摘要
目的鉴定果蝇品系mf917的突变基因slmb,并探究其造成果蝇翅及复眼发育缺陷的原因。方法突变品系mf917进行直接镶嵌克隆(direct mosaic clone)分析和镶嵌克隆(mosaic clone)分析,观察成虫翅及复眼的表型并解剖翅成虫盘;使用免疫荧光(immunofluorescent,IF)染色方法检测Notch、Wnt、Hippo和Hedgehog信号通路的靶基因或报告基因在mf917克隆中的表达情况,判断mf917与这些信号通路的相关性;使用反向PCR鉴定mf917中Minos转座子的插入位点及所影响的基因;利用转座子精确剪切方法检测插入mf917转座子跳除后表型是否恢复;使用mf917与已报道的slmb突变进行互补实验,进一步验证mf917与slmb的相关性。结果突变品系mf917存在翅发育缺陷以及复眼增生等表型,三龄幼虫翅成虫盘野生型区域出现明显增生;在mf917克隆中,Notch信号靶基因Cut和Wingless(Wg)表达无明显变化,Wnt信号报告基因Dll-lacZ和fz3-lacZ表达升高,Hippo信号报告基因CycE-lacZ和ex697-lacZ表达下降、bantam sensor-GFP表达升高,Hedgehog信号报告基因ptc-lacZ和Ci蛋白在翅成虫盘前部腔室克隆中表达升高;mf917突变影响的基因是slmb。结论mf917通过调控Wnt、Hippo和Hedgehog信号通路,从而影响果蝇翅和复眼的发育,mf917鉴定为一个slmb新突变体。
Objective To identify the mutated gene slmb in the Drosophila strain mf 917,and investigate the causes of developmental defects in the wings and compound eyes of Drosophila.Methods The mutant strain mf 917 was analyzed by direct mosaic clone and mosaic clone.The phenotypes of the adult wings and compound eyes were observed,and the adult wing discs were dissected.The expression of target genes or reporter genes of Notch,Wnt,Hippo,and Hedgehog signaling pathways in mf 917 homozygous mutant clones were detected by immunofluorescent(IF)staining to assess the correlation between mf 917 and these signaling pathways.Utilized inverse PCR to identify the insertion sites of Minos transposons in mf 917 and the genes affected by them.Utilized the transposable element precise excision method to detect whether the phenotype recovers after the mf 917 transposons are removed by jumping out.Conducted complementary experiments using mf 917 and reported slmb mutations to further verify the correlation between mf 917 and slmb.Results The mutant strain mf 917 had phenotypes such as wing development defects and compound eye hyperplasia.Significant proliferation occurs in the wild-type region of the adult disc in the wings of third-instar larvae.In the mf 917 clone,there were no significant changes in the expression of Notch signaling target genes Cut and Wingless(Wg)while the expression of Wnt signaling reporter genes Dll-lacZ and fz3-lacZ increased.The Hippo signaling reporter genes CycE-lacZ and ex697-lacZ were down-regulated and bantam sensor-GFP was up-regulated in mf 917 mutant clone.The Hedgehog signaling reporter gene ptc-lacZ and Ci protein exhibit elevated expression in the anterior compartment clone of the wing imaginal disc.The gene affected by the mf917 mutation is slmb.Conclusion mf 917 affects the development of Drosophila wings and compound eyes by regulating the Wnt,Hippo,and Hedgehog signaling pathways,and is identified as a novel slmb allele.
作者
冯春霞
杨敏
罗俊鹏
严江松
安云飞
符纯海
张博文
杨颜静
魏微
张庆海
FENG Chunxia;YANG Min;LUO Junpeng;YAN Jiangsong;AN Yunfei;FU Chunhai;ZHANG Bowen;YANG Yanjing;WEI Wei;ZHANG Qinghai(Department of Biology,School of Basic Medicine,Guizhou Medical University,Guiyang 550025,Guizhou,China;Key Laboratory of Medical Insects,School of Basic Medicine,Guizhou Medical University,Guiyang 550025,Guizhou,China;Research Center of Basic Medical Science,Guizhou Medical University,Guiyang 550025,Guizhou,China;Collegeof Biology and Engineering,Guizhou Medical University,Guiyang 550025,Guizhou,China;Clinical College,Guizhou Medical University,Guiyang 550025,Guizhou,China;Laboratory of Multimedia Morphology,School of Basic Medicine,Guizhou Medical University,Guiyang 550025,Guizhou,China)
出处
《贵州医科大学学报》
2025年第8期1144-1155,共12页
Journal of Guizhou Medical University
基金
国家自然科学基金项目(31601898,31960128)
贵州省科学技术基金项目(黔科合基础[2017]1158
黔科合基础[2020]1Y082)。
