Nature Genetics,
Год журнала:
2022,
Номер
54(12), С. 1919 - 1932
Опубликована: Дек. 1, 2022
It
remains
unclear
why
acute
depletion
of
CTCF
(CCCTC-binding
factor)
and
cohesin
only
marginally
affects
expression
most
genes
despite
substantially
perturbing
three-dimensional
(3D)
genome
folding
at
the
level
domains
structural
loops.
To
address
this
conundrum,
we
used
high-resolution
Micro-C
nascent
transcript
profiling
in
mouse
embryonic
stem
cells.
We
find
that
enhancer-promoter
(E-P)
interactions
are
largely
insensitive
to
(3-h)
CTCF,
or
WAPL.
YY1
has
been
proposed
as
a
regulator
E-P
loops,
but
also
had
minimal
effects
on
transcription
3D
folding.
Strikingly,
live-cell,
single-molecule
imaging
revealed
reduced
factor
(TF)
binding
chromatin.
Thus,
although
cohesin,
WAPL
is
not
required
for
short-term
maintenance
gene
expression,
our
results
suggest
may
facilitate
TFs
search
bind
their
targets
more
efficiently.
Nature Reviews Methods Primers,
Год журнала:
2021,
Номер
1(1)
Опубликована: Июнь 3, 2021
Single-molecule
localization
microscopy
(SMLM)
describes
a
family
of
powerful
imaging
techniques
that
dramatically
improve
spatial
resolution
over
standard,
diffraction-limited
and
can
image
biological
structures
at
the
molecular
scale.
In
SMLM,
individual
fluorescent
molecules
are
computationally
localized
from
sequences
localizations
used
to
generate
super-resolution
or
time
course
images,
define
trajectories.
this
Primer,
we
introduce
basic
principles
SMLM
before
describing
main
experimental
considerations
when
performing
including
labelling,
sample
preparation,
hardware
requirements
acquisition
in
fixed
live
cells.
We
then
explain
how
low-resolution
processed
reconstruct
images
and/or
extract
quantitative
information,
highlight
selection
discoveries
enabled
by
closely
related
methods.
discuss
some
limitations
potential
artefacts
as
well
ways
alleviate
them.
Finally,
present
an
outlook
on
advanced
promising
new
developments
fast-evolving
field
SMLM.
hope
Primer
will
be
useful
reference
for
both
newcomers
practitioners
