Nonlatching positive feedback enables robust bimodality by decoupling expression noise from the mean

Fundamental to biological decision-making is the ability to generate bimodal expression patterns where 2 alternate expression states simultaneously exist. Here, we use a combination of single-cell analysis and mathematical modeling to examine the sources of bimodality in the transcriptional program controlling HIV’s fate decision between active replication and viral latency. We find that the HIV transactivator of transcription (Tat) protein manipulates the intrinsic toggling of HIV’s promoter, the long terminal repeat (LTR), to generate bimodal ON-OFF expression and that transcriptional positive feedback from Tat shifts and expands the regime of LTR bimodality. This result holds for both minimal synthetic viral circuits and full-length virus. Strikingly, computational analysis indicates that the Tat circuit’s noncooperative “nonlatching” feedback architecture is optimized to slow the promoter’s toggling and generate bimodality by stochastic extinction of Tat. In contrast to the standard Poisson model, theory and experiment show that nonlatching positive feedback substantially dampens the inverse noise-mean relationship to maintain stochastic bimodality despite increasing mean expression levels. Given the rapid evolution of HIV, the presence of a circuit optimized to robustly generate bimodal expression appears consistent with the hypothesis that HIV’s decision between active replication and latency provides a viral fitness advantage. More broadly, the results suggest that positive-feedback circuits may have evolved not only for signal amplification but also for robustly generating bimodality by decoupling expression fluctuations (noise) from mean expression levels.生成双峰表达模式——即同时存在两种交替的表达状态——是生物决策的核心基础之一。本研究结合单细胞分析与数学建模，探究了调控人类免疫缺陷病毒（HIV）在活跃复制与病毒潜伏（viral latency）之间命运抉择的转录程序中，双峰性的来源。研究发现，HIV转录反式激活因子（transactivator of transcription, Tat）可调控HIV启动子——长末端重复序列（long terminal repeat, LTR）——的固有切换过程，从而生成ON-OFF双峰表达；且Tat介导的转录正反馈可偏移并拓宽LTR双峰性的发生区间。该结论在最小合成病毒回路与完整病毒体系中均成立。令人瞩目的是，计算分析显示，Tat回路的非协同“非锁存（nonlatching）”反馈架构经过优化，可通过Tat的随机消亡减缓启动子的切换过程，从而催生双峰表达。与经典泊松模型（Poisson model）不同，理论与实验结果均表明，非锁存型正反馈可显著抑制噪声与均值的反向关联，从而在表达均值持续升高的情况下仍维持随机双峰性。鉴于HIV的快速进化特性，该回路被优化以稳健生成双峰表达的现象，与“HIV在活跃复制与潜伏间的抉择可赋予病毒适合度优势”的假说相一致。从更广泛的视角来看，本研究结果提示，正反馈回路的进化意义不仅在于信号放大，还可通过将表达波动（噪声）与表达均值解耦，稳健地生成双峰表达模式。