Different roles of the human Orc6 protein in the replication initiation process

Abstract

In eukaryotes, binding of the six-subunit origin recognition complex (ORC) to DNA provides an interactive platform for the sequential assembly of pre-replicative complexes. This process licenses replication origins competent for the subsequent initiation step. Here, we analyze the contribution of human Orc6, the smallest subunit of ORC, to DNA binding and pre-replicative complex formation. We show that Orc6 not only interacts with Orc1-Orc5 but also with the initiation factor Cdc6. Biochemical and imaging experiments reveal that this interaction is required for licensing DNA replication competent. Furthermore, we demonstrate that Orc6 contributes to the interaction of ORC with the chaperone protein HMGA1a (high mobility group protein A1a). Binding of human ORC to replication origins is not specified at the level of DNA sequence and the functional organization of origins is poorly understood. We have identified HMGA1a as one factor that might direct ORC to AT-rich heterochromatic regions. The systematic analysis of the interaction between ORC and HMGA1a revealed that Orc6 interacts with the acidic C-terminus of HMGA1a and also with its AT-hooks. Both domains support autonomous replication if targeted to DNA templates. As such, Orc6 functions at different stages of the replication initiation process. Orc6 can interact with ORC chaperone proteins such as HMGA1a to facilitate chromatin binding of ORC and is also an essential factor for pre-RC formation.

Chaperoning HMGA2 Protein Protects Stalled Replication Forks in Stem and Cancer Cells

“…It has been described that the highly homologous HMGA1a protein interacts with subunits of the human origin recognition complex (ORC) via its C-terminal domain (Thomae et al, 2008(Thomae et al, , 2011. Given the high degree of conservation, it is possible that HMGA2 also interacts with ORC and remains there after initiation of replication has occurred (Natsume and Tanaka, 2010).…”

Regulating DNA Replication in Eukarya

“…Despite significant homology between metazoan Orc6 proteins , human Orc6 interacts weakly with ORC, compared with its fly counterpart (Chesnokov et al 1999;Vashee et al 2003;Siddiqui and Stillman 2007). However, it is still required for DNA synthesis and may have evolved to perform other roles in origin recognition (Prasanth et al 2002;Balasov et al 2007;Thomae et al 2008Thomae et al , 2011.…”

Helicase loading: How to build a MCM2-7 double-hexamer

“…It remains unclear whether these structural features are required for helicase loading. Protein factors like HMGA or ORCA/Lrwd1, which bind to ORC, facilitate recruitment of ORC to DNA and in part can promote pre-RC assembly [32][33][34][35]. Interestingly, even tethering of ORC to non-origin DNA is sufficient to create an artificial origin of DNA replication [36].…”