The Evolution of Lymphocyte Recognition

本研究的长期目标是了解鲫鱼免疫球蛋白（Ig）重链基因的结构、组织和表达。在噬菌体中构建了基因组文库；三个VH的初步测序。并通过寡核苷酸探针杂交鉴定出推测的JH区。这些研究将扩大到三个领域。首先，将鉴定和测序具有代表性的VH、DH、JH和CH区域，以阐明其种系组织。将使用标准的DNA克隆、制图和测序策略。为了鉴定CH区域，将对金鱼IgM重链的肽进行微测序，并合成特异性的混合寡核苷酸探针，以对应其编码序列。其次，对金鱼的VH基因序列进行计算机辅助分析，并将这些序列与其他脊椎动物的VH序列进行比较，试图推断选择、突变、漂变和非漂变的相对贡献。在这些基因进化过程中的相互交换。第三，研究金鱼Ig重链基因的调控区域的功能。我们将构建合适的表达载体来测试金鱼重链启动子（与哺乳动物Ig增强子区相关）是否允许基因在哺乳动物淋巴样细胞中表达。除哺乳动物外，对动物免疫功能的机制了解甚少。了解鱼体内免疫球蛋白基因的结构、组织和表达调控的基本知识不仅对其内在的利益具有重要意义，同时也因为了解鱼类的免疫功能对于鱼类在商业养殖中生长和维持的能力以及了解鱼类在自然环境中生存的成功或失败的因素至关重要。从这些研究中获得的免疫功能进化的知识也可能揭示高等脊椎动物免疫系统的功能。

The long term goals of this research are to understand the structure, organization, and expression of immunoglobulin (Ig) heavy chain genes in the goldfish, Carassius auratus. A genomic library has been constructed in lambda phage; initial sequencing of three VH.related genes has been completed, and putative JH regions have been identified by oligonucleotide probe hybridization. These studies will be expanded in three areas. First, representative VH, DH, JH, and CH regions will be identified and sequenced to elucidate their germline organization. Standard DNA cloning, mapping, and sequencing strategies will be used. To identify CH regions, peptides of goldfish IgM heavy chains will be microsequenced, and specific mixed oligonucleotide probes will be synthesized to correspond to their coding sequences. Second, computer assisted analyses of goldfish VH gene sequences and comparison of these with each other and with published VH sequences for other vertebrates will be undertaken in an attempt to infer the relative contributions of selection, mutation, drift, and non.reciprocal exchange in the evolution of these genes. Third, the function of putative regulatory regions of the goldfish Ig heavy chain genes will be studied. Appropriate expression vectors will be constructed to test whether the goldfish heavy chain promoter (in association with a mammalian Ig enhancer region) permits gene expression in mammalian lymphoid cells. The mechanisms of immune function in animals other than mammals are as yet poorly understood. Basic knowledge of the structure, organization and regulation of expression of immunoglobulin genes in fish is of importance not only for its intrinsic interest, but also because understanding of immune function in fish is fundamental for the capability to grow and maintain of fish in commercial culture and for understanding the factors involved in the success or failure of fish to survive in natural environments. Knowledge of the evolution of immune function obtained from these studies may also shed light on the function of immune systems in higher vertebrates.