Considering a possible correlation between the diverse functions of Fis and its large concentration fluctuations in physiological conditions, Fis emerges as an ideal candidate to study the generic effects of TF-concentration variations on protein unbinding kinetics and chromosome condensation.įis is also one of the first TFs identified to undergo a concentration-dependent unbinding via a mechanism referred to as facilitated dissociation (FD) from its nonspecific and specific binding sites. Through bending the DNA or bridging of remote regions of the chromosome, Fis alters the nucleoid structure and stabilizes DNA loops in vitro. Fis can also bind to DNA in a non-specific manner with a lower affinity than that of specific binding.
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During this concentration rise and fall, Fis binds to DNA sequence specifically and participates in the regulation of more than 200 genes (either as an activator or repressor) necessary for growth, replication, and energy metabolism. Fis in vivo has growth-dependent concentration levels, and under nutrient-rich conditions, it can increase its cellular concentration level from less than a hundred to 60.000 copies per cell and returns its initial limits by the end of the exponential phase. coli) nucleoid binding proteins, and it functions both as a TF and a structural protein. However, whether such a concentration-dependent mechanism can be functional in cellular conditions for structural TFs such as Fis, which has the ability to significantly alter the chromosome structure in a concentration-dependent manner while undergoing large concentration fluctuations, has been elusive.įis is one of the most extensively studied dual-purpose Escherichia coli ( E. The single-molecule tracking experiments in living cells with relatively low-abundance metalloregulatory TFs such as CueR and ZuR have provided some evidence in this direction. Independently, the recent in vitro single-molecule experiments show that off-rates (i.e., reciprocal of the residence time) of major bacterial TF proteins, including the factor for inversion stimulus (Fis), which participates in a variety of cellular functions, accelerate as the concentration of solution-phase proteins increases, suggesting that cellular concentration level of a TF could operate as a regulatory feedback mechanism. However, a growing body of work has demonstrated that the time that a TF spends along the genome (i.e., residence time) could regulate the TF functions, and this effect is more dramatic if the TF has diverse biological functionality. The formation of a DNA-TF complex upon binding is often considered sufficient for the biomolecular functionality of the corresponding TF (i.e., activation or repression of the target gene). Overall, our results indicate that cellular-concentration levels of a structural DNA-binding protein is intermingled with the genome architecture and DNA residence times, thereby providing a basis for understanding the complex effects of dynamic protein-DNA interactions on gene regulation.īacterial transcription factors (TFs) are DNA-binding proteins that can dynamically regulate RNA polymerase’s involvement in the transcription machinery by transiently binding to DNA. As a result, at the physiologically observed maximum levels of Fis, the off-rates significantly slow down. However, Fis significantly changes the chromosome structure at higher concentrations by forming dense protein clusters bridging specific sites and juxtaposing remote DNA segments. Particularly, when nutrient-rich conditions are emulated with Fis concentrations around micromolar levels, the off-rates increase one order of magnitude compared to the lower Fis levels.
Our simulations show that FD of Fis can well occur in confinement at physiological concentrations. coli, which is a dual-purpose protein with a diverse functionality, we model the unbinding of Fis from specific bindings sites along a high-molecular-weight circular DNA in a cylindrical structure mimicking the cellular confinement of chromosome. In this study, inspired by the previous single-molecule studies on the factor for inversion stimulation (Fis) protein of E. In the meantime, single-molecule experiments showed that the off-rates of a wide array of DNA-binding proteins accelerate as the bulk concentration of the protein increases via a concentration-dependent mechanism (i.e., facilitated dissociation, FD). Recent experimental studies demonstrate that residence time (i.e., inverse off-rate) of a transcription factor protein can be a contributor to the functional diversity of the protein. Transcription machinery ultimately depends on the temporal formation of protein-DNA complexes.
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