Domain-Selective BET Ligands Yield Next-Generation Synthetic Genome Readers/Regulators with Nonidentical Cellular Functions
Domain-Selective BET Ligands Yield Next-Generation Synthetic Genome Readers/Regulators with Nonidentical Cellular Functions
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DOI：
10.1021/jacs.3c06297
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发表时间：
2023-11-03
影响因子：
15
通讯作者：
Ansari，Aseem Z.
中科院分区：
文献类型：
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作者：
Mohammed，Ashraf;Waddell，M. Brett;Ansari，Aseem Z.
SynTEF1是一种原型合成基因组读取器/调节剂（SynGR），旨在靶向GAA三联体重复序列并恢复弗里德赖希共济失调患者frataxin （FXN）的表达。它通过使用序列选择性dna结合聚酰胺，通过泛bet配体（JQ1）将BRD4招募到GAA重复序列，从而实现了这一复杂的任务。当以这种方式结合到特定的基因组位点时，JQ1作为乙酰赖氨酸残基的化学假体，这些残基是溴端和外端结构域（BET）蛋白中两个串联溴结构域（BD1和BD2）的天然靶点。随着下一代BET配体的公开，我们在SynGR设计中测试了一组具有改进的物理化学、药理学和溴域选择性的配体作为JQ1的替代品。在这里，我们报告了两个意想不到的发现：(1)携带pan-BET或bd2选择性配体的syngr允许在fxnlocus上转录，而那些携带bd1选择性配体的syngr则不允许；(2)非束缚的bd1选择性配体（GSK778）大大增强了所有活性syngr的活性，而不是中性或抑制性的。bd1选择性SynGRs招募BRD4/BET蛋白的失败表明，活性SynGRs不是作为“表观遗传/染色质模拟物”发挥作用，而是模仿天然转录因子通过BD2结合参与BET蛋白的功能。此外，SynGRs与BD1选择性配体共处理后的活性增强表明，天然转录因子竞争有限的非染色质结合的BET蛋白，阻断BD1可使泛BET配体更有效地与BD2结合。综上所述，SynGRs作为化学探针提供了对自然因素精确调节基因表达的分子识别原理的独特见解，并指导设计更复杂的合成基因调节剂，具有更大的治疗潜力。
SynTEF1, a prototype synthetic genome reader/regulator (SynGR), was designed to target GAA triplet repeats and restore the expression of frataxin (FXN) in Friedreich’s ataxia patients. It achieves this complex task by recruiting BRD4, via a pan-BET ligand (JQ1), to the GAA repeats by using a sequence-selective DNA-binding polyamide. When bound to specific genomic loci in this way, JQ1 functions as a chemical prosthetic for acetyl-lysine residues that are natural targets of the two tandem bromodomains (BD1 and BD2) in bromo- and extra-terminal domain (BET) proteins. As next-generation BET ligands were disclosed, we tested a select set with improved physicochemical, pharmacological, and bromodomain-selective properties as substitutes for JQ1 in the SynGR design. Here, we report two unexpected findings: (1) SynGRs bearing pan-BET or BD2-selective ligands license transcription at theFXNlocus, whereas those bearing BD1-selective ligands do not, and (2) rather than being neutral or inhibitory, an untethered BD1-selective ligand (GSK778) substantively enhances the activity of all active SynGRs. The failure of BD1-selective SynGRs to recruit BRD4/BET proteins suggests that rather than functioning as “epigenetic/chromatin mimics,” active SynGRs mimic the functions of natural transcription factors in engaging BET proteins through BD2 binding. Moreover, the enhanced activity of SynGRs upon cotreatment with the BD1-selective ligand suggests that natural transcription factors compete for a limited pool of nonchromatin-bound BET proteins, and blocking BD1 directs pan-BET ligands to more effectively engage BD2. Taken together, SynGRs as chemical probes provide unique insights into the molecular recognition principles utilized by natural factors to precisely regulate gene expression, and they guide the design of more sophisticated synthetic gene regulators with greater therapeutic potential.