The Origin Recognition Complex: A Biochemical and Structural View

Abstract

The origin recognition complex (ORC) was first discovered in the baker’s yeast in 1992. Identification of ORC opened up a path for subsequent molecular level investigations on how eukaryotic cells initiate and control genome duplication each cell cycle. Twenty years after the first biochemical isolation, ORC is now taking on a three-dimensional shape, although a very blurry shape at the moment, thanks to the recent electron microscopy and image reconstruction efforts. In this chapter, we outline the current biochemical knowledge about ORC from several eukaryotic systems, with emphasis on the most recent structural and biochemical studies. Despite many species-specific properties, an emerging consensus is that ORC is a ATP-dependent machine that recruits other key proteins to form pre-Replicative Complexes (pre-RCs) at many origins of DNA replication, enabling the subsequent initiation of DNA replication in S phase.

The Dynamics of Eukaryotic Replication Initiation: Origin Specificity, Licensing, and Firing at the Single-Molecule Level

“…ORC binding to ARS1 was confirmed by comparison to the binding distribution on a λ substrate lacking the ARS1 site (Figure 2B). ORC’s preference for AT-rich sequences agrees with expectations based on the conserved property of eukaryotic ORCs for binding AT-rich DNA (Li and Stillman, 2012) and the high AT content of ARS sequences (Siow et al, 2012). Furthermore, unlabeled ORC can load Mcm2-7 at non- ARS1 sites (albeit with low efficiency), indicating that non- ARS1 -bound ORC is at least partially biochemically functional (see below).…”

RPA Interacts with HIRA and Regulates H3.3 Deposition at Gene Regulatory Elements in Mammalian Cells

“…Importantly, depletion of RPA1 also led to reduced H3.3 deposition in the G1 phase cells (Figure S2A–D). In addition, we observed that depletion of ORC3, a subunit of the Origin Recognition Complex involved in DNA initiation of replication (Li and Stillman, 2012), while also leading to defects in cell cycle progression and spontaneous DNA damage (Figure S2E–G), had no apparent effect on H3.3 deposition (Figure S2H–I). Thus, it is unlikely that the effect of RPA1 depletion on H3.3 deposition is the secondary effect arising from defects in DNA replication, DNA repair or cell cycle progression in RPA1 depleted cells.…”

ORC4 Surrounds Extruded Chromatin in Female Meiosis

“…However, recent models for ORC suggest it might be more transiently associated with DNA than previously thought [Li and Stillman, 2012], so it is possible that we missed time points in which ORC subunits were associated with DNA. Our own data for ORC4 (discussed below) and ORC2 (discussed in ref.…”

“…from bacteria, in which DnaA is structurally and functionally homologous to ORC to mammals and are part of a group of proteins termed replication initiators [Scholefield et al, 2011]. The six ORC proteins were originally discovered in yeast and named according to size, with ORC1 being the largest, and the subunits in other species names according to the homology with their yeast counterparts [Li and Stillman, 2012]. More studies have been done on human ORC than mouse, but the close homology between these two species suggests that similar mechanisms are involved in both [Kneissl et al, 2003; Springer et al, 1999a; Springer et al, 1999b].…”