REGULATION OF ERYTHROPOIESIS BY CELL SURFACE COMPONENTS

我们发现促进红系爆裂的活性(BPA)表达为 淋巴细胞膜。我们现在建议利用一种生物化学方法 明确的文化体系，以： 1.纯化膜相关双酚A。我们将利用各种不同的 生化方法，包括免疫亲和力、凝集素亲和力和凝胶 高效液相色谱过滤和反相 高效液相色谱法从淋巴细胞质膜和淋巴提取液中分离纯化双酚A 水泡。提纯的组分将通过等电聚焦来表征 蔗糖梯度分级。最终，纯化的双酚A将被 进行氨基酸分析。 2.测定可溶性BPA与囊泡性BPA的相关性。我们将聘用 与双酚A反应的抗血清和单抗 从膜和溶液中纯化的双酚A在抗原性上是相关的。 将进行研究以确定双酚A的两种物理形式是否 由类似的机制从相似的细胞类型释放，如果它们作用于 相似的骨髓亚群。每个生长因子的目标细胞将是 具有组织化学和形态特征。 3.定义囊泡相关双酚A的释放动力学。累计金额 以及产生双酚A的细胞群体的囊泡形成和释放双酚A的速率 将会被确定。我们的发现将与旨在 测定培养物中选定的膜成分的生物合成。 4.研究双酚A的作用模式。我们将确定双酚A的脱落是否 免疫反应免疫反应的构成过程或部分研究将会是 检查囊泡形成是否涉及细胞骨架成分 以及膜脂和磷脂组成是否对细胞周期有重要影响 剥离过程。我们将确定水泡是否与 优先使用骨髓细胞，如果是的话，这是什么性质的 协会可能使用了电子显微镜的方法。最终，我们 将定位和纯化BPA受体，计划的长期目标是 检查受体激活的细胞内机制。

We have found that erythroid burst promoting activity (BPA) is expressed by lymphocyte plasma membranes. We now propose to utilize a biochemically defined culture system to: 1. Purify membrane-associated BPA. We will utilize a variety of biochemical methods, including immunoaffinity, lectin affinity and gel filtration high-performance liquid chromatography (HPLC) and reverse-phase HPLC, to purify BPA from extracts of lymphocyte plasma membranes and shed vesicles. Purified fractions will be characterized by isoelectric focusing and sucrose gradient fractionation. Ultimately, purified BPA will be subjected to amino acid analysis. 2. Determine relatedness of soluble and vesicular BPA. We will employ antisera and monoclonal antibodies that react with BPA to detect whether purified BPA from membranes and in solution are antigenically related. Studies will be performed to determine if both physical forms of BPA are released by similar mechanisms from similar cell types, and if they act on similar marrow subpopulations. Target cells for each growth factor will be characterized histochemically and morphologically. 3. Define kinetics of vesicle-associated BPA release. Cumulative amounts and rates of vesiculation and BPA release by BPA-producing cell populations will be determined. Our findings will be correlated with studies aimed at measuring the biosynthesis of selected membrane components in culture. 4. Examine modes of BPA action. We will determine whether BPA shedding is a constitutive process or part of an immune reaction. Studies will be performed to examine whether vesiculation involves cytoskeletal components and whether membrane lipid and phospholipid composition is important to the shedding process. We will determine whether vesicles associate preferentially with marrow cells and, if so, what the nature of this association might be using electronmicroscopic methods. Ultimately, we will locate and purify BPA receptors with the projected long-range goal to examine intracellular mechanisms for receptor activation.