Chromatin condensation at the invisible length scale
Lead Research Organisation:
University of Cambridge
Abstract
Around 30% of the eukaryotic proteome consists of proteins or protein regions that are disordered, but nevertheless have important functions. In biology, we have become accustomed to linking sequence to structure, and structure to function, but where there is no defined structure, we must think of the protein as adopting a dynamic ensemble of conformations. The paradigm therefore becomes sequence -> ensemble -> function, but studying protein ensembles can be very challenging experimentally, as they are heterogeneous and often invisible by diffraction methods or electron microscopy, and can suffer from severe peak overlap in NMR spectroscopy. Another issue is that the linear sequence of disordered regions is often not as well conserved over evolution as their global, physicochemical properties, making it difficult to track, understand and predict the sequence -> ensemble -> function relationships.
Linker histones are abundant nuclear proteins that have a well-established function in the phenomenological sense: they, along with the core histones, bind and compact genomic DNA to make chromatin. Linker histones orchestrate a unique stage of this process: the condensation of the so-called 11nm fibre into a more compact structure, which reduces its transcriptional activity. The extreme endpoint of this process is a thicker, 30nm fibre, which has been observed in transcriptionally insert cells that use specialised linker histones. However, in transcriptionally active cells, the fibre appears more open, flexible and heterogeneous, somewhere in between the 11nm and 30nm fibre states in compaction level, and self-assembling into liquid-like globules. This “top-down” view, from super-resolution imaging, is resonant with our “bottom-up” findings, from NMR and molecular biophysics applied to a minimal in vitro model. Linker histones have a long, disordered C-terminus. We have established that this disorder is preserved when it binds short DNAs, explaining why its structure – termed a “fuzzy complex” – has defied the usual structural biology toolkits. We also discovered that these fuzzy complexes tend to coalesce into dense liquid condensates, which may explain how the dynamics of the 11nm fibre are retained in the liquid globules seen by microscopy. Linker histones therefore appear to function like a DNA “liquid glue”, and understanding exactly how the glue works will be an important step forward in chromatin biology.
Our aim is now to answer two overarching questions:
Do minimal models of linker histone tails and short DNAs accurately recapitulate the region of interest in the context of full chromatin?
What are the factors controlling linker histones in their role as DNA glues, and how are they coded by the protein sequence?
These define our new objectives:
To develop experimental models of full chromatin.
To capture the local structure and dynamics at all stages of condensation.
To measure the viscoelastic properties of the condensed chromatin fibre.
To quantify the thermodynamics of condensation.
To use the results of 2, 3 and 4 to establish the sequence -> ensemble -> function rules that encompass all linker histones.
Achieving these objectives requires a fully joined-up approach of solution- and solid-state NMR, and new biophysical tools. This project is therefore not only about addressing a significant knowledge gap in chromatin biology by characterising a key disordered region, but an example of how integration of various methodologies can produce a picture of a disordered assembly that is far more informative than the individual techniques alone.
Linker histones are abundant nuclear proteins that have a well-established function in the phenomenological sense: they, along with the core histones, bind and compact genomic DNA to make chromatin. Linker histones orchestrate a unique stage of this process: the condensation of the so-called 11nm fibre into a more compact structure, which reduces its transcriptional activity. The extreme endpoint of this process is a thicker, 30nm fibre, which has been observed in transcriptionally insert cells that use specialised linker histones. However, in transcriptionally active cells, the fibre appears more open, flexible and heterogeneous, somewhere in between the 11nm and 30nm fibre states in compaction level, and self-assembling into liquid-like globules. This “top-down” view, from super-resolution imaging, is resonant with our “bottom-up” findings, from NMR and molecular biophysics applied to a minimal in vitro model. Linker histones have a long, disordered C-terminus. We have established that this disorder is preserved when it binds short DNAs, explaining why its structure – termed a “fuzzy complex” – has defied the usual structural biology toolkits. We also discovered that these fuzzy complexes tend to coalesce into dense liquid condensates, which may explain how the dynamics of the 11nm fibre are retained in the liquid globules seen by microscopy. Linker histones therefore appear to function like a DNA “liquid glue”, and understanding exactly how the glue works will be an important step forward in chromatin biology.
Our aim is now to answer two overarching questions:
Do minimal models of linker histone tails and short DNAs accurately recapitulate the region of interest in the context of full chromatin?
What are the factors controlling linker histones in their role as DNA glues, and how are they coded by the protein sequence?
These define our new objectives:
To develop experimental models of full chromatin.
To capture the local structure and dynamics at all stages of condensation.
To measure the viscoelastic properties of the condensed chromatin fibre.
To quantify the thermodynamics of condensation.
To use the results of 2, 3 and 4 to establish the sequence -> ensemble -> function rules that encompass all linker histones.
Achieving these objectives requires a fully joined-up approach of solution- and solid-state NMR, and new biophysical tools. This project is therefore not only about addressing a significant knowledge gap in chromatin biology by characterising a key disordered region, but an example of how integration of various methodologies can produce a picture of a disordered assembly that is far more informative than the individual techniques alone.
Organisations
People |
ORCID iD |
| Katherine Stott (Principal Investigator) |