Bienz-Tadmor B, Tolias P, Stebbins-Boaz B, Mariani B D, Gerbi S A, Kafatos F C
Department of Cellular and Developmental Biology, Harvard University, Cambridge, MA 02138.
Chromosoma. 1992 Aug;101(9):538-48. doi: 10.1007/BF00660313.
P element mediated germ-line transformation was used to study the developmental specificity of Drosophila chorion gene regulatory sequences directing expression of the bacterial reporter genes for chloramphenicol acetyltransferase (CAT) and beta-galactosidase (lacZ). DNA fragments containing 5' flanking plus the entire 5' untranslated and the beginning of the coding region of either the s36 or the s15 chorion gene are able to confer on the reporter genes normal tissue as well as temporal specificity of expression, exclusively in the ovary of transformed female flies. However, if 5' untranslated and coding regions are omitted, normal ovarian expression is maintained but tissue specificity is relaxed: expression of the reporter gene is detected both in the ovary and in specific non-ovarian tissues of transformed females and males. The evidence suggests that the missing 5' untranslated and coding sequences may include negative elements that normally suppress expression in non-ovarian tissues, and that these putative elements are distinct from those that prevent premature expression in the ovarian follicles. The exact location of ectopic lacZ expression within the internal male genitalia depends on the constellation of 5' flanking chorion regulatory sequences included in the P element constructs. Ectopic expression of the CAT gene in the male genitalia under s15 promoter control can be abolished by mutating the hexamer TCACGT, a sequence previously shown to be essential for the normal expression of this chorion gene in the ovary.
P 因子介导的种系转化被用于研究果蝇绒毛膜基因调控序列的发育特异性,这些调控序列指导氯霉素乙酰转移酶(CAT)和β-半乳糖苷酶(lacZ)这两种细菌报告基因的表达。含有 s36 或 s15 绒毛膜基因 5'侧翼加上整个 5'非翻译区及编码区起始部分的 DNA 片段,能够赋予报告基因正常的组织以及时间特异性表达,且仅在转化雌蝇的卵巢中表达。然而,如果省略 5'非翻译区和编码区,正常的卵巢表达得以维持,但组织特异性会减弱:在转化的雌蝇和雄蝇的卵巢以及特定的非卵巢组织中都能检测到报告基因的表达。证据表明,缺失的 5'非翻译区和编码序列可能包含通常抑制非卵巢组织中表达的负调控元件,并且这些假定的元件与那些防止卵巢卵泡过早表达的元件不同。异位 lacZ 在雄性内生殖器中的表达的确切位置取决于 P 因子构建体中包含的 5'侧翼绒毛膜调控序列的组合。在 s15 启动子控制下,CAT 基因在雄性生殖器中的异位表达可以通过突变六聚体 TCACGT 来消除,该序列先前已被证明对于该绒毛膜基因在卵巢中的正常表达至关重要。