Nuclear architecture: structure, function and disease

Recently scientists have developed tools for identifying mutations in individuals with complex neuro-development disorders. Surprisingly, many of these mutations are found in proteins that have a role in fundamental cellular functions. To understand the molecular basis of these diseases, with a hope of developing new prognostic markers and potential disease treatments, it is crucial to understand how the specific proteins function in the complex environment of the cell. My team is expert in analysing how DNA is packaged inside part of a human cell called the nucleus and we hypothesise that in some individuals their disease is caused by altered packaging of DNA so it does not function correctly. However, there are many steps to this process from packaging DNA to how the DNA is read to make RNA and then how the RNA is folded and exported to other parts of the cell to make protein. In this study we will focus on one specific protein, HNRNPU, that when mutated causes learning difficulties, autism, and epilepsy. So far, our research has shown that HNRNPU interacts with another molecule in the nucleus called RNA and together they form a gel that surrounds the DNA, similar to protective cotton wool. Together this gel facilitates processes that occur on the DNA such as reading into RNA, or repairing damage to the DNA. We will study how mutations in HNRNPU alter DNA packaging and how this affects many important cellular processes and disease.

In mammalian cells nuclear architecture is underpinned by a mesh comprised of proteins and RNA, that is often considered to be relatively passive. However, recent evidence suggests that structural proteins, such as HNRNPU (also called Scaffold Attachment Factor A; SAF-A), interact with chromatin-associated RNA to form a series of interconnected clusters, creating a dynamic nuclear environment, but little is known about its organisation, regulation and function. We hypothesise that the ribonucleoprotein mesh creates an active environment that facilitates critical processes such as transcription, chromatin compaction and RNA processing, but when misregulated causes disease.

HNRNPU is an abundant evolutionarily conserved protein that is required for normal development across multiple organisms. Our preliminary data shows that it interacts with additional members of the hnRNP family including HNRNPUL1 and HNRNPUL2, and other structural proteins. Recently mutations in HNRNPU have been identified as causing HNRNPU-related disorder, with symptoms that include learning difficulties, autism, and epilepsy, but the molecular link between protein function and phenotype is unknown.

To develop a new paradigm for understanding the mechanisms linking nuclear architecture and function our aim is to investigate how a nuclear mesh can create an active environment that facilitates nuclear processes and how alterations in this milieu can cause disease. To achieve this our specific objectives are:
1. Investigate the molecular basis of a nuclear mesh
To understand how a nuclear mesh is formed we will analyse the structure of HNRNPU and its homologues and investigate how they interact with RNA in a cell model. We will then map the positions of clinical mutations onto HNRNPU to identify critical parts of the protein. Using super-resolution and advanced imaging techniques we will assess how HNRNPU mutations affect the structure of a nuclear mesh and how they alter its biophysical properties. Finally, by mutating HNRNPU and its homologues to disrupt the nuclear mesh we will determine how this microenvironment is necessary for chromatin structure, transcription and RNA processing.
2. How do HNRNPU mutations affect neuronal function and cause disease?
We hypothesise that the molecular basis for HNRNPU-related disorder is a disrupted or disorganised nuclear mesh that affects chromatin folding or RNA processing. To test this we will analyse chromatin structure, RNA processing and concomitant protein synthesis in a model of induced neuronal cells that have mutations in HNRNPU.

Genetic analysis of individuals with neuro-developmental disorders indicates that many of the mutations occur in proteins important for regulating fundamental nuclear processes. The molecular basis for this is not clear, which severely affects the ability of clinicians to make a clear prognosis for patients. Furthermore, without knowing the root cause of the disease it is not possible to develop potential treatments. This fundamental research will investigate mutations in a specific type of protein, HNRNPU, to provide a paradigm for understanding the molecular basis of diseases affecting chromatin packaging and nuclear organisation and provide new insight for the future development of prognostic markers and disease treatments.