Emerging Microbes & Infections,
Год журнала:
2021,
Номер
10(1), С. 1626 - 1637
Опубликована: Янв. 1, 2021
Coronaviruses
(CoVs)
can
infect
a
variety
of
hosts,
including
humans,
livestock
and
companion
animals,
pose
serious
threat
to
human
health
the
economy.
The
current
COVID-19
pandemic,
which
is
caused
by
severe
acute
respiratory
syndrome
coronavirus
2
(SARS-CoV-2),
has
killed
millions
people.
Unfortunately,
effective
treatments
for
CoVs
infection
are
still
lacking,
suggesting
importance
vaccines.
Our
previous
work
showed
that
CoV
nonstuctural
protein
14
(nsp14)
functions
as
(guanine-N7)-methyltransferase
(N7-MTase),
involved
in
RNA
cap
formation.
Moreover,
we
found
N7-MTase
well
conserved
among
different
universal
target
developing
antivirals
against
CoVs.
Here,
show
be
an
ideal
designing
live
attenuated
Using
murine
hepatitis
virus
strain
A59
(MHV-A59),
representative
well-studied
model
coronaviruses,
constructed
N7-MTase-deficient
recombinant
MHV
D330A
Y414A.
These
two
mutants
highly
mice
exhibit
similar
replication
efficiency
wild-type
(WT)
cell
culture.
Furthermore,
single
dose
immunization
or
Y414A
induce
long-term
humoral
immune
responses
robust
CD4+
CD8+
T
responses,
provide
full
protection
challenge
lethal-dose
MHV-A59.
Collectively,
this
study
provides
strategy
design
vaccines
abolishing
viral
activity.
This
approach
may
apply
other
viruses
encode
their
own
conservative
N7-methyltransferase.
Nature Reviews Molecular Cell Biology,
Год журнала:
2021,
Номер
23(1), С. 21 - 39
Опубликована: Ноя. 25, 2021
Severe
acute
respiratory
syndrome
coronavirus
2
(SARS-CoV-2)
has
killed
millions
of
people
and
continues
to
cause
massive
global
upheaval.
Coronaviruses
are
positive-strand
RNA
viruses
with
an
unusually
large
genome
~30
kb.
They
express
RNA-dependent
polymerase
a
cohort
other
replication
enzymes
supporting
factors
transcribe
replicate
their
genomes.
The
proteins
performing
these
essential
processes
prime
antiviral
drug
targets,
but
discovery
is
hindered
by
our
incomplete
understanding
synthesis
processing.
In
infected
cells,
the
must
coordinate
viral
host
produce
both
mRNAs
new
Recent
research
aiming
decipher
contextualize
structures,
functions
interplay
subunits
SARS-CoV-2
transcription
complex
burgeoned.
this
Review,
we
discuss
recent
advancements
in
molecular
basis
complexity
RNA-synthesizing
machinery.
Specifically,
outline
mechanisms
regulation
translation,
transcription.
We
also
composition
complexes
suitability
as
targets
for
therapy.
coronaviruses
rely
on
specialized
studies
have
improved
transcription,
offer
therapeutic
targets.
Abstract
SARS-CoV-2
is
an
extremely
contagious
respiratory
virus
causing
adult
atypical
pneumonia
COVID-19
with
severe
acute
syndrome
(SARS).
has
a
single-stranded,
positive-sense
RNA
(+RNA)
genome
of
~
29.9
kb
and
exhibits
significant
genetic
shift
from
different
isolates.
After
entering
the
susceptible
cells
expressing
both
ACE2
TMPRSS2,
directly
functions
as
mRNA
to
translate
two
polyproteins
ORF1a
ORF1b
region,
which
are
cleaved
by
viral
proteases
into
sixteen
non-structural
proteins
(nsp1-16)
initiate
replication
transcription.
The
also
encodes
four
structural
(S,
E,
M
N)
up
six
accessory
(3a,
6,
7a,
7b,
8,
9b)
proteins,
but
their
translation
requires
newly
synthesized
individual
subgenomic
RNAs
(sgRNA)
in
infected
cells.
Synthesis
full-length
genomic
(gRNA)
sgRNAs
conducted
inside
double-membrane
vesicles
(DMVs)
transcription
complex
(RTC),
comprises
nsp7,
nsp8,
nsp9,
nsp12,
nsp13
short
primer.
To
produce
sgRNAs,
RTC
starts
synthesis
highly
structured
gRNA
3'
end
switches
template
at
various
regulatory
sequence
(TRS
B
)
sites
along
body
probably
mediated
long-distance
RNA–RNA
interaction.
TRS
motif
5'
leader
L
responsible
for
interaction
upstream
each
ORF
skipping
between
them
sgRNAs.
Abundance
depend
on
location
read-through
efficiency
.
Although
more
studies
needed,
unprecedented
pandemic
taught
world
painful
lesson
that
invest
proactively
prepare
future
emergence
other
types
coronaviruses
any
possible
biological
horrors.
High-throughput
sequencing
dramatically
changed
our
view
of
transcriptome
architectures
and
allowed
for
ground-breaking
discoveries
in
RNA
biology.
Recently,
full-length
transcripts
based
on
the
single-molecule
platform
from
Oxford
Nanopore
Technologies
(ONT)
was
introduced
is
widely
used
to
sequence
eukaryotic
viral
RNAs.
However,
experimental
approaches
implementing
this
technique
prokaryotic
transcriptomes
remain
scarce.
Here,
we
present
an
bioinformatic
workflow
ONT
RNA-seq
bacterial
model
organism
Escherichia
coli
,
which
can
be
applied
any
microorganism.
Our
study
highlights
critical
steps
library
preparation
computational
analysis
compares
results
gold
standards
field.
Furthermore,
comprehensively
evaluate
applicability
advantages
different
ONT-based
protocols,
including
direct
RNA,
cDNA,
PCR-cDNA.
We
find
that
(PCR)-cDNA-seq
offers
improved
yield
accuracy
compared
sequencing.
Notably,
suitable
quantitative
measurements
readily
simultaneous
accurate
detection
transcript
5′
3′
boundaries,
transcriptional
units,
heterogeneity.
In
summary,
comprehensive
study,
show
nanopore
a
ready-to-use
tool
allowing
rapid,
cost-effective,
annotation
multiple
transcriptomic
features.
Thereby
holds
potential
become
valuable
alternative
method
prokaryotes.
Viruses,
Год журнала:
2021,
Номер
13(10), С. 1923 - 1923
Опубликована: Сен. 24, 2021
SARS-CoV-2,
the
etiologic
agent
at
root
of
ongoing
COVID-19
pandemic,
harbors
a
large
RNA
genome
from
which
tiered
ensemble
subgenomic
RNAs
(sgRNAs)
is
generated.
Comprehensive
definition
and
investigation
these
products
are
important
for
understanding
SARS-CoV-2
pathogenesis.
This
review
summarizes
recent
progress
on
sgRNA
identification,
characterization,
application
as
viral
replication
marker.
The
significance
findings
potential
future
research
areas
interest
discussed.
Despite
an
unprecedented
global
research
effort
on
SARS-CoV-2,
early
replication
events
remain
poorly
understood.
Given
the
clinical
importance
of
emergent
viral
variants
with
increased
transmission,
there
is
urgent
need
to
understand
stages
and
transcription.
We
used
single-molecule
fluorescence
in
situ
hybridisation
(smFISH)
quantify
positive
sense
RNA
genomes
95%
detection
efficiency,
while
simultaneously
visualising
negative
genomes,
subgenomic
RNAs,
proteins.
Our
absolute
quantification
RNAs
factories
revealed
that
SARS-CoV-2
genomic
long-lived
after
entry,
suggesting
it
avoids
degradation
by
cellular
nucleases.
Moreover,
we
observed
highly
variable
between
cells,
only
a
small
cell
population
displaying
high
burden
RNA.
Unexpectedly,
B.1.1.7
variant,
first
identified
UK,
exhibits
significantly
slower
kinetics
than
Victoria
strain,
novel
mechanism
contributing
its
higher
transmissibility
important
implications.
Abstract
Since
the
development
of
Sanger
sequencing
in
1977,
technology
has
played
a
pivotal
role
molecular
biology
research
by
enabling
interpretation
biological
genetic
codes.
Today,
nanopore
is
one
leading
third‐generation
technologies.
With
its
long
reads,
portability,
and
low
cost,
widely
used
various
scientific
fields
including
epidemic
prevention
control,
disease
diagnosis,
animal
plant
breeding.
Despite
initial
concerns
about
high
error
rates,
continuous
innovation
platforms
algorithm
analysis
effectively
addressed
accuracy.
During
coronavirus
(COVID‐19)
pandemic,
critical
detecting
severe
acute
respiratory
syndrome
coronavirus‐2
virus
genome
containing
pandemic.
However,
lack
understanding
this
may
limit
popularization
application.
Nanopore
poised
to
become
mainstream
choice
for
preventing
controlling
COVID‐19
future
epidemics
while
creating
value
other
such
as
oncology
botany.
This
work
introduces
contributions
during
pandemic
promote
public
use
emerging
outbreaks
worldwide.
We
discuss
application
microbial
detection,
cancer
genomes,
genomes
summarize
strategies
improve
bioRxiv (Cold Spring Harbor Laboratory),
Год журнала:
2025,
Номер
unknown
Опубликована: Янв. 8, 2025
Abstract
RNA
viruses
like
SARS-CoV-2
have
a
high
mutation
rate,
which
contributes
to
their
rapid
evolution.
The
rate
of
mutations
depends
on
the
type
(e.g.,
A
→
C
,
G
etc.)
and
can
vary
between
sites
in
viral
genome.
Understanding
this
variation
shed
light
mutational
processes
at
play,
is
crucial
for
quantitative
modeling
Using
millions
available
full-genome
sequences,
we
estimate
rates
synonymous
all
12
possible
nucleotide
types
examine
how
much
these
sites.
We
find
surprisingly
level
variability
several
striking
patterns:
four
suddenly
increase
one
two
gene
boundaries;
most
strongly
depend
site’s
local
sequence
context,
with
up
56-fold
differences
contexts;
consistent
previous
study,
some
are
lower
engaged
secondary
structure.
simple
log-linear
model
features
explains
∼15-60%
fold-variation
sites,
depending
type;
more
complex
models
only
modestly
improve
predictive
power
out
sample.
fitness
effect
each
based
number
times
it
actually
occurs
versus
expected
occur
model.
identify
small
regions
genome
where
or
noncoding
less
often
than
expected,
indicative
strong
purifying
selection
that
independent
protein
sequence.
Overall,
work
expands
our
basic
understanding
SARS-CoV-2’s
evolution
by
characterizing
virus’s
process
individual
uncovering
patterns
arise
from
unknown
mechanisms.
Nature Communications,
Год журнала:
2021,
Номер
12(1)
Опубликована: Сен. 28, 2021
Abstract
The
dynamics
of
SARS-CoV-2
RNA
structure
and
their
functional
relevance
are
largely
unknown.
Here
we
develop
a
simplified
SPLASH
assay
comprehensively
map
the
in
vivo
RNA-RNA
interactome
genome
across
viral
life
cycle.
We
report
canonical
alternative
structures
including
5′-UTR
3′-UTR,
frameshifting
element
(FSE)
pseudoknot
cyclization
both
cells
virions.
provide
direct
evidence
interactions
between
Transcription
Regulating
Sequences,
which
facilitate
discontinuous
transcription.
In
addition,
reveal
short
long
distance
arches
around
FSE.
More
importantly,
find
that
within
virions,
while
undergoes
intensive
compaction,
domains
remain
stable
but
with
strengthened
demarcation
local
weakened
global
cyclization.
Taken
together,
our
analysis
reveals
structural
basis
for
regulation
replication,
transcription
translational
frameshifting,
conformations
maintenance
organization
during
whole
cycle
SARS-CoV-2,
anticipate
will
help
better
antiviral
strategies.