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  3. Fakultät für Biologie - Digitale Hochschulschriften der LMU - Teil 05/06 Podcast
  4. The role of ORC6 in the human cell
Orc6 is a crucial component of the replication initiation machinery
in eukaryotes. Its study helps us to better understand one of the
most basic cell functions: DNA replication. The first step in
replication initiation is the assembly and DNA binding of the
origin recognition complex (ORC). Although members of ORC are
highly conserved and well studied, Orc6 evolved faster than the
other members of the complex, has a different structure and is less
well understood than Orc1-5. The low number of studies on Orc6 in
human cells and the low predictability of results obtained from
model systems necessitates an in depth analysis of HsOrc6. The aim
of this study is to better understand the role of Orc6 in the human
cell by exploring the mechanism of Orc6-Orc1-5 interaction and
Orc6-DNA binding, studying the function of Orc6 in live cells, and
mapping its interaction network to find new pathways and functions
of the protein. The study concludes that Orc6 interacts via its C
terminus with Orc1-5 in vitro. It might also have a second pre-RC
interaction domain, probably close to the nuclear localization
signal. Orc6 is not required for Orc1-5-DNA binding, but is able to
enhance the process in vitro. It is also able to bind DNA in the
absence of Orc1-5 with high affinity. In vivo experiments present
evidence on the ability of Orc6 to recruit the replication
machinery to itself on DNA. Although this ability is less efficient
than that of EBNA1, a viral factor, it is sufficiently strong to
support autonomous replication of plasmids in human cell lines for
at least four weeks. Truncated variants of the protein containing
at least one of the predicted Orc1-5 interaction sites are
sufficient to allow replication. This finding can be used to design
autonomous-replicating plasmids for use in gene therapy. Mapping
the interaction network of Orc6 leads to the conclusion that HsOrc6
is only weakly attached to pre-RC factors, and is likely to have an
important role in mitosis and possibly be involved in other
pathways.

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