Rapid and quantitative functional interrogation of human enhancer variant activity in live mice
Nature Communications,
Год журнала:
2025,
Номер
16(1)
Опубликована: Янв. 6, 2025
Abstract
Functional
analysis
of
non-coding
variants
associated
with
congenital
disorders
remains
challenging
due
to
the
lack
efficient
in
vivo
models.
Here
we
introduce
dual-enSERT,
a
robust
Cas9-based
two-color
fluorescent
reporter
system
which
enables
rapid,
quantitative
comparison
enhancer
allele
activities
live
mice
less
than
two
weeks.
We
use
this
technology
examine
and
measure
gain-
loss-of-function
effects
previously
linked
limb
polydactyly,
autism
spectrum
disorder,
craniofacial
malformation.
By
combining
dual-enSERT
single-cell
transcriptomics,
characterise
gene
expression
cells
where
is
normally
ectopically
active,
revealing
candidate
pathways
that
may
lead
misregulation.
Finally,
demonstrate
widespread
utility
by
testing
fifteen
uncharacterised
rare
common
neurodevelopmental
disorders.
In
doing
so
identify
reproducibly
alter
activity
OTX2
MIR9-2
brain
enhancers,
implicating
them
autism.
Dual-enSERT
thus
allows
researchers
go
from
identifying
comparative
embryos
under
Язык: Английский
Massively parallel reporter assays and mouse transgenic assays provide complementary information about neuronal enhancer activity
bioRxiv (Cold Spring Harbor Laboratory),
Год журнала:
2024,
Номер
unknown
Опубликована: Апрель 23, 2024
Genetic
studies
find
hundreds
of
thousands
noncoding
variants
associated
with
psychiatric
disorders.
Massively
parallel
reporter
assays
(MPRAs)
and
Язык: Английский
Dissecting the biology of feeding and eating disorders
Trends in Molecular Medicine,
Год журнала:
2024,
Номер
30(4), С. 380 - 391
Опубликована: Март 1, 2024
Feeding
and
eating
disorders
(FEDs)
are
heterogenous
characterized
by
varying
patterns
of
dysregulated
weight.
Genome-wide
association
studies
(GWASs)
clarifying
their
underlying
biology
genetic
relationship
to
other
psychiatric
metabolic/anthropometric
traits.
Genetic
research
on
anorexia
nervosa
(AN)
has
identified
eight
significant
loci
uncovered
correlations
implicating
both
risk
factors.
Careful
explication
these
metabolic
contributors
may
be
key
developing
effective
enduring
treatments
for
devastating,
life-altering,
frequently
lethal
illnesses.
We
discuss
clinical
phenomenology,
genomics,
phenomics,
intestinal
microbiota,
functional
genomics
propose
a
path
that
translates
variants
genes,
genes
pathways,
pathways
outcomes
advance
the
science
eventually
treatment
FEDs.
Язык: Английский
An in vivo systemic massively parallel platform for deciphering animal tissue-specific regulatory function
Frontiers in Genetics,
Год журнала:
2025,
Номер
16
Опубликована: Апрель 9, 2025
Introduction:
Transcriptional
regulation
is
an
important
process
wherein
non-protein
coding
enhancer
sequences
play
a
key
role
in
determining
cell
type
identity
and
phenotypic
diversity.
In
neural
tissue,
these
gene
regulatory
processes
are
crucial
for
coordinating
plethora
of
interconnected
regionally
specialized
types,
ensuring
their
synchronized
activity
generating
behavior.
Recognizing
the
intricate
interplay
brain
imperative,
as
mounting
evidence
links
neurodevelopment
neurological
disorders
to
non-coding
genome
regions.
While
genome-wide
association
studies
swiftly
identifying
human
disease-associated
loci,
decoding
mechanisms
challenging
due
causal
variant
ambiguity
specific
tissue
impacts.
Methods:
Massively
parallel
reporter
assays
(MPRAs)
widely
used
culture
study
regions,
linking
sequence
differences
tissue-specific
function.
However,
widespread
use
animals
encounters
significant
challenges,
including
insufficient
viral
library
delivery
quantification,
irregular
transduction
rates,
injection
site
inflammation
disrupting
expression.
Here,
we
introduce
systemic
MPRA
(sysMPRA)
address
challenges
through
intravenous
AAV
delivery.
Results:
We
demonstrate
successful
into
diverse
mouse
tissues,
efficiently
specificity
candidate
enhancers
aligning
well
with
predictions
from
machine
learning
models.
highlight
that
sysMPRA
effectively
uncovers
effects
stemming
disruption
MEF2C
transcription
factor
binding
sites,
single-nucleotide
polymorphisms,
consequences
genetic
variations
associated
late-onset
Alzheimer‘s
disease.
Conclusion:
SysMPRA
effective
delivering
method
simultaneously
determines
transcriptional
functions
hundreds
vivo
across
multiple
tissues.
Язык: Английский
Just a SNP Away: The Future of in vivo Massively Parallel Reporter Assay
Cell Insight,
Год журнала:
2024,
Номер
4(1), С. 100214 - 100214
Опубликована: Окт. 10, 2024
The
human
genome
is
largely
noncoding,
yet
the
field
still
grasping
to
understand
how
noncoding
variants
impact
transcription
and
contribute
disease
etiology.
massively
parallel
reporter
assay
(MPRA)
has
been
employed
characterize
function
of
at
unprecedented
scales,
but
its
application
limited
by
Язык: Английский
Novelty versus innovation of gene regulatory elements in human evolution and disease
Current Opinion in Genetics & Development,
Год журнала:
2024,
Номер
90, С. 102279 - 102279
Опубликована: Ноя. 26, 2024
Язык: Английский
How our brains are built: emerging approaches to understand human-specific features
Current Opinion in Genetics & Development,
Год журнала:
2024,
Номер
89, С. 102278 - 102278
Опубликована: Ноя. 15, 2024
Язык: Английский
Rapid and Quantitative Functional Interrogation of Human Enhancer Variant Activity in Live Mice
bioRxiv (Cold Spring Harbor Laboratory),
Год журнала:
2023,
Номер
unknown
Опубликована: Дек. 10, 2023
Abstract
Functional
analysis
of
non-coding
variants
associated
with
human
congenital
disorders
remains
challenging
due
to
the
lack
efficient
in
vivo
models.
Here
we
introduce
dual-enSERT,
a
robust
Cas9-based
two-color
fluorescent
reporter
system
which
enables
rapid,
quantitative
comparison
enhancer
allele
activities
live
mice
any
genetic
background.
We
use
this
new
technology
examine
and
measure
gain-
loss-of-function
effects
linked
limb
polydactyly,
autism,
craniofacial
malformation.
By
combining
dual-enSERT
single-cell
transcriptomics,
characterize
variant
alleles
at
cellular
resolution,
thereby
implicating
candidate
molecular
pathways
pathogenic
misregulation.
further
show
that
independent,
polydactyly-linked
lead
ectopic
expression
same
cell
populations,
indicating
shared
mechanisms
underlying
pathogenesis.
Finally,
streamline
for
F0
animals
by
placing
both
reporters
on
transgene
separated
synthetic
insulator.
Dual-enSERT
allows
researchers
go
from
identifying
comparative
activity
embryos
under
two
weeks.
Язык: Английский
Advances in computational and experimental approaches for deciphering transcriptional regulatory networks
BioEssays,
Год журнала:
2024,
Номер
46(7)
Опубликована: Май 8, 2024
Understanding
the
influence
of
cis-regulatory
elements
on
gene
regulation
poses
numerous
challenges
given
complexities
stemming
from
variations
in
transcription
factor
(TF)
binding,
chromatin
accessibility,
structural
constraints,
and
cell-type
differences.
This
review
discusses
role
regulatory
networks
enhancing
understanding
transcriptional
covers
construction
methods
ranging
expression-based
approaches
to
supervised
machine
learning.
Additionally,
key
experimental
methods,
including
MPRAs
CRISPR-Cas9-based
screening,
which
have
significantly
contributed
TF
binding
preferences
element
functions,
are
explored.
Lastly,
potential
learning
artificial
intelligence
unravel
logic
is
analyzed.
These
computational
advances
far-reaching
implications
for
precision
medicine,
therapeutic
target
discovery,
study
genetic
health
disease.
Язык: Английский