Lakhotia Subhash C, Mallik Moushami, Singh Anand K, Ray Mukulika
Cytogenetics Laboratory, Department of Zoology, Banaras Hindu University, Varanasi 221005, India.
Chromosoma. 2012 Feb;121(1):49-70. doi: 10.1007/s00412-011-0341-x. Epub 2011 Sep 9.
The hs-GAL4(t)-driven expression of the hsrω-RNAi transgene or EP93D allele of the noncoding hsrω resulted in global down- or upregulation, respectively, of the large hsrω-n transcripts following heat shock. Subsequent to temperature shock, hsrω-null or those expressing hsrω-RNAi or the EP93D allele displayed delayed lethality of most embryos, first or third instar larvae. Three-day-old hsrω-null flies mostly died immediately or within a day after heat shock. Heat-shock-induced RNAi or EP expression in flies caused only a marginal lethality but severely affected oogenesis. EP allele or hsrω-RNAi expression after heat shock did not affect heat shock puffs and Hsp70 synthesis. Both down- and upregulation of hsrω-n transcripts suppressed reappearance of the hsrω-n transcript-dependent nucleoplasmic omega speckles during recovery from heat shock. Hrp36, heterochromatin protein 1, and active RNA pol II in unstressed or heat-shocked wild-type or hsrω-null larvae or those expressing the hs-GAL4(t)-driven hsrω-RNAi or the EP93D allele were comparably distributed on polytene chromosomes. Redistribution of these proteins to pre-stress locations after a 1- or 2-h recovery was severely compromised in glands with down- or upregulated levels of hsrω-n transcripts after heat shock. The hsrω-null unstressed cells always lacked omega speckles and little Hrp36 moved to any chromosome region following heat shock, and its relocation to chromosome regions during recovery was also incomplete. This present study reveals for the first time that the spatial restoration of key regulatory factors like hnRNPs, HP1, or RNA pol II to their pre-stress nuclear targets in cells recovering from thermal stress is dependent upon critical level of the large hsrω-n noncoding RNA. In the absence of their relocation to pre-stress chromosome sites, normal developmental gene activity fails to be restored, which finally results in delayed organismal death.
由hs-GAL4(t)驱动的hsrω-RNAi转基因或非编码hsrω的EP93D等位基因的表达，分别导致热休克后大型hsrω-n转录本的整体下调或上调。温度休克后，hsrω缺失或表达hsrω-RNAi或EP93D等位基因的个体，大多数胚胎、一龄或三龄幼虫表现出延迟致死性。三日龄的hsrω缺失果蝇大多在热休克后立即或一天内死亡。热休克诱导的果蝇RNAi或EP表达仅导致轻微致死性，但严重影响卵子发生。热休克后EP等位基因或hsrω-RNAi表达不影响热休克胀泡和Hsp70合成。hsrω-n转录本的下调和上调均抑制了热休克恢复过程中hsrω-n转录本依赖性核质ω斑点的重新出现。在未受应激或热休克的野生型或hsrω缺失幼虫，或表达由hs-GAL4(t)驱动的hsrω-RNAi或EP93D等位基因的幼虫中，Hrp36、异染色质蛋白1和活性RNA聚合酶II在多线染色体上的分布相当。热休克后hsrω-n转录本水平下调或上调的腺体中，这些蛋白质在1或2小时恢复后重新分布到应激前位置的过程受到严重损害。未受应激的hsrω缺失细胞总是缺乏ω斑点，热休克后很少有Hrp36迁移到任何染色体区域，其在恢复过程中向染色体区域的重新定位也不完整。本研究首次揭示，从热应激恢复的细胞中，关键调节因子如hnRNPs、HP1或RNA聚合酶II向其应激前核靶点的空间恢复，依赖于大型hsrω-n非编码RNA的临界水平。在它们未重新定位到应激前染色体位点的情况下，正常的发育基因活性无法恢复，最终导致生物体延迟死亡。
