Occlusion of Regulatory Sequences by Promoter Nucleosomes In Vivo

Nucleosomes are believed to inhibit DNA binding by transcription factors. Theoretical attempts to understand the significance of nucleosomes in gene expression and regulation are based upon this assumption. However, nucleosomal inhibition of transcription factor binding to DNA is not complete. Rather, access to nucleosomal DNA depends on a number of factors, including the stereochemistry of transcription factor-DNA interaction, the in vivo kinetics of thermal fluctuations in nucleosome structure, and the intracellular concentration of the transcription factor. In vitro binding studies must therefore be complemented with in vivo measurements. The inducible PHO5 promoter of yeast has played a prominent role in this discussion. It bears two binding sites for the transcriptional activator Pho4, which at the repressed promoter are positioned within a nucleosome and in the linker region between two nucleosomes, respectively. Earlier studies suggested that the nucleosomal binding site is inaccessible to Pho4 binding in the absence of chromatin remodeling. However, this notion has been challenged by several recent reports. We therefore have reanalyzed transcription factor binding to the PHO5 promoter in vivo, using ‘chromatin endogenous cleavage’ (ChEC). Our results unambiguously demonstrate that nucleosomes effectively interfere with the binding of Pho4 and other critical transcription factors to regulatory sequences of the PHO5 promoter. Our data furthermore suggest that Pho4 recruits the TATA box binding protein to the PHO5 promoter

Dissecting Nucleosome Free Regions by a Segmental Semi-Markov Model

BACKGROUND: Nucleosome free regions (NFRs) play important roles in diverse biological processes including gene regulation. A genome-wide quantitative portrait of each individual NFR, with their starting and ending positions, lengths, and degrees of nucleosome depletion is critical for revealing the heterogeneity of gene regulation and chromatin organization. By averaging nucleosome occupancy levels, previous studies have identified the presence of NFRs in the promoter regions across many genes. However, evaluation of the quantitative characteristics of individual NFRs requires an NFR calling method. METHODOLOGY: In this study, we propose a statistical method to identify the patterns of NFRs from a genome-wide measurement of nucleosome occupancy. This method is based on an appropriately designed segmental semi-Markov model, which can capture each NFR pattern and output its quantitative characterizations. Our results show that the majority of the NFRs are located in intergenic regions or promoters with a length of about 400-600bp and varying degrees of nucleosome depletion. Our quantitative NFR mapping allows for an investigation of the relative impacts of transcription machinery and DNA sequence in evicting histones from NFRs. We show that while both factors have significant overall effects, their specific contributions vary across different subtypes of NFRs. CONCLUSION: The emphasis of our approach on the variation rather than the consensus of nucleosome free regions sets the tone for enabling the exploration of many subtler dynamic aspects of chromatin biology

Adjusting Phenotypes by Noise Control

Genetically identical cells can show phenotypic variability. This is often caused by stochastic events that originate from randomness in biochemical processes involving in gene expression and other extrinsic cellular processes. From an engineering perspective, there have been efforts focused on theory and experiments to control noise levels by perturbing and replacing gene network components. However, systematic methods for noise control are lacking mainly due to the intractable mathematical structure of noise propagation through reaction networks. Here, we provide a numerical analysis method by quantifying the parametric sensitivity of noise characteristics at the level of the linear noise approximation. Our analysis is readily applicable to various types of noise control and to different types of system; for example, we can orthogonally control the mean and noise levels and can control system dynamics such as noisy oscillations. As an illustration we applied our method to HIV and yeast gene expression systems and metabolic networks. The oscillatory signal control was applied to p53 oscillations from DNA damage. Furthermore, we showed that the efficiency of orthogonal control can be enhanced by applying extrinsic noise and feedback. Our noise control analysis can be applied to any stochastic model belonging to continuous time Markovian systems such as biological and chemical reaction systems, and even computer and social networks. We anticipate the proposed analysis to be a useful tool for designing and controlling synthetic gene networks

Chromatin remodeling by the SWI/SNF complex is essential for transcription mediated by the yeast cell wall integrity MAPK pathway.

In Saccharomyces cerevisiae, the transcriptional program triggered by cell wall stress is coordinated by Slt2/Mpk1, the mitogen-activated protein kinase (MAPK) of the cell wall integrity (CWI) pathway, and is mostly mediated by the transcription factor Rlm1. Here we show that the SWI/SNF chromatin-remodeling complex plays a critical role in orchestrating the transcriptional response regulated by Rlm1. swi/snf mutants show drastically reduced expression of cell wall stress-responsive genes and hypersensitivity to cell wall-interfering compounds. On stress, binding of RNA Pol II to the promoters of these genes depends on Rlm1, Slt2, and SWI/SNF. Rlm1 physically interacts with SWI/SNF to direct its association to target promoters. Finally, we observe nucleosome displacement at the CWI-responsive gene MLP1/KDX1, which relies on the SWI/SNF complex. Taken together, our results identify the SWI/SNF complex as a key element of the CWI MAPK pathway that mediates the chromatin remodeling necessary for adequate transcriptional response to cell wall stress