Design of Synthetic Combinatorial Transcription Factors

描述（由申请人提供）：许多生物学决定是由复杂的转录回路控制的。能够专门调节这种电路的试剂将是非常有用的，作为剖析复杂过程的顺式调控逻辑的工具，并最终作为治疗方法。最先进的工具是合成DNA结合蛋白，但这些工具缺乏在天然转录因子中观察到的组合特异性。最终，更精确的监管可以通过一系列协同作用的设计因素来实现。许多天然组合转录因子起着变构开关的作用——该因子只有在另一辅助因子存在时才有活性。最近，一种引入变构控制蛋白质活性的新策略被开发出来。这种策略包括在蛋白质中引入构象自抑制相互作用；Lautoinhibition可以通过与合适的伴侣蛋白结合来缓解。我的具体目标是使用这种自抑制策略来生成合成转录因子，其中：1)DNA结合活性由相邻的DNA结合辅助因子变构调节。2)转录激活功能受邻近DNA结合辅因子的变构调节。这些设计因子应该产生极其严格的转录控制——激活依赖于分子精确组合的同时存在。

DESCRIPTION (provided by applicant): Many biological decisions are controlled by complex transcriptional circuits. Reagents that can specifically modulate such circuits would be extremely useful, as tools for dissecting the cis-regulatory logic of complex processes, and ultimately as therapeutics. The best developed tools are synthetic DNA binding proteins, but these lack the combinatorial specificity observed in natural transcription factors. Ultimately, more precise regulation could be achieved with sets of designer factors that act in concert. Many natural combinatorial transcription factors act as allosteric switches -- the factor is only active in the presence of another co-factor. Recently a novel strategy for introducing allosteric control over a protein activity has been developed. This strategy involves the introduction of conformational autoinhibitory interactions into a protein; lautoinhibition can be relieved by association with an appropriate partner protein. My specific aims are to use Ithis autoinhibitory strategy to generate synthetic transcription factors in which: 1) DNA binding activity is allosterically regulated by an adjacent DNA binding co-factor. 2) Transcriptional activation function is allosterically regulated by an adjacent DNA binding co-factor. These designer factors should yield extremely tight transcriptional control -- activation is dependent on the simultaneous presence of a precise combination of molecules.