CHARACTERIZATION OF HOMEO BOX HOMOLOGIES IN THE LEECH

最近已经证明，存在直接的DNA序列， 果蝇同源异型基因之间的同源性。 该地区已被 被称为homeo盒，并已被证明编码一个保守的，约60 氨基酸残基序列 基因组DNA的交叉杂交， 各种其他物种的探针含有果蝇同源盒， 认为同源盒的区域在进化上是 保守的 交叉杂交也可用于分离 青蛙、老鼠和人类中含有这个区域的一些基因。 因为果蝇的基因中含有同源盒， 发展过程，特别是分段和分段 为了区分，已经提出，homeo盒也可以是 在其他物种中发现这些基因。 一组动物， 最合适的例子是水蛭。 水蛭，就像 所有其他环节动物和昆虫一样， 组织他们的身体。 目前的进化理论认为昆虫 从环节动物的祖先，这表明， 昆虫同源异型基因是从相应的环节动物基因进化而来的。 此外，我们的初步观察表明，水蛭也有 含有与果蝇同源盒交叉杂交的区域的基因。 本项目的目的是鉴定和分离这些基因 几种水蛭，并确定它们的空间和 表达的时间模式与果蝇相似， 含有同源盒的基因。 我们将首先实现这一目标， 确定同源盒交叉杂交区域的近似数目， 用Southern杂交分析水蛭的DNA。 然后我们将确定 与同源盒的同源性，通过测序水蛭DNA的区域， 并将通过北方分析来表征其转录本。 最后，我们将在连续的组织切片上进行原位杂交， 水蛭胚胎，以评估表达的空间和时间模式， 水蛭的基因 特别是，我们将研究 这些基因在个体的水平上， CNS，以确定差异基因表达是否与 神经元表型的节段分化 表征了 这次调查的结果应该有一个重要的 影响了我们对基因进化的理解， 这些基因在表型遗传中的作用 CNS的异质性。

It has been recently demonstrated that there is direct DNA sequence homology between several Drosophila homeotic genes. This region has been called the homeo box and has been shown to code for a conserved, about 60 amino acid residue sequence. Cross-hybridization of genomic DNA from various other species with probes containing the Drosophila homeo box has suggested that regions homologous to the homeo box are evolutionarily conserved. Cross-hybridization has also been useful in the isolation of some of the genes that contain this region in frogs, mice and humans. Because the Drosophila genes that contain the homeo box affect developmental processes, in particular segmentation and segmental differentiation, it has been proposed that the homeo box could also be a marker for such genes in other species. A group of animals in which a test of this idea would be particularly appropriate are leeches. Leeches, like all other annelids and like the insects, display an obvious segmental organization of their bodies. Current evolutionary theory has the insects descending from an annelid ancestor, suggesting the possibility that the insect homeotic genes have evolved from corresponding annelid genes. Furthermore, our preliminary observations indicate that leeches also have genes that contain regions which cross-hybridize with the fly homeo box. The aim of the project proposed here is to identify and isolate such genes in a few species of leeches, and to determine whether their spatial and temporal patterns of expression are similar to those of the Drosophila genes that contain the homeo box. We will pursue this goal by first determining the approximate number of homeo box cross-hybridizing regions in the leech by Southern blot analysis. We will then determine the degree of homology with the homeo box by sequencing the regions of leech DNA of interest and will characterize their transcripts by Northern analysis. Finally, we will do in situ hybridization on serial tissue sections of leech embryos to assess the spatial and temporal patterns of expression of the identified leech genes. In particular, we will study the expression of these genes at the level of individual, identified neurons in the leech CNS, to determine whether differential gene expression correlates with the segmental differentiation of neuronal phenotypes that we have characterized. The results of this investigation should have a significant bearing on our understanding of the evolution of genes that control early development and of the role of such genes in the genesis of phenotypic heterogeneity in the CNS.