Process for formation of chromatin functional domains using Drosophila

除携带UAS-DREF和UAS-Mi-2外，还建立了携带UAS-BEAF和UAS-BEAF RNAi的转基因果蝇品系。利用这些系，BEAF蛋白的过度表达或下调是以组织特异性的方式实现的。BEAF蛋白在眼成像盘细胞中的过度表达导致了粗眼表型。另一方面，当BEAF转录本被BEAF特异的双链RNA还原到眼睛想象盘或翼盘的细胞中时，成人眼或翼的形态似乎是正常的。早期胚胎中BEAF蛋白水平降低导致发育迟缓，1龄幼虫全部死亡。这些结果表明，BEAF可能在发育的早期阶段发挥重要作用，但在2龄幼虫或3龄幼虫之后的后期可能发挥重要作用。我们分析了热休克前后DREF、BEAF和Mi-2在热休克蛋白70(HSP70)基因上的重新合成。众所周知，37℃处理动物在HSP70基因周围建立了特殊的染色质结构，并强烈诱导HSP70基因的瞬时转录，但对参与HSP70基因特殊染色质区域形成和转录激活的因素知之甚少。热休克后，DREF和Mi-2均在HSP70位点迅速募集。现在，我们正在准备一套系统来检查HSP70基因座发生了什么，以及Dref和Mi-2在这一现象中扮演了什么角色。

Transgenic fly lines carrying UAS-BEAF and UAS-BEAF RNAi have been established in addition to the lines carrying UAS-DREF and UAS-Mi-2. Using these lines, over-expression or knock-down of BEAF protein is realized in tissues-specific manner. Over-expression of BEAF protein in cells of the eye-imaginal disc resulted in rough-eye phenotype. On the other hand, when BEAF transcript was reduced by BEAF-specific double-stranded RNA in cells of the eye imaginal disc or wing disc, morphology of adult eye or wing seems to be normal.. Reducing level of BEAF protein in early embryo caused retardation of development and all of animals were dead at first instar larval stage. These result suggest that BEAF might have important roles in early stages of development but in the later stages after 2^<nd> or 3^<rd> -instar larva.Recruitment of DREF, BEAF, and Mi-2 at the genes for heat shock protein 70 (hsp70) were analyzed before and after heat shock.It is well known that treatment of animals at 37 C establishes special chromatin structure around the hsp70 gene and strong and induces transient transcription of the hsp70 gene, but little is known about factors involved in formation of the special chromatin domain and activation of transcription of the hsp70 gene locus. Both DREF and Mi-2 were rapidly recruited at the hsp70 locus after heat shock.. Now, we are preparing system for examining what is happened at the hsp70 locus and what roles DREF and Mi-2 have in that phenomena.