Paraspeckles are ribonucleoprotein bodies found in mammalian cell nuclei, which play a role in the regulation of gene expression by retention of RNA. The core proteins of paraspeckles are Drosophila behavior/ human splicing (DBHS) proteins SFPQ, NONO and PSPC1. These proteins have essential cellular functions and genetic defects that affect DBHS proteins can cause severe phenotypes, such as cancer progression. Two approaches are being taken to investigate functional interactions of DBHS proteins.

Firstly, SFPQ is a human tumor suppressor protein that has the ability to form linear polymer molecules via its extended coiled-coil interactions, which is important for nucleic acid binding and gene regulation. We are studying polymerization of SFPQ by creating truncated protein constructs at the coiled-coil region and mutating key residues.  SFPQ coiled-coil interactions will be studied by Small-Angle X-Ray scattering (SAXS) and transmission electron microscopy (TEM).

Secondly, interactions between two of the core domains of DBHS proteins, termed NOPS and RRM2, dictate the formation of functional homo- and heterodimers. The RRM2 is a non-canonical RNA recognition motif where three typically conserved residues are substituted. This indicates that RRM2 either does not bind RNA or binds it in an unexpected manner. We have designed NOPS-RRM2 fusion protein constructs in order to study RNA binding and with a view to obtaining a high resolution crystal structure.