Control of Ig Gene Expression in Ig Transgenic Mice

描述(由申请人提供)：我们提出两个目标，其中Ig 转基因小鼠将用于研究V基因表达的调控 发育中的B细胞。第一个目标是关于VH基因的表达，这是 成年小鼠和胎鼠的B细胞差异很大。在发育中的B细胞 在成年小鼠中，VH基因的表达已知依赖于 不同H(Mu)链与L链配对的效率 常规)以及在给出一种特定的 穆某和L成对链条。VH表达是否同样依赖于这样的 胎儿B细胞发育过程中的细胞选择尚不清楚。要解决这个问题 ，我们将选择性地培育两个IGL链转基因小鼠系 (V-kappa-8和V-lambda-L小鼠)缺乏替代轻(SL)链和 内源L(Kappa)链。在该模型中，V-kappa-8或V-lambda-L链将 替换SL链。新生的u链基本上将有选择 仅与单个L链(V-kappa-8或V-lambda-L)配对和形成 B细胞受体(BCR)。我们将测试V-kappa-8和V-lambda-L小鼠的使用情况 不同VH基因的差异Mu-L链配对及其证据 基于bcr特异性的B细胞选择。使用一种新的检测系统，我们将 还要测量不同的VH基因家族的相对频率 重新编排的目标。通过这种方式，我们将评估表达式是否 胎儿和成人B细胞中不同的VH基因主要反映了不同的 细胞选择的结果(假设-1)或可能没有 胎儿B细胞发育中的细胞选择(假设2)。 第二个目的是控制VL基因的表达。我们观察到了一个 V-Lambda-L/J-Lambda-L重排差分控制的显著例子 在两个免疫球蛋白(MU)转基因小鼠(M54和3H9小鼠)中。MM，但不是3H9 小鼠，V-lambda-1/J-lambda-L重排受到抑制。我们假设 这些转基因的差异效应是由于vh相关的差异造成的。 在M54和3H9MU链中或TG诱导的基因表达差异 在M54和3H9B系细胞之间。我们将使用以下工具测试这些假设 几种不同的策略。这将包括测试M54亩 链，或通过减去M54之间的基因产物鉴定的任何基因产物 和3H9前B细胞，可抑制温度诱导的V-lambda-1/J-lambda-1 转化的PM-B细胞系中的重排。

DESCRIPTION (Provided by the Applicant): We propose two aims in which Ig transgenic mice will be used to investigate the control of V gene expression in developing B-cells. The first aim deals with the expression of VH genes, which differs dramatically in B-cells of adult and fetal mice. In developing B-cells of adult mice, the expression of VH genes is known to be dependent on the efficiency with which different H (mu) chains pair with L chains (surrogate or conventional) and on the antigen-binding specificity that results when a given mu and L chain pair. Whether VH expression is similarly dependent on such cellular selection in developing fetal B-cells is not known. To address this issue, we will selectively breed two lines of IgL chain transgenic mice (V-kappa-8 and V-lambda-l mice) lacking the surrogate light (SL) chain and endogenous L(kappa) chains. In this model, V-kappa-8 or V-lambda-l chains will substitute for the SL chain. Nascent u chains will essentially have the choice of only a single L chain (V-kappa-8 or V-lambda-l) with which to pair and form a B-cell receptor (BCR). We will test V-kappa-8 and V-lambda-l mice for usage of different VH genes, differential mu-L chain pairing and for evidence of B-cell selection based on BCR specificity. Using a novel assay system, we will also measure the relative frequency at which different VH gene families are targeted for rearrangement. In this way, we will assess whether the expression of different VH genes in fetal and adult B-cells primarily reflects different outcomes of cellular selection (hypothesis-1) or the possible absence of cellular selection in fetal B-cell development (hypothesis-2). The second aim deals with the control of VL gene expression. We have observed a striking example of differential control of V-lambda-l/J-lambda-l rearrangement in two lines of IgH (mu) transgenic mice (M54 and 3H9 mice). In MM but not 3H9 mice, V-lambda-1/J-lambda-l rearrangement is suppressed. We hypothesize that the differential effect of these transgenes is due to a VH-related difference in the M54 and 3H9 mu chains or a tg-induced difference in gene expression between M54 and 3H9 B lineage cells. We will test these hypotheses using several different strategies. This will include testing whether the M54 mu chain, or any of the gene products identified by cDNA subtraction between M54 and 3H9 pre-B-cells, can inhibit temperature-induced V-lambda-1/J-lambda-1 rearrangement in transformed pm-B-cell lines.