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Insight into the Complexity of the i‑Motif and G‑Quadruplex DNA Structures Formed in the KRAS Promoter and Subsequent Drug-Induced Gene Repression
journal contribution
posted on 2017-06-01, 00:00 authored by Christine
E. Kaiser, Natalie A. Van Ert, Prashansa Agrawal, Reena Chawla, Danzhou Yang, Laurence H. HurleyActivating KRAS mutations
frequently occur in
pancreatic, colorectal, and lung adenocarcinomas. While many attempts
have been made to target oncogenic KRAS, no clinically useful therapies
currently exist. Most efforts to target KRAS have focused on inhibiting
the mutant protein; a less explored approach involves targeting KRAS
at the transcriptional level. The promoter element of the KRAS gene contains a GC-rich nuclease hypersensitive site
with three potential DNA secondary structure-forming regions. These
are referred to as the Near-, Mid-, and Far-regions, on the basis
of their proximity to the transcription start site. As a result of
transcription-induced negative superhelicity, these regions can open
up to form unique DNA secondary structures: G-quadruplexes on the
G-rich strand and i-motifs on the C-rich strand. While the G-quadruplexes
have been well characterized, the i-motifs have not been investigated
as thoroughly. Here we show that the i-motif that forms in the C-rich
Mid-region is the most stable and exists in a dynamic equilibrium
with a hybrid i-motif/hairpin species and an unfolded hairpin species.
The transcription factor heterogeneous nuclear ribonucleoprotein K
(hnRNP K) was found to bind selectively to the i-motif species and
to positively modulate KRAS transcription. Additionally,
we identified a benzophenanthridine alkaloid that dissipates the hairpin
species and destabilizes the interaction of hnRNP K with the Mid-region
i-motif. This same compound stabilizes the three existing KRAS G-quadruplexes. The combined effect of the compound
on the Mid-region i-motif and the G-quadruplexes leads to downregulation
of KRAS gene expression. This dual i-motif/G-quadruplex-interactive
compound presents a new mechanism to modulate gene expression.