Small regulatory RNAs (sRNAs) in eukaryotes and bacteria play an important role in the regulation of gene expression either by binding to regulatory proteins or directly to target mRNAs. Two of the best-characterized bacterial sRNAs, Spot42 and RyhB, form a complementary pair with the ribosome binding region of their target mRNAs, thereby inhibiting translation or promoting mRNA degradation. To investigate the steady-state and dynamic potential of such sRNAs, we examine the 2 key parameters characterizing sRNA regulation: the capacity to overexpress the sRNA relative to its target mRNA and the speed at which the target mRNA is irreversibly inactivated. We demonstrate different methods to determine these 2 key parameters, for Spot42 and RyhB, which combine biochemical and genetic experiments with computational analysis. We have developed a mathematical model that describes the functional properties of sRNAs with various characteristic parameters. We observed that Spot42 and RyhB function in distinctive parameter regimes, which result in divergent mechanisms.gene regulation ͉ mRNA silencing ͉ RyhB ͉ small RNA ͉ Spot42

Optimal response to environmental stimuli often requires activation of certain genes and repression of others. Dual function regulatory proteins play a key role in the differential regulation of gene expression. While repression can be achieved by any DNA binding protein through steric occlusion of RNA polymerase in the promoter region, activation often requires a surface on the regulatory protein to contact RNAP and thus facilitate transcription initiation. RNAP itself is also a DNA binding protein, therefore it can function as a transcriptional repressor. Searching the Escherichia coli promoter database we found that ∼14% of the identified ‘forward’ promoters overlap with a promoter oriented in the opposite direction. In this article we combine a mathematical model with experimental analysis of synthetic regulatory regions to investigate interference of overlapping promoters. We find that promoter interference depends on the characteristics of overlapping promoters. The model predicts that promoter strength and interference can be regulated separately, which provides unique opportunities for regulation. Our experimental data suggest that in principle any DNA binding protein can be used for both activation and repression of promoter transcription, depending on the context. These findings can be exploited in the construction of synthetic networks.

The lactose operon of Escherichia coli is a paradigm system for quantitative understanding of gene regulation in prokaryotes. Yet, none of the many mathematical models built so far to study the dynamics of this system considered the fact that the Lac repressor regulates its own transcription by forming a transcriptional roadblock at the O3 operator site. Here we study the effect of autoregulation on intracellular LacI levels and also show that cAMP-CRP binding does not affect the efficiency of autoregulation. We built a mathematical model to study the role of LacI autoregulation in the lactose utilization system. Previously, it has been argued that negative autoregulation can significantly reduce noise as well as increase the speed of response. We show that the particular molecular mechanism, a transcriptional roadblock, used to achieve self-repression in the lac system does neither. Instead, LacI autoregulation balances two opposing states, one that allows quicker response to smaller pulses of external lactose, and the other that minimizes production costs in the absence of lactose.