SMFnet reveals principles of combinatorial transcription factor assembly

Combinatorial transcription factors (TFs) assembly plays a crucial role in opening chromatin at specific regions known as cis-regulatory elements (cREs). This opening enables cell-type-specific gene expression. The variability in TF binding-site arrangements on cREs has made it challenging to understand individual TF functions and their collective behavior.

Researchers have discovered that a single regulatory sequence can support various configurations of TF and nucleosome occupancy, suggesting that different chromatin states may correspond to distinct TF assemblies. However, predictive models that link regulatory sequences to chromatin accessibility average out these diverse molecular states, making it difficult to determine how specific TF assemblies influence accessible chromatin.

To address this issue, scientists have developed a new tool called SMFNet, a generative diffusion model capable of predicting distributions of single-molecule chromatin accessibility profiles based on DNA sequence. Complementing SMFNet is PRISM, a tool that identifies sequence features associated with individual molecular states.

By combining these two approaches, researchers can systematically link single-molecule chromatin states at cREs with specific TF binding motifs. Altering these motifs in the laboratory can redistribute the corresponding chromatin states, as predicted by SMFNet, demonstrating their interpretation as TF-dependent configurations underlying chromatin opening.

Further investigation through controlled in silico motif screens has allowed researchers to dissect individual and combinatorial TF contributions to chromatin opening. Each TF exhibits a unique chromatin-opening signature, characterized by different accessible-region widths and nucleosome-positioning precision, even when associated with similar levels of chromatin accessibility.

The combination of motifs reveals partner- and spacing-dependent effects on chromatin accessibility, TF occupancy, and nucleosome positioning. Genetic perturbation of TF binding sites using variations found in endogenous cREs confirms that specific TF properties shape chromatin accessibility in living organisms.

In conclusion, these findings shed light on the sequence-encoded principles of combinatorial TF assembly during chromatin opening, providing valuable insights into the molecular mechanisms governing cell-type-specific gene expression.