SL12 T-LYMPHOMA--A NEW MODEL FOR GENE CONTROL IN TUMORS

从小鼠T淋巴瘤细胞系SL12.4中分离到新的cDNA克隆 通过与姐妹细胞克隆进行减法杂交 这些基因的表达。这项建议主要集中在三个最 有趣的克隆，具有意想不到和惊人的特征。 这三个都是新奇的，因为它们与已知的没有明显的相似之处 基因或基因产物。它们在胚胎发育过程中受到调控， 在成年动物中显示组织特异性表达，并可在 在体外系统中定义良好。其中一种新基因具有 一种致癌胎儿基因。另一种仅在T淋巴系中表达 转录本在T细胞激活过程中被强烈诱导。这个 第三种是在经历成熟的SL12.4细胞中可诱导的，并且高度 在卵巢组织和人卵巢癌细胞系中表达。 基因表达的调控。对每个基因的调控将是 在诱导性和非诱导性条件下，在相关和 不相关的谱系，并在体细胞杂交种之间表达 和非表达细胞。将对转录速度进行评估。5英尺 将分离每个基因的调节区，并测试其功能 瞬时转染法。评估DNA-蛋白质的体外实验 我们将检查控制区中的相互作用。 蛋白质产品的鉴定。有人建议进行研究，以分离和 确定蛋白质产品的特征。其中一个基因预测一种蛋白质 有4个假定的跨膜结构域，很可能是 定位于细胞表面。另外两个可能是细胞内的 蛋白质。将针对预测的蛋白质制备抗体以 评估蛋白质的物理性质，潜在的糖基化， 亚基结构，并确定每个亚细胞的定位。 新基因表达的分析。基因的表达将是 在淋巴瘤发展的离散阶段的AKR动物中使用一种 由Esther Hays博士(将提供细胞)开发的模型系统。 基因的表达将在选定的成人组织中进行检测，并 在胚胎发育过程中使用原位杂交。

Novel cDNA clones were isolated from the SL12.4 murine T-lymphoma cell line by subtraction hybridization against a sister cell clone which lacks expression of these genes. This proposal focuses on the three most interesting clones which possess unexpected and striking characteristics. All three are novel in that they have no significant similarity with known genes or gene products. They are regulated during embryonic development, show tissue specific expression in adult animals, and can be induced in well defined in vitro systems. One of the novel genes has the hallmarks of an onco-fetal gene. Another is expressed only in the T-lymphoid lineage and the transcripts are strongly induced during T cell activation. The third is inducible in SL12.4 cells undergoing maturation and is highly expressed in ovarian tissues and human ovarian carcinoma cell lines. Regulation of gene expression. The regulation of each of the genes will be examined under inducing and non-inducing conditions, in related and unrelated lineages, and in somatic cell hybrids formed between expressing and non-expressing cells. Rates of transcription will be assessed. The 5' regulatory region of each gene will be isolated and tested for function in transient transfection assays. In vitro experiments to assess DNA-protein interactions in the control regions will be examined. Identification of the protein product. Studies are proposed to isolate and characterize the protein products. One of the genes predicts a protein with 4 putative transmembrane spanning domains and is likely to be localized on the cell surface. The other two likely to be intracellular proteins. Antibodies will be prepared against the predicted proteins to assess the physical properties of the protein, potential glycosylation, subunit structure and to determine the subcellular localization of each. Analysis of novel gene expression. The expression of the genes will be examined in AKR animals at discrete stages of lymphoma development using a model system developed by Dr. Esther Hays (who will provide the cells). Expression of the genes will be examined in selected adult tissue and during embryogenesis using in situ hybridization.