Biomedical Engineering Reference
In-Depth Information
Rop dimer
RNA
I
ρ
ρ
ρ
RNAI
RNA
I
Origin
rop
5'
3'
3'
5'
ρ
RNA
II
RNA
II
Infrequently
Frequently
RNA
I
3'
5'
RNA
II
5'
5'
RNA
II
RNase H
RNA
II
Fig. 4.4
Regulation of replication of
Col E1-derived plasmids. RNA II must
be processed by RNase H before it can
prime replication. 'Origin' indicates
the transition point between the RNA
primer and DNA. Most of the time,
RNA I binds to RNA II and inhibits
the processing, thereby regulating the
copy number. ρ
RNAI
and ρ
RNA II
are
the promoters for RNA I and RNA II
transcription, respectively. RNA I is
coloured pale red and RNA II dark red.
The Rop protein dimer enhances the
initial pairing of RNA I and RNA II.
5'
3'
3'
5'
5'
RNA
I
-RNA
II
duplex
Pol
I
RNA
II
primes
DNA synthesis
RNA
II
inactivated for
primer function
Replication
No replication
ribonucleotide molecule called RNA II, which forms
an RNA-DNA hybrid at the replication origin
(Tomizawa & Itoh 1982), RNA II can only act as
a primer if it is cleaved by RNase H to leave a free
3
(Fig. 4.4). Since RNA I is encoded by the plasmid,
more of it will be synthesized when the copy number
of the plasmid is high. As the host cell grows and
divides, so the concentration of RNA I will fall and
the plasmid will begin to replicate again (Cesarini
et al.
1991, Eguchi
et al.
1991).
In addition to RNA I, a plasmid-encoded protein
called Rop helps maintain the copy number (Cesarini
et al.
1982). This protein, which forms a dimer,
enhances the pairing between RNA I and RNA II so
that processing of the primer can be inhibited even
at relatively low concentrations of RNA I. Deletion of
the
ROP
gene (Twigg & Sherratt 1980) or mutations
in RNA I (Muesing
et al.
1981) result in increased
copy numbers.
hydroxyl group. Unless RNA II is processed in this
way, it will not function as a primer and replication
will not ensue. Replication control is mediated by
another small (108-base) RNA molecule called RNA
I (Tomizawa & Itoh 1981), which is encoded by
the same region of DNA as RNA II but by the com-
plementary strand. Thus RNA I and RNA II are
complementary to each other and can hybridize to
form a double-stranded RNA helix. The formation of
this duplex interferes with the processing of RNA II
by RNase H and hence replication does not ensue
′
