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In rho-dependent termination in prokaryotes, how does RNA polymerase “know” that it has reached the end of a gene and that it has to stop so that the rho-factor can bind mRNA’s rut site? Is there a termination sequence on the template DNA that “signals” the end of a gene?

And consequently, how does the rho-factor “know” when to bind to mRNA?... how does rho-factor recognize that the RNA polymerase has stopped so it can start its ATPase activity to dissociate the transcription complex from DNA and release the mRNA?

P...
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Termination of the synthesis of the RNA molecule in bacteria is also signaled by a sequence in the template strand of the DNA molecule.
This sequence of nucleotide in the template strand is the RNA polymerase pause site. When the RNAP continues its transcription it reaches this site and pauses it's transcription activity over here. The longevity of this pause can also be affected by various factors like high $Mg^{2+}$ ions in some cases.

Meanwhile, rho factor which after binding to rut-site, continues it's translocation along the mRNA (supposing mRNA is synthesized here) in 5'-3' direction using its ATPase activity. The rho factor utilises this opportunity when RNAP has paused it's transcription activity at the RNAP pause site to reach this ternary transcription elongation complex. When it had reached this complex it starts it's ATP dependent helicase activity which separates the RNA-DNA hybrid helix, causing the release of the mRNA.

Sikander
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In prokaryotes you have transcription and translation happen almost simultaneously. Downstream of the translational stop codon, there exists a C rich site on the mRNA called the rho-utilisation-site (rut) and another site called transcription-stop-point (tsp).

Rho-dependent Termination of Transcription Is Governed Primarily by the Upstream Rho Utilization (rut) Sequences of a Terminator

How does RNA polymerase know it has reached the end? RNA polymerase pauses at the tsp site during transcription. The rut site functions as an mRNA loading site and it also works to activate the rho-factor. The activated rho-factor travels down the mRNA while still attached to the rut site and catches up with the paused RNA polymerase. Contact with the RNA polymerase leads to RNA polymerase to dissociate from the transcriptional complex You can read about the dissociation here, the article is behind a paywall though.

I think I answered the "when does rho know?" too. Read the wikipedia article on this as well

prab4th
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