Main text
At first sight, RNA polymerase II has evolved to function in a rather
curious way with respect to genome integrity. Intuitively, we might
expect the holoenzyme once loaded to simply follow the helical turns of
the template strand, displacing histones and creating a single stranded
bubble in a very localised region but causing minimal disruption to
surrounding genome topology. This is not how RNA polymerase II operates:
it passages DNA through the active site, creating positive supercoils
ahead of the holoenzyme and negative supercoils behind that must be
either resolved by topoisomerases or allowed to relax when the
polymerase dissociates (Tsao et al., 1989). Quite how RNA polymerase
achieves this remains unclear – even the bacterial RNA polymerase II
working on a naked DNA can induce supercoiling (Janissen et al., 2023)
– and although this behaviour facilitates the release of the nascent
RNA (which would otherwise become wound around the DNA) it also creates
a set of vulnerabilities for mutation:
- Transcription requires the DNA helix to be transiently melted, and the
single-stranded DNA (ssDNA) formed is more sensitive to chemical
insults than dsDNA.
- Supercoiling around the polymerase must be relaxed by topoisomerases,
which can be mutagenic when topoisomerase action is not completed.
- The transcription regulatory machinery and RNA polymerases must be
removed to allow passage of replication forks.
- Negative supercoiling behind the polymerase enhances the formation of
non-B DNA structures, such as G-quadruplexes and RNA:DNA hybrids
called R-loops, which are considered to be a threat to genome
stability.
Many of these vulnerabilities arise from interactions between DNA
replication and direct or indirect outcomes of transcription. The
replisome moves extremely rapidly and must traverse transcriptional
units almost without pause to duplicate chromosomes sufficiently fast
not to become the rate limiting step of cell division. Here we review
the mechanisms by which transcription may induce DNA damage in the
budding yeast S. cerevisiae , focusing particularly on
interactions with DNA replication, and assess whether genome structure
has evolved to enhance genetic heterogeneity at particular times and
places by increasing vulnerability to transcription-induced genome
instability.