Genomics and Pharmacogenomics

DNA replication origins are licensed by the pre-replication complex, which encompasses six components of the origin recognition complex (ORC), CDT1, CDC6, and the MCM helicase. Mutations in pre-replication complex components have been shown to cause Meier-Gorlin Syndrome (MGS) a disorder characterized by microcephaly, short stature, and patellar abnormalities.  ORC components have also been shown to localize to heterochromatin and impact on the heterochromatin superstructure. Here, we show that deficiency in ORC1 in MGS patient cells impairs sister chromatid cohesion and formation of heterochromatin via a process we refer to as 'decompaction'. Comparison to other human cell lines with disordered chromatin, ICF (Immunodeficiency- Centromere instability-Facial anomalies syndrome) and RETT syndrome, revealed that the effect of Orc1 deficiency on heterochromatin structure is profound in human cell lines. Depletion of ORC1 by siRNA knockdown affects heterochromatin assembly and diminishes levels of HP1 and H3K9Me3.  Similar changes were observed in MGS patient cells with mutations in ORC1. We investigated the cohesion of sister chromatids using a fluorescent in-situ hybridization assay and observed an impairment in the cohesion of sister chromatids.  We examined aspects of the DNA damage response which are known to be sensitive to heterochromatin status to provide evidence for a functional impact of the disordered heterochromatin. ORC1 deficiency impairs DSB repair carried out by homologous recombination.  We suggest that these results can be explained by our 'decompaction' model. Where ORC1 deficiency leads to improperly structured heterochromatin.  These results suggest that ORC has a role in regulating heterochromatin structure and DSB repair by homologous recombination which is distinct to the role in origin licensing.