Ribosomal DNA promoter recognition is determined in vivo by cooperation between UBTF1 and SL1 and is compromised in the UBTF-E210K neuroregression syndrome.
Ribosomal DNA promoter recognition is determined in vivo by cooperation between UBTF1 and SL1 and is compromised in the UBTF-E210K neuroregression syndrome.
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DOI：
10.1371/journal.pgen.1009644
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发表时间：
2022-03
期刊：
影响因子：
4.5
通讯作者：
Moss T
中科院分区：
文献类型：
--
作者：
Tremblay MG;Sibai DS;Valère M;Mars JC;Lessard F;Hori RT;Khan MM;Stefanovsky VY;LeDoux MS;Moss T
通过RNA聚合酶I（RPI/PolR 1）转录约200个小鼠和人类核糖体RNA基因（rDNA），占总细胞RNA的80%，约占所有核RNA合成的35%，并决定细胞质核糖体互补。因此，它是控制细胞生长的主要因素，其功能障碍与肥大和发育障碍有关。每个rDNA重复序列的激活需要核小体被结构性多HMGbox因子UBTF替换，以产生15.7 kbp的无核小体区（NFR）。该NFR的形成对于TBP-TAFI因子SL 1的募集和在rDNA的基因和增强子相关启动子处的前起始复合物（PIC）形成也是必不可少的。然而，这些启动子几乎没有显示出序列共性，并且UBTF和SL 1都没有显示出显著的DNA序列结合特异性，使得驱动PIC形成的原因成为一个谜。在这里，我们表明，SL 1和较长的UBTF 1剪接变体之间的合作产生的特异性所需的rDNA启动子识别细胞。我们发现，SL 1的TAF 1B亚基的条件性缺失导致两个rDNA启动子处的UBTF的显著耗尽，但不在rDNA的其他地方。我们还发现，虽然UBTF 1和UBTF-2变体都结合在整个rDNA NFR中，但只有UBTF 1与启动子处的SL 1一起存在。这些数据有力地表明了一个诱导拟合模型的RPI启动子识别，其中UBTF 1发挥了建筑作用。有趣的是，复发性UBTF-E210 K突变和儿科神经退行性综合征的原因为该模型提供了间接支持。E210 K敲入细胞显示UBTF 1剪接变体的水平增加，并且伴随着活性rDNA拷贝的增加。相反，它们还显示减少的rDNA转录和启动子募集SL 1。因此，我们认为UBTF-E210 K综合征的根本原因是合作UBTF 1-SL 1启动子募集的减少，这可能部分由增强的rDNA激活补偿。200个核糖体RNA基因拷贝（rDNA）占总细胞RNA的80%，决定了细胞的核糖体补体，因此也决定了其合成蛋白质的能力。因此，这些基因直接控制细胞生长，它们的功能失调导致肥大和发育障碍。然而，目前还不清楚是什么决定了哪些rDNA拷贝被激活的选择，以及这与转录前起始复合物的形成有何关系。我们的研究基于TBP复合物SL 1的一个亚基TAF 1B的条件性缺失，揭示了DNA序列非特异性HMGbox因子UBTF如何在SL 1募集和preinitiation复合物形成中发挥序列特异性作用。我们进一步展示了E210 K复发性小儿神经退行性综合征的原因突变是如何降低这种作用的。这些数据表明，诱导适合模型preinitiation复合物的形成，其中UBTF创建一个DNA折叠，是由SL 1特异性识别。
Transcription of the ~200 mouse and human ribosomal RNA genes (rDNA) by RNA Polymerase I (RPI/PolR1) accounts for 80% of total cellular RNA, around 35% of all nuclear RNA synthesis, and determines the cytoplasmic ribosome complement. It is therefore a major factor controlling cell growth and its misfunction has been implicated in hypertrophic and developmental disorders. Activation of each rDNA repeat requires nucleosome replacement by the architectural multi-HMGbox factor UBTF to create a 15.7 kbp nucleosome free region (NFR). Formation of this NFR is also essential for recruitment of the TBP-TAFI factor SL1 and for preinitiation complex (PIC) formation at the gene and enhancer-associated promoters of the rDNA. However, these promoters show little sequence commonality and neither UBTF nor SL1 display significant DNA sequence binding specificity, making what drives PIC formation a mystery. Here we show that cooperation between SL1 and the longer UBTF1 splice variant generates the specificity required for rDNA promoter recognition in cell. We find that conditional deletion of the TAF1B subunit of SL1 causes a striking depletion of UBTF at both rDNA promoters but not elsewhere across the rDNA. We also find that while both UBTF1 and -2 variants bind throughout the rDNA NFR, only UBTF1 is present with SL1 at the promoters. The data strongly suggest an induced-fit model of RPI promoter recognition in which UBTF1 plays an architectural role. Interestingly, a recurrent UBTF-E210K mutation and the cause of a pediatric neurodegeneration syndrome provides indirect support for this model. E210K knock-in cells show enhanced levels of the UBTF1 splice variant and a concomitant increase in active rDNA copies. In contrast, they also display reduced rDNA transcription and promoter recruitment of SL1. We suggest the underlying cause of the UBTF-E210K syndrome is therefore a reduction in cooperative UBTF1-SL1 promoter recruitment that may be partially compensated by enhanced rDNA activation. The 200 Ribosomal RNA gene copies (the rDNA) account for 80% of total cellular RNA and determine the cell’s ribosome complement and hence also its capacity to synthesize proteins. As a consequence, these genes directly control cell growth and their misfunction leads both to hypertrophic and developmental disorders. However, it is still unclear what determines the choice of which rDNA copies are activated and how this relates to the formation of the transcription preinitiation complex. Our study, based on the conditional deletion of TAF1B, a subunit of the TBP complex SL1, reveals how the DNA sequence non-specific HMGbox factor UBTF plays a sequence-specific role in SL1 recruitment and preinitiation complex formation. We further show how this role is degraded by a mutation that is the cause of the E210K recurrent pediatric neuroregression syndrome. The data suggest an induced-fit model of preinitiation complex formation in which UBTF creates a DNA folding that is specifically recognized by SL1.
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