An allosteric mechanism of Rho-dependent

transcription termination
Rho is the essential RNA helicase that sets the borders between transcription units
and adjusts transcriptional yield to translational needs in bacteria 1, 2, 3. Although Rho
was the first termination factor to be discovered 4, the actual mechanism by which it
reaches and disrupts the elongation complex (EC) is unknown. Here we show that
the termination-committed Rho molecule associates with RNA polymerase (RNAP)
throughout the transcription cycle; that is, it does not require the nascent transcript
for initial binding. Moreover, the formation of the RNAP–Rho complex is crucial for
termination. We show further that Rho-dependent termination is a two-step process
that involves rapid EC inactivation (trap) and a relatively slow dissociation.
Inactivation is the critical rate-limiting step that establishes the position of the
termination site. The trap mechanism depends on the allosterically induced
rearrangement of the RNAP catalytic centre by means of the evolutionarily
conserved mobile trigger-loop domain, which is also required for EC dissociation.
The key structural and functional similarities, which we found between Rho-
dependent and intrinsic (Rho-independent) termination pathways, argue that the
allosteric mechanism of termination is general and likely to be preserved for all
cellular RNAPs throughout evolution. (i) RNA polymerase (RNA Pol) binds to
template DNA and begins synthesizing RNA. The nascent RNA forms a duplex with
the template DNA, stabilizing RNA Pol on the template DNA. (ii) RNA Pol reaches a
Rho-utilization (rut) site and transcribes this sequence. (iii) Rho recognizes this
sequence and binds to it. This binding does not occur if a ribosome is bound to the
rut site and is translating it. (iv) Using energy from ATP hydrolysis, Rho moves along
the RNA transcript towards RNA Pol. (v) While RNA Pol pauses at a Rho-termination
site in the DNA, Rho catches up to RNA Pol and uses its RNA-DNA helicase activity to
cause RNA Pol to dissociate. These events result in the termination of transcription.

Termination Sites

There are a number of proteins involved in termination of transcription,

particularly a protein called rho. However the mechanism by which rho
terminates transcription is not understood, and there are as yet no
apparent sequence similarities in these rho-dependent termination
regions.
Figure 1: A rho-independent method of transcription termination. The direction of transcription
is from left to right, the RNA polymerase encounters a G-C rich inverted repeat which forms a
hairpin loop, causeing the RNA polymerase to pause. The DNA reanneals, knocking off the
RNA polymerase and the newly synthesised chain.

There is a rho-independent method of termination which is understood, (depicted in figure 1,

page ) in which the RNA polymerase reaches a GC rich region with an inverted repeat -
such as CCCACT AGTGGG . When such a feature is encountered, the newly-synthesised
RNA forms a hairpin loop and it is believed that this may cause the polymerase to pause,
allowing the DNA to reanneal locally and breaking off the synthesised RNA chain. This is
associated in particular with a sequence of U's in the RNA, the A-U bonds being weaker than G-
C (2, as opposed to 3 hydrogen bonds), making it more likely for the RNA to break off.