Recent studies have
proven that the genetic landscape of pancreatic
cancer is dominated by the
KRAS
oncogene. Its transcription
is controlled by a G-rich motif (called 32R) located immediately upstream
of the TSS. 32R may fold into a G-quadruplex (G4) in equilibrium between
two G4 conformers: G9T (
T
M
= 61.2 °C)
and G25T (
T
M
= 54.7 °C). We found
that both G4s bind to hnRNPA1 and its proteolytic fragment UP1, promoting
several contacts with the RRM protein domains. 1D NMR analysis of
DNA imino protons shows that, upon binding to UP1, G25T is readily
unfolded at both 5′ and 3′ tetrads, while G9T is only
partially unfolded. The impact of hnRNPA1 on
KRAS
expression was determined by comparing Panc-1 cells with two Panc-1
knockout cell lines in which hnRNPA1 was deleted by the CRISPR/Cas9
technology. The results showed that the expression of
KRAS
is inhibited in the knockout cell lines, indicating that hnRNPA1
is essential for the transcription of
KRAS
. In addition,
the knockout cell lines, compared to normal Panc-1 cells, show a dramatic
decrease in cell growth and capacity of colony formation. Pull-down
and Western blot experiments indicate that conformer G25T is a better
platform than conformer G9T for the assembly of the transcription
preinitiation complex with PARP1, Ku70, MAZ, and hnRNPA1. Together,
our data prove that hnRNPA1, being a key transcription factor for
the activation of
KRAS
, can be a new therapeutic
target for the rational design of anticancer strategies.
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