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 转基因小鼠将被用于研究V基因表达的控制， B细胞发育第一个目的涉及VH基因的表达， 在成年小鼠和胎儿小鼠的B细胞中有显著差异。在B细胞的发育过程中 在成年小鼠中，已知VH基因的表达依赖于 不同的H（mu）链与L链配对的效率（替代或 常规的）和当给定的抗原结合特异性时产生的抗原结合特异性。 mu和L链对。VH表达是否类似地依赖于这种 发育中胎儿B细胞的细胞选择是未知的。为了解决这个 本课题将选育两系IgL链转基因小鼠 （V-κ-8和V-λ-1小鼠）缺乏替代轻链（SL）， 内源性L（κ）链。在该模型中，V-κ-8或V-λ-1链将 替代SL链。新生的u链基本上可以选择 只有一条L链（V-κ-8或V-λ-1）与之配对并形成 B细胞受体（BCR）。我们将测试V-kappa-8和V-lambda-1小鼠的使用情况 不同的VH基因，不同的mu-L链配对和证据， 基于BCR特异性的B细胞选择。使用新的分析系统，我们将 还测量了不同VH基因家族的相对频率， 以重组为目标。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VH基因的差异主要反映了不同的 细胞选择的结果（假设-1）或可能缺乏 胎儿B细胞发育中的细胞选择（假说2）。 第二个目的涉及VL基因表达的控制。我们观察到 V-lambda-1/J-lambda-1重排微分控制一个突出例子 在两种IgH（mu）转基因小鼠（M54和3 H9小鼠）中。在MM中，但不是3 H9 在小鼠中，V-lambda-1/J-lambda-1重排被抑制。我们假设 这些转基因的差异效应是由于VH相关的差异 M54和3 H9 mu链或tg诱导的基因表达差异 M54和3 H9 B系细胞之间的差异。我们将测试这些假设使用 几种不同的策略。这将包括测试M54 mu是否 链，或通过M54和M55之间的cDNA扣除鉴定的任何基因产物。 和3 H9前B细胞，可以抑制温度诱导的V-λ-1/J-λ-1 在转化的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.