Solution structure and behaviour of the Arabidopsis thaliana HYL1 protein.

In plants, microRNA biogenesis involves the complex assembly of molecular processes that are mostly governed by three proteins: RNase III protein DCL1 and two RNA binding proteins, SERRATE and HYL1. HYL1 protein is a double stranded RNA binding protein that is needed for the precise excision of miRNA/miRNA* duplex from the stem-loop containing primary miRNA gene transcripts. Moreover, HYL1 protein partners with HSP90 and CARP9 proteins to load the miRNA molecules onto the AGO1 endonuclease. HYL1 protein as a crucial player in the biogenesis pathway is regulated by its phosphorylation status to fine tune the levels of miRNA in various physiological conditions. HYL1 protein consists of two dsRNA binding domains (dsRBD) that are involved in RNA binding and dimerization and a C-terminal disordered tail of unknown function. Although the spatial structures of the individual dsRBDs have been determined there is a lack of information about the behaviour and structure of the full length protein. Using small the angle X-ray scattering (SAXS) technique we investigated the structure and dynamic of the HYL1 protein from Arabidopsis thaliana in solution. We show that the C-terminal domain is disordered and dynamic in solution and that HYL1 protein dimerization is dependent on the concentration. HYL1 protein lacking a C-terminal tail and a nuclear localisation signal (NLS) fragment is almost exclusively monomeric and similarly to full-length protein has a dynamic nature in solution. Our results point for the first time to the role of the C-terminal fragment in stabilisation of HYL1 dimer formation.