Non-canonical role of roX RNA in X-chromosome targeting

果蝇的X染色体剂量补偿过程保证了两性基因组表达的相似性。雄性果蝇只有在其单个X染色体上的基因转录在两倍的Fange中得到增强的情况下才能存活。探索这一高度进化过程的机制基础，继续揭示基因组调控的基本原则。雄性果蝇X染色体基因的激活是通过剂量补偿复合体(DCC)实现的，DCC是一个由5个雄性特异性致死(MSL)蛋白和长长的非编码Rox RNA组成的多酶组合。最初，DCC与X染色体的高度选择性结合是忠实剂量补偿的关键。这一过程的损害会导致不分青红皂白的染色体结合，这对受影响的雄性果蝇是致命的。ROX RNA和介导该lncRNA组装到DCC的解旋酶MLE的作用尚不清楚。DCC实际上是极少数与染色质结合的lncRNPs之一，其所有RNA和蛋白质亚单位都是已知的，因此为详细研究lncRNA的功能提供了巨大的机会。我们最近发现了roX2 RNA在X染色体选择性中的重要作用。奇怪的是，这类RNA最直观的功能似乎并不适用于Rox。ROX显然不能以三螺旋或R环的形式读出Dann序列。ROX也不能作为促进复合体形成的支架，因为MSL蛋白可以在没有RNA的情况下形成活性复合体。我们还发现，在ROX存在的情况下，DCC与X染色体的特异性结合与采用紧密结合模式有关，这在FRAP实验中被异常长的停留时间所证明。我们试图揭示新的原理和潜在的分子机制，通过这些原理和潜在的分子机制，Rox RNAs促进MSL复合体与X染色体的强健和排他性结合，并为X染色体上DCC的“紧结合”状态奠定基础。

The process of X chromosome dosage compensation in Drosophila melanogaster assures that genomes of both sexes are expressed similarly. Male flies can only live if the transcription of genes on their single X chromosome is boosted in the two-fold fange. Exploring the mechanistic underpinnings of this highly evolved process continues to uncover fundamental principles of genome regulation. The activation of X chromosomal genes in male flies is achieved by the dosage compensation complex (DCC), a multi-enzyme assembly consisting of five male-specific-lethal (MSL) proteins and long, non-coding roX RNA. The initial, highly selective binding of the DCC to the X chromosome is key to faithful dosage compensation. Impairment of this process leads to indiscriminate chromosome binding, which is lethal for affected male flies. The role of roX RNA and the helicase MLE that mediates assembly of this lncRNA into the DCC is poorly understood. The DCC is actually one of the very few chromatin-bound lncRNPs, for which all RNA and protein subunits are known, thus providing a huge opportunity to investigate the function of a lncRNA in detail. We recently discovered an important role for roX2 RNA in X chromosome selectivity. Curiously, the most intuitive functions for such RNAs appear not to apply to roX. RoX apparently does not read out DANN sequence by triple-helix or R-loop formation. RoX also does not function as a scaffold to facilitate complex formation, since the MSL proteins can from active complexes in the absence of RNA. We also found that the specific binding of the DCC to the X chromosome in the presence of roX correlated with the adoption of a tight-binding mode, documented by unusually long residence times in FRAP experiments. We seek to uncover novel principles and underlying molecular mechanism, through which roX RNAs promote the robust and exclusive binding of the MSL complex to the X chromosome, and which underlie the ‘tight-binding’ state of the DCC on the X chromosome.