and tDNAmet do not compete with 5S DNA in the S150 but do compete in the nuclear extract. Do these findings reflect differences in the factor populations? 6. to investigate why 5S, E190, and tDNAmet have similar activities in a heterologous extract (HeLa), but radically different activities in the homologous extracts.

客观的。 拟议研究的长期目标是 了解III类基因的分子机制 受不同的监管。 在上一个赠款期间，我们证明了该顺序 在体细胞和卵母细胞5S启动子效应之前的差异 这些基因在体外和体内的相对活性。 这 仅在条件下观察到调节作用 体细胞5s基因具有大（100-200倍）的转录 优于卵母细胞5s的优势。 我们已经证明了这些序列 差异增加了对公共转录因子的亲和力 从而有助于稳定的复合形成。 我们还有更多 证明了共同因素IIIA，B和C， TFIIIA的亲和力不受这些序列差异的影响。 这一发现很重要，因为它表明差异 这些基因的表达不仅是由5S特异性介导的 因子tfiiia，也通过优先IIIB的优先关联 与体细胞5S基因。 我们现在希望确定哪个因素 优先与体细胞5S基因相互作用。 这些序列差异。 我们也希望确定为什么 序列差异仅在整个卵母细胞S150中影响基因活性 体外提取物（不在核提取物中）和某些 体内发育阶段。 这表明特定于阶段 聚合酶III因子的形式？ 在一个相关的发现中，我们有 表明其他两个III类基因TDNAMET和OAX也是 在整个卵母细胞和卵母细胞中差异表达 核提取物。 最有趣的是，这些DNA与5s竞争 DNA对于核提取物中的因素，但在整个卵母细胞中却没有 提炼。 我们希望确定这一发现是否反映了多个 整体中聚合酶III因子的基因特异性形式 卵母细胞S150提取物。 该提案的具体目的是： 1。要优先确定S150提取物中的哪个因素 序列与体细胞5S基因相互作用 发起人之前的差异。 2。研究为什么这些序列差异提供 卵母细胞S150提取物中的转录优势，但不在 核提取物。 3。更准确地确定哪些序列差异最大 对这种影响很重要。 4。研究为什么E190和TDNAMET基因差异化 在S150和核提取物中表达。 5。调查为什么E190和TDNAMET不与5S DNA竞争 在S150中，但确实在核提取物中竞争。 做这些 发现反映了因子种群的差异？ 6。研究为什么5s，E190和TDNAMET具有类似的活动 在异源提取物（HELA）中，但根本不同 同源提取物中的活动。