Transcriptional Kinetic Synergy: A Complex Landscape Revealed by Integrating Modelling and Synthetic Biology

Martinez-Corral, Rosa; Park, Min Hee; Biette, Kelly M.; Friedrich, Dhana; Scholes, Clarissa; Khalil, Ahmad S.; Gunawardena, Jeremy; DePace, Angela H.

doi:10.2139/ssrn.3985163

Cited by 4 publications

(11 citation statements)

References 71 publications

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“…[29]) or difficulties with TF overexpression. Accordingly, during preliminary work for a previous project [56], we attributed to these some non-monotonic input-output responses we observed. However, the theoretical results presented above suggest an alternative, where non-monotonicity reflects underlying TF incoherent regulation.…”

Section: Experimental Evidence For Non-monotonicity and Affinity-depe...mentioning

confidence: 66%

“…Input TF concentration is controlled by transfection (Fig. S9B), [56], and the transcriptional 10 response by monitoring reporter mRNA levels by qRT-PCR. We first selected a range of synTF plasmid concentration over which increases in input TF does not affect the expression of GAPDH nor p21 (Fig.…”

Section: Experimental Evidence For Non-monotonicity and Affinity-depe...mentioning

confidence: 99%

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Emergence of activation or repression in transcriptional control under a fixed molecular context

Martinez-Corral,

Friedrich,

Frömel

et al. 2024

Preprint

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For decades, studies have noted that transcription factors (TFs) can behave as either activators or repressors of different target genes. More recently, evidence suggests TFs can act on transcription simultaneously in positive and negative ways. Here we use biophysical models of gene regulation to define, conceptualize and explore these two aspects of TF action: “duality”, where TFs can be overall both activators and repressors at the level of the transcriptional response, and “coherent and incoherent” modes of regulation, where TFs act mechanistically on a given target gene either as an activator or a repressor (coherent) or as both (incoherent). For incoherent TFs, the overall response depends on three kinds of features: the TF’s mechanistic effects, the dynamics and effects of additional regulatory molecules or the transcriptional machinery, and the occupancy of the TF on DNA. Therefore, activation or repression can be tuned by just the TF-DNA binding affinity, or the number of TF binding sites, given an otherwise fixed molecular context. Moreover, incoherent TFs can cause non-monotonic transcriptional responses, increasing over a certain concentration range and decreasing outside the range, and we clarify the relationship between non-monotonicity and common assumptions of gene regulation models. Using the mammalian SP1 as a case study and well controlled, synthetically designed target sequences, we find experimental evidence for incoherent action and activation, repression or non-monotonicity tuned by affinity. Our work highlights the importance of moving from a TF-centric view to a systems view when reasoning about transcriptional control.

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Section: Experimental Evidence For Non-monotonicity and Affinity-depe...mentioning

confidence: 66%

Section: Experimental Evidence For Non-monotonicity and Affinity-depe...mentioning

confidence: 99%

Emergence of activation or repression in transcriptional control under a fixed molecular context

Martinez-Corral,

Friedrich,

Frömel

et al. 2024

Preprint

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show abstract

“…We elaborated the techniques previously introduced to estimate ( p, s ) regions like that in Fig.1C [1, 10, 32] to plot the universal ( p, s ) region, Ω m , for input-output responses at t.e. with m input binding sites (Fig.3A).…”

Section: Resultsmentioning

confidence: 99%

“…1C [1,10,32] to plot the universal (p, s) region, Ω m , for input-output responses at t.e. with m input binding sites (Fig.…”

Section: Universal Position-steepness Region and The Hopfield Barriermentioning

confidence: 99%

The Hill function is the universal Hopfield barrier for sharpness of input-output responses

Martinez-Corral,

Nam,

DePace

et al. 2024

Preprint

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The Hill functions, Hh(x) = xh/(1 + xh), have been widely used in biology for over a century but, with the exception of H1, they have had no justification other than as a convenient fit to empirical data. Here, we show that they are the universal limit for the sharpness of any input-output response arising from a Markov process model at thermodynamic equilibrium. Models may represent arbitrary molecular complexity, with multiple ligands, internal states, conformations, co-regulators, etc, under core assumptions that are detailed in the paper. The model output may be any linear combination of steady-state probabilities, with components other than the chosen input ligand held constant. This formulation generalises most of the responses in the literature. We use a coarse-graining method in the graph-theoretic linear framework to show that two sharpness measures for input-output responses fall within an effectively bounded region of the positive quadrant, Ωm⊂ (R+)2, for any equilibrium model with m input binding sites. Ωmexhibits a cusp which approaches, but never exceeds, the sharpness of Hmbut the region and the cusp can be exceeded when models are taken away from thermodynamic equilibrium. Such fundamental thermodynamic limits are called Hopfield barriers and our results provide a biophysical justification for the Hill functions as the universal Hopfield barriers for sharpness. Our results also introduce an object, Ωm, whose structure may be of mathematical interest, and suggest the importance of characterising Hopfield barriers for other forms of cellular information processing.

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“…Synergy might arise at the level of TF binding, either through direct physical interactions between TFs 5 , from a thermodynamic competition between TFs and nucleosomes for DNA occupancy 6 , or through the effects of cofactors (such as chromatin remodelers or acetyltransferases) which aid in nucleosome eviction 7,8 . Alternatively, synergy could be the result of TFs acting on different steps of the transcription cycle and cooperating to drive super-additive expression once bound 9,10 .…”

Section: Introductionmentioning

confidence: 99%

Single-molecule chromatin configurations link transcription factor binding to expression in human cells

Doughty,

Hinks,

Schaepe

et al. 2024

Preprint

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The binding of multiple transcription factors (TFs) to genomic enhancers activates gene expression in mammalian cells. However, the molecular details that link enhancer sequence to TF binding, promoter state, and gene expression levels remain opaque. We applied single-molecule footprinting (SMF) to measure the simultaneous occupancy of TFs, nucleosomes, and components of the transcription machinery on engineered enhancer/promoter constructs with variable numbers of TF binding sites for both a synthetic and an endogenous TF. We find that activation domains enhance a TF's capacity to compete with nucleosomes for binding to DNA in a BAF-dependent manner, TF binding on nucleosome-free DNA is consistent with independent binding between TFs, and average TF occupancy linearly contributes to promoter activation rates. We also decompose TF strength into separable binding and activation terms, which can be tuned and perturbed independently. Finally, we develop thermodynamic and kinetic models that quantitatively predict both the binding microstates observed at the enhancer and subsequent time-dependent gene expression. This work provides a template for quantitative dissection of distinct contributors to gene activation, including the activity of chromatin remodelers, TF activation domains, chromatin acetylation, TF concentration, TF binding affinity, and TF binding site configuration.

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Transcriptional Kinetic Synergy: A Complex Landscape Revealed by Integrating Modelling and Synthetic Biology

Cited by 4 publications

References 71 publications

Emergence of activation or repression in transcriptional control under a fixed molecular context

Emergence of activation or repression in transcriptional control under a fixed molecular context

The Hill function is the universal Hopfield barrier for sharpness of input-output responses

Single-molecule chromatin configurations link transcription factor binding to expression in human cells

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