Molecular origins of transcriptional heterogeneity in diazotrophic K. oxytoca
Molecular origins of transcriptional heterogeneity in diazotrophic K. oxytoca
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DOI：
10.1101/2020.02.18.955476
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发表时间：
2020-02
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通讯作者：
Tufail Bashir;Rowan D. Brackston;Christopher J. Waite;I. Kotta-Loizou;C. Engl;M. Buck;J. Schumacher
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作者：
Tufail Bashir;Rowan D. Brackston;Christopher J. Waite;I. Kotta-Loizou;C. Engl;M. Buck;J. Schumacher
克隆细菌批量培养中的表型异质性是适应环境变化的重要策略，包括具有将双氮转化为生物可利用的铵的独特能力的固氮菌。在固氮型产酸克雷伯菌中，我们使用双分子、单细胞 RNA-FISH 同时测量了建立固氮所需的关键调节操纵子（glnK-amtB、nifLA）和结构操纵子（nifHDK）的 mRNA 水平。通过随机转录模型和互信息分析，我们揭示了固氮酶表达异质性的可能分子起源。在野生型和调控变异菌株中，我们推断了内在和外在噪声的贡献，发现 nifHDK 转录本质上是突发性的，但通过信号传导的噪声传播也很重要。调节基因 glnK 对 nifHDK 方差的影响最大，而来自调节途径之外因素的噪音可以忽略不计。结果证明异质性是该调节系统的基本特性，表明工程同质固氮酶表达的潜在限制。
Phenotypic heterogeneity in clonal bacterial batch cultures is an important adaptive strategy to changing environments, including in diazotrophs with the unique capacity to convert di-nitrogen into bio-available ammonium. In diazotrophic Klebsiella oxytoca we simultaneously measured mRNA levels of key regulatory (glnK-amtB, nifLA) and structural (nifHDK) operons required for establishing nitrogen fixation, using dual molecule, single cell RNA-FISH. Through stochastic transcription models and mutual information analysis we revealed likely molecular origins for heterogeneity in nitrogenase expression. In wildtype and regulatory variant strains we inferred contributions from intrinsic and extrinsic noise, finding that nifHDK transcription is inherently bursty, but that noise propagation through signalling is also significant. The regulatory gene glnK had the highest discernible effect on nifHDK variance, while noise from factors outside of the regulatory pathway were negligible. Results provide evidence that heterogeneity is a fundamental property of this regulatory system, indicating potential constraints for engineering homogeneous nitrogenase expression.