Poster Presentation The 43rd Lorne Conference on Protein Structure and Function 2018

Structural and functional analysis of the C. elegans TRIM protein NHL-2 (#115)

Rhys N Colson 1 , Peter R Boag 1 , Jacqueline A Wilce 1
  1. Biochemistry and Molecular Biology, Monash Biomedicine Discovery Institute , Melbourne, VIC, Australia

TRIM-NHL proteins play a role in various cellular functions from muscular and neuronal tissue development to controlling stem cell fate. While the NHL domain of these proteins was originally thought to facilitate protein-protein interactions, it has recently been discovered that several TRIM-NHL proteins have sequence specific interactions with RNA through the domain. The C. elegans protein NHL-2 is involved in small RNA pathways, interacting with the DEAD-box RNA helicase’s CGH-1 and DRH-3. I have shown that, like other TRIM-NHL proteins, NHL-2 binds directly to U-rich RNA and identified residues key to RNA binding, which when mutated show a significant reduction in affinity. These mutations will be tested in vivo, where the phenotypes of RNA binding mutant worms will be compared to nhl-2 knock out, assessing the biological importance of NHL-2’s RNA binding.

Structural analysis of NHL-2 is also being undertaken, with aim to use using X-ray crystallography to solve the structure of the NHL domain both apo and in complex with RNA. Currently the only solved structure of the NHL domain is from the drosophila protein BRAT. Solving the structure of NHL-2’s NHL domain will provide insight into the structural differences between TRIM-NHL proteins that underlie their functional differences. SAXS is also being used to investigate the quaternary structure of NHL-2, looking at the way in which homodimerization, that is facilitated by the coiled coil domain, affects the positioning of other domains.