GASTROINTESTINAL-LIVER ALKALINE PHOSPHATASES

人碱性磷酸酶是一个重要的多基因家族， 多达五种组织特异性形式。 碱性磷酸酶具有 广泛应用于诊断医学和研究， 在过去的50年里一直在深入研究。 三 染色体位点已经确定。 另外两个假设 PLAP样和IAP样在肿瘤组织中均有表达 因此是潜在的肿瘤标记物。 表征 在DNA水平上这两种假定的形式将提供特定的DNA 进行临床研究以评估其潜力的探针 作为肿瘤和疾病标记物的应用。 我们实验室的长期目标是继续解开 其余假定的人碱性磷酸酶的结构 同工酶;使用这个多基因家族作为研究组织的模型 特异性，并研究一些这些“异常”表达 来自癌组织和肿瘤细胞系的同工酶。 我们之前 分离了PLAP cDNA，并从北方分析中得到证据， PCR扩增的mRNA表明， 碱性磷酸酶在胎盘组织中表达。 这种形式 可能是IAP样碱性磷酸酶。 我们 目前，我们的PLAP和PLAP样基因， 密切相关，但具有不同的组织特异性。 因此， 它们将是研究组织特异性的良好模型。 在本申请中，我们建议：1）完成我们的 PLAP和推定的PLAP样基因的表征， 对这两个基因进行测序，以绘制染色体位置， PLAP样基因; 2）表征这种多基因的其他形式 家族，包括IAP样形式，在JEG中表达的其他形式， 3（绒毛膜癌）和LS 174 T（结肠腺癌）人癌 细胞系和肝组织形式的非特异性碱性 磷酸酶;和3）使用我们的PLAP研究组织特异性， PLAP样基因。 我们构建了cDNA文库， 我们的cDNA和基因组的主要cDNA和寡核苷酸探针 筛选 序列分析将由 克隆后双脱氧核苷酸链终止法 验证 我们将研究组织特异性， 上游调控区与报告基因，氯霉素 乙酰转移酶，并在细胞系中进行缺失研究 表达了这两个密切相关的一个或两个 基因.

Human alkaline phosphatase is an important multigene family with as many as five tissue-specific forms. Alkaline phosphatase have extensive application in diagnostic medicine and research and has been under intensive study during the past 50 years. Three chromosomal loci have been determined. The other two putative forms, PLAP-like and IAP-like are both expressed in tumor tissue and are therefore potential tumor markers. Characterization of these two putative forms at the DNA level will provide specific DNA probes to carry out clinical studies to evaluate their potential applications as tumor and disease markers. The long-term goal of our laboratory is to continue to unravel the structures of the remaining putative human alkaline phosphatase isozymes; to use this multigene family as a model to study tissue specificity and to study the "aberrant" expression of some these isozymes from cancer tissue and tumor cell lines. We previously isolated the PLAP cDNA and have evidence from Northern analysis and PCR amplification of mRNA to suggest that na intestinal form of alkaline phosphatase is expressed in placental tissue. This form may be the putative IAP-like alkaline phosphatase. We are currently characterizing our PLAP and PLAP-like genes which are closely related but with different tissue specificity. There fore, they will be a good model to study tissue specificity. In this application we propose to: 1) complete our characterization of the PLAP and the putative PLAP-like genes by sequencing these two genes, to map the chromosome location of the PLAP-like gene; 2) characterize other forms of this multigene family including the IAP-like form, other forms expressed in JEG- 3 (choriocarcinoma) and LS174T (colon adenocarcinoma) human cancer cell lines and the liver form of tissue unspecific alkaline phosphatase; and 3) study tissue specificity using our PLAP and PLAP-like genes. We have constructed cDNA libraries and will use primarily cDNA and oligonucleotide probes for our cDNA and genomic screening. Sequence analysis will be performed by dideoxynucleotide chain termination method after clone verification. We will study tissue specificity by linking the 5' upstream regulatory region with a reporter gene, chloramphenicol acetyl-transferase and to carry out deletion studies in cell lines which express either one or both of these two closely related genes.