Normal and aberrant switch recombination to the murine alpha heavy chain gene

描述(申请人提供)：重链开关是一种受调控的DNA重组缺失，对抵抗感染性病原体具有重要意义。在B淋巴细胞中的正常事件中，缺失事件开始于位于Mu重链恒定区上游的DNA序列，结束于位于γ、epsilon或α恒定区基因上游的DNA序列。这一缺失事件使编码可变区的外显子与新的恒定区并列，使B淋巴细胞能够表达一种可能更适合于特定病原体的新型抗体。在极少数情况下，其中一个重链基因与位于不同染色体上的c-myc癌基因发生重组事件。这种异常的重链开关导致了染色体的相互易位，是小鼠浆细胞瘤和人类淋巴瘤的潜在原因之一。我们的长期目标是了解生理性开关重组和导致肿瘤的异常开关重组的序列要求。为了研究开关重组的规律和机制，我们开发了一个230kb的重链恒定区基因全长的转基因，它可以像内源重链基因一样进行开关重组。在这里提出的可行性项目中，我们将确定转基因重链基因是否也会发生染色体易位到c-myc，从而导致浆细胞瘤。在目标1中，我们将测试在用Pristane诱导浆细胞瘤后，转基因重链基因是否易位到具有3到4个拷贝的转基因小鼠。在目标2和目标3中，我们将使用包括共表达bc1-xl转基因的诱导方案来测试转基因的易位。我们将把重链转基因的易位能力与染色体插入位置、方向和等位基因排斥相关联。如果重链基因可以与c-myc一起发生易位，那么在转基因中进行任何所需的突变、缺失或插入的能力为我们提供了一个机会来确定重链基因中的哪些元件对异常重组事件和随后的c-myc转录失控起关键作用。与研究所的使命相关：从这些研究中获得的信息将提高我们对肿瘤中常见的染色体易位如何发生的理解。这些信息也可能与正常的开关重组有关，并可能对更好的疫苗设计产生影响。

DESCRIPTION (provided by applicant): The heavy chain switch is a regulated DNA recombinational deletion that has important implications for protection against infectious agents. In the normal event in B lymphocytes, a deletion event begins in DNA sequences that lie upstream of the mu heavy chain constant region and ends in DNA sequences that lie upstream of the gamma, epsilon, or alpha constant region genes. This deletion event bring the exon encoding the variable region into juxtaposition with a new constant region, allowing B lymphocytes to express a new type of antibody that may be more suitable for a given pathogen. In rare instances, one of the heavy chain genes undergoes a recombination event with the c-myc oncogene on a different chromosome. This aberrant heavy chain switch, resulting in a reciprocal chromosomal translocation, is one of the underlying causes of plasmacytomas in mice and lymphomas in humans. Our long term goal is to understand the sequence requirements of both physiologic switch recombination and the aberrant switch recombination that leads to tumors. In order to study the regulation and mechanism of switch recombination, we have developed a 230 kb transgene of the entire heavy chain constant region locus that is able to undergo switch recombination like that of the endogenous heavy chain genes. In the feasibility project proposed here, we will determine if the transgenic heavy chain locus can also undergo chromosomal translocations to c-myc, resulting in plasmacytomas. In Aim 1 we will test for translocations, after induction of plasmacytomas with pristane, to the transgenic heavy chain genes of mice with three or four copies of the transgene. In Aims 2 and 3, we will test for translocations to the transgene with an induction protocol that includes co-expression of a bcl-xL transgene. We will correlate the ability of the heavy chain transgene to translocate with chromosomal insertion site, orientation, and allelic exclusion. If the heavy chain locus transgene can undergo translocations with c-myc, the ability to make any desired mutation, deletion, or insertion in the transgene provides us with an opportunity to determine which elements in the heavy chain locus are critical for the aberrant recombination event and subsequent deregulation of c-myc transcription. Relevance to mission of the Institutes: Information gained from these studies will improve our understanding of how the chromosomal translocations common in tumors occur. The information is also likely to be relevant to normal switch recombination, with implications for better vaccine design.