Juvenile hormone receptor and molecular mechanism of gene expresion

为了研究昆虫幼虫激素（JH）调控基因表达的分子机制，本研究利用mRNA差异显示的方法分析了虫幼虫激素（JH）的应答基因。将JH类似物甲基戊二烯应用于滞育雌成虫，并从脂肪体中提取RNA。我们发现许多cDNA条带在JHA和溶剂丙酮处理动物的mRNA差异显示上存在差异。我们通过JH处理提取了诱导或抑制的cDNA。利用这些cDNA，我们对JH或丙酮处理的脂肪体RNA进行了Northern Blot分析，鉴定出两个JH诱导基因（ji -1和JI-2）和三个JH抑制基因(JR-！JR-2和JR-3)。对这些cDNA的部分序列分析和数据库同源性检索表明，JI-1和JI-2分别与cyanoprotein-a和vitellogenin具有较高的同源性，为血淋巴储存六聚体和蛋黄蛋白前体。JR-1与脊椎动物和昆虫的转铁蛋白具有高度同源性。另一方面，未发现与JR-2和JR-3有同源性的基因，推断这些基因为新基因。我们克隆并测序了JI-2和JR-1的cDNA全长，并通过NOrthern Blot分析JH处理后mRNA水平的变化。克隆了基因组DNA，分析了上游调控基因JI-1 （CP-a）、JI-2 （Vg）和JR-1 （Transferrin）的结构。目前已有3个JH诱导基因和3个抑制基因，可以比较相同激素条件下JH在同一昆虫中的表达调控。

To study the molecular mechanism of gene expression regulation by insect juvenile hormone (JH), we have analyzed JH responsive genes by mRNA Differential Display in the bean bug, Riptortus clavatus. JH analog, methoprene, was applied on the diapause female adults and extracted RNA from the fat body. We found many cDNA bands which showed differnence between JHA and solvent acetone treated animals in mRNA Differential Display. We picked up the cDNA which were induced or repressed cDNA by JH treatment. Using these cDNA we performed Northern Blot analyzes for JH or acetone treated fat body RNA,and identified two JH inducible genes (JI-1and JI-2) and three JH repressible genes (JR-! , JR-2 and JR-3). The analysis of the partial sequeces of these cDNA and data base homology search indicated that JI-1 and JI-2 had high homology to cyanoprotein-a and vitellogenin, respectively, which we have showed hemolymph storage hexamer and egg yolk protein precursor. JR-1 was showed high homology to transferrin in vertebrate and insects. On the other hand we could not find any genes which have homology to JR-2 and JR-3 and concluded these gene are novel. We cloned and sequenced full length of cDNA of JI-2 and JR-1 and analyzed the changes of mRNA level after JH tretment by NOrthern Blot analyzes. We also cloned genomic DNA and analyzed the upstream regulatory gene structure of JI-1 (CP-a), JI-2 (Vg) and JR-1 (Transferrin). Now three JH inducible and three repressible genes are available to compare the regulation of expression by JH in the same insect under the same hormonal condition.