Structural Maintenance of Chromosomes Hinge Domain-containing protein 1 (SMCHD1) has been established as an epigenetic regulator, with critical roles in X-chromosome inactivation, autosomal gene silencing and genomic imprinting. Recently, variations in SMCHD1 have been associated with two human conditions: facioscapulohumeral muscular dystrophy (FSHD) and Bosma arhinia microphthalmia syndrome (BAMS). There has therefore been a growing interest in unveiling SMCHD1’s atomic structure and the molecular mechanisms underlying its function in both a healthy and diseased state.
To provide a better understanding of Smchd1’s molecular structure and function, I successfully expressed and purified the full-length 2007-amino acid Smchd1 protein. Electron microscopy analyses of the Smchd1 dimer revealed an elongated rod-like structure that displays a high conformational flexibility, similar to that of other structural maintenance of chromosomes (SMC) proteins. In follow-up studies of the two individual domains, we revealed the first atomic-resolution structure of Smchd1’s hinge domain, providing a novel insight into its DNA-binding and dimerisation modes. Contrary to previously suggested models describing the DNA interaction mode of canonical SMC proteins, I showed that nucleic acids are not threaded through the central pore region of the Smchd1 hinge domain. Subsequent immunofluorescence studies additionally revealed that the hinge domain targets full-length Smchd1 to chromatin, and that a functional hotspot within the hinge is required for chromatin localisation in cells. Using analytical ultracentrifugation, I demonstrated that the wild-type SMCHD1 ATPase undergoes dimerisation, which was reliant on the inclusion of both the UBL domain and the presence of substrate, ATP. Follow-up cellular studies revealed that Smchd1’s catalytic activity, as well as the presence of the newly- identified UBL domain, are both necessary for the localisation of full-length Smchd1 to chromatin. Together, these studies provide an insight into the molecular basis of Smchd1 function and highlight how chromatin binding may be compromised in human disease.