Minor spliceosome and disease

Abstract

The U12-type dependent (minor) spliceosome excises a rare group of introns that are characterized by a highly conserved 5' splice site and branch point sequence. Several new congenital or somatic diseases have recently been associated with mutations in components of the minor spliceosome. A common theme in these diseases is the detection of elevated levels of transcripts containing U12-type introns, of which a subset is associated with other splicing defects. Here we review the present understanding of the minor spliceosome diseases, particularly those associated with the specific components of the minor spliceosome. We also present a model for interpreting the molecular-level consequences of the different diseases.

Full text

Please
cite
this
article
in
press
as:
B.
Verma,
et
al.,
Minor
spliceosome
and
disease,
Semin
Cell
Dev
Biol
(2017),
https://doi.org/10.1016/j.semcdb.2017.09.036
ARTICLE IN PRESS
G Model
YSCDB-2404;
No.
of
Pages
10
Seminars
in
Cell
&
Developmental
Biology
xxx
(2017)
xxx–xxx
Contents
lists
available
at
ScienceDirect
Seminars
in
Cell
&
Developmental
Biology
j
ourna
l
h
o
me
page:
www.elsevier.com/locate/semcdb
Review
Minor
spliceosome
and
disease
Bhupendra
Verma1,
Maureen
V.
Akinyi,
Antto
J.
Norppa,
Mikko
J.
Frilander∗
Institute
of
Biotechnology,
PL56
(Viikinkaari
5),
000014,
University
of
Helsinki,
Finland
a
r
t
i
c
l
e
i
n
f
o
Article
history:
Received
4
July
2017
Received
in
revised
form
21
September
2017
Accepted
27
September
2017
Available
online
xxx
Keywords:
U12-type
introns
Minor
spliceosome
Human
diseases
Pre-mRNA
splicing
Cryptic
splice
sites
Exon
skipping
Intron
retention
a
b
s
t
r
a
c
t
The
U12-dependent
(minor)
spliceosome
excises
a
rare
group
of
introns
that
are
characterized
by
a
highly
conserved
5splice
site
and
branch
point
sequence.
Several
new
congenital
or
somatic
diseases
have
recently
been
associated
with
mutations
in
components
of
the
minor
spliceosome.
A
common
theme
in
these
diseases
is
the
detection
of
elevated
levels
of
transcripts
containing
U12-type
introns,
of
which
a
subset
is
associated
with
other
splicing
defects.
Here
we
review
the
present
understanding
of
minor
spliceosome
diseases,
particularly
those
associated
with
the
specific
components
of
the
minor
spliceosome.
We
also
present
a
model
for
interpreting
the
molecular-level
consequences
of
the
different
diseases.
©
2017
The
Authors.
Published
by
Elsevier
Ltd.
This
is
an
open
access
article
under
the
CC
BY
license
(http://creativecommons.org/licenses/by/4.0/).
Contents
1.
Introduction
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2.
Minor
spliceosome
in
human
disease
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2.1.
Diseases
affecting
the
intron
recognition
step
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2.1.1.
Isolated
growth
hormone
deficiency
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2.1.2.
Cerebellar
ataxia
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2.1.3.
Myelodysplastic
syndrome
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2.2.
Diseases
affecting
the
formation
of
catalytically
active
spliceosome
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2.2.1.
MOPD1/TALS.
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2.2.2.
Roifman
syndrome
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2.3.
Diseases
with
mutations
in
the
U12-type
splice
sites
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2.4.
Other
spliceosome
mutations
affecting
minor
spliceosome
activity
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3.
Minor
spliceosome
components
as
biomarkers
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4.
Consequences
of
minor
spliceosome-specific
mutations
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4.1.
The
fate
of
the
mRNA:
intron
retention
vs
cryptic
splicing
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4.2.
Fate
of
the
mRNA
and
disease
severity
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4.3.
Defects
with
individual
genes
vs
a
global
splicing
defect
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5.
Future
perspective
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Acknowledgements.
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References
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00
∗Corresponding
author.
E-mail
address:
Mikko.Frilander@Helsinki.Fi
(M.J.
Frilander).
1Present
address:
Department
of
Biotechnology,
All
India
Institute
of
Medical
Sciences,
New
Delhi,
India.
https://doi.org/10.1016/j.semcdb.2017.09.036
1084-9521/©
2017
The
Authors.
Published
by
Elsevier
Ltd.
This
is
an
open
access
article
under
the
CC
BY
license
(http://creativecommons.org/licenses/by/4.0/).

Please
cite
this
article
in
press
as:
B.
Verma,
et
al.,
Minor
spliceosome
and
disease,
Semin
Cell
Dev
Biol
(2017),
https://doi.org/10.1016/j.semcdb.2017.09.036
ARTICLE IN PRESS
G Model
YSCDB-2404;
No.
of
Pages
10
2
B.
Verma
et
al.
/
Seminars
in
Cell
&
Developmental
Biology
xxx
(2017)
xxx–xxx
1.
Introduction
Pre-mRNA
splicing
is
an
essential
step
in
the
gene
expression
pathway
of
all
eukaryotes.
During
splicing,
the
non-coding
intron
sequences
are
recognized
and
removed
from
precursor
mRNA
(pre-
mRNA)
by
the
spliceosome,
a
large
ribonucleoprotein
complex.
Defects
in
splicing
of
one
or
more
mRNA
species
are
a
major
cause
of
human
diseases
and
present
estimates
suggest
that
up
to
60%,
are
associated
with
pre-mRNA
splicing
[1,2].
In
addition
to
mono-
genic
disorders,
genetic
variants
altering
splicing
are
also
thought
to
be
an
important
contributor
to
complex
diseases
and
cancer
[3,4].
The
majority
of
disease-associated
splicing
defects
are
cis-
acting
mutations
within
a
single
gene
that
disrupt
either
the
splice
site
consensus
sequences
or
splicing
regulatory
elements
located
in
introns
or
exons
[5].
Notably,
exonic
mutations
interpreted
as
mis-
sense,
nonsense
or
silent
also
commonly
affect
splicing
[4,6,7].
The
outcome
of
cis-acting
splicing
mutations
is
often
either
a
formation
of
abnormal
or
non-functional
protein
species
or
accelerated
decay
of
the
affected
individual
mRNAs.
In
contrast,
mutations
in
splicing
factors,
spliceosome
components
or
spliceosome
assembly
factors
often
lead
to
widespread
defects
in
the
processing
of
large
numbers
of
pre-mRNAs
[8–10].
Most
metazoan
species
contain
two
distinct
pre-mRNA
splicing
machineries
known
as
the
major
(U2-dependent)
and
minor
(U12-
dependent)
spliceosomes,
which
recognize
and
excise
either
the
major
(U2-type)
or
minor
(U12-type)
class
of
introns,
respectively.
In
contrast
to
the
major
introns,
U12-type
introns
are
character-
ized
by
divergent
and
highly
conserved
5splice
site
(5ss)
and
branch
point
sequences
(BPS)
(Fig.
1A;
[11]).
These
introns
also
lack
the
characteristic
polypyrimidine
tract
(PPT)
that
is
present
in
U2-type
introns
immediately
upstream
of
the
3splice
site
(3ss).
Minor
introns
constitute
only
∼0.35%
of
all
human
introns
and
have
been
reported
to
be
present
in
700–800
genes,
each
of
which
typ-
ically
carry
only
a
single
U12-type
intron
and
multiple
U2-type
introns.
U12-type
introns
are
enriched
in
genes
that
represent
a
rather
restricted
set
of
functional
classes
and
pathways.
Particu-
larly
they
are
present
in
genes
related
to
‘information
processing
functions’,
such
as
DNA
replication
and
repair,
transcription,
RNA
processing,
and
translation,
but
can
also
be
found
in
genes
related
to
cytoskeletal
organization,
vesicular
transport,
and
voltage-gated
ion
channel
activity,
as
suggested
originally
by
Burge
et
al.,
[12]
and
verified
later
[13,14].
Both
the
identities
of
genes
carrying
U12-
type
introns
and
their
positions
within
the
genes
are
evolutionarily
conserved
[15].
The
overall
organisational
and
functional
features
of
both
spliceosomes
are
highly
similar.
Both
are
composed
of
five
small
nuclear
RNA
(snRNA)
molecules
that
associate
with
protein
factors
to
give
rise
to
small
nuclear
ribonucleoproteins
(snRNPs).
Within
the
minor
spliceosome,
four
of
the
five
snRNAs
are
unique.
Specifi-
cally,
U11,
U12,
U4atac,
and
U6atac
replace
the
major
spliceosome
counterparts
U1,
U2,
U4
and
U6
snRNAs,
respectively.
U5
snRNA
is
shared
between
the
two
spliceosomes.
Of
the
200–300
pro-
teins
associated
with
spliceosomes,
most
are
thought
to
be
shared
between
the
two
systems
and
only
7
proteins,
associated
with
U11
and
U12
snRNPs,
are
unique
to
the
minor
spliceosome
[16,17].
The
highly
similar
spliceosome
composition
is
reflected
in
the
conserved
assembly
pathway
and
catalytic
mechanism.
Both
spliceosomes
are
assembled
sequentially
starting
from
intron
recognition,
followed
by
formation
of
a
catalytically
active
spliceo-
some
and
joining
of
the
exons
flanking
the
excised
intron.
With
minor
introns,
the
5ss
and
BPS
are
co-operatively
recognized
by
a
pre-formed
U11/U12
di-snRNP
[18],
contrary
to
the
sequential
recognition
of
these
sequences
by
individual
U1
and
U2
snRNPs
in
major
introns.
Since
the
PPT
is
lacking
in
minor
introns,
the
U2AF1/2
heterodimer
that
recognizes
the
PPT
and
3ss
of
major
introns
does
not
associate
with
minor
introns.
Instead,
an
integral
U11/U12
di-snRNP
protein
component,
Urp/ZRSR2
takes
up
the
role
of
3ss
recognition
with
minor
introns
[19].
Following
this
initial
recognition,
both
splicing
pathways
proceed
with
association
of
a
tri-snRNP,
either
U4atac/U6atac.U5
or
U4/U6.U5
(Fig.
1B;
[20,21]).
Further
rearrangements
in
RNA–RNA
and
RNA–protein
interac-
tions
lead
to
the
formation
of
a
catalytically
active
spliceosome
and
catalytic
excision
of
the
intron
[20,22,23].
Here,
we
review
the
role
of
the
minor
spliceosome
in
human
diseases,
with
a
specific
focus
on
diseases
caused
by
mutations
in
the
integral
components
of
the
minor
spliceosome.
Given
that
most
protein
components
and
U5
snRNA
are
shared
with
the
major
spliceosome,
there
are
also
several
diseases
where
mutations
dis-
rupting
shared
components
can
potentially
affect
the
functions
of
both
spliceosomes.
Those
have
been
discussed
in
detail
elsewhere
[24].
2.
Minor
spliceosome
in
human
disease
The
direct
targets
for
human
diseases
specific
for
the
minor
spliceosome
are
the
unique
snRNA
and
protein
components.
This
includes
several
components
of
the
U11/U12
intron
recognition
complex
and
the
U4atac
and
U6atac
snRNAs
in
the
minor
tri-snRNP.
The
U11/U12
di-snRNP
contains,
in
addition
to
the
U11
and
U12
snRNAs,
seven
integral
proteins
(65K,
48K,
59K,
35K,
31K,
25K
and
20K)
that
are
unique
to
the
minor
spliceosome
[25,26].
Addition-
ally,
the
Urp/ZRSR2
protein
associated
with
the
U11/U12
di-snRNP
has
been
reported
to
function
in
both
minor
and
major
spliceo-
somes,
with
an
essential
role
for
U12-type
intron
3ss
recognition
[19].
To
date
five
human
diseases
with
mutations
in
the
spe-
cific
components
of
the
minor
spliceosome
have
been
described
(Table
1).
Three
of
them
affect
the
components
of
the
U11/U12
di-
snRNP,
namely
the
U11/U12-65K
protein
(RNPC3;
[27]),
U12
snRNA
(RNU12;
[28])
and
Urp
protein
(ZRSR2;
[29]);
while
two
diseases
are
attributed
to
mutations
in
the
U4atac
snRNA
(RNU4ATAC;
[30–32]).
Each
of
these
diseases
is
hypomorphic,
leading
only
to
a
partial
loss
of
minor
spliceosome
function
because
correctly
spliced
mRNAs
can
be
detected
in
the
patient
cells.
We
briefly
introduce
each
disease
and
later
discuss
the
impact
of
disease
mutations
on
the
assembly
of
the
minor
spliceosome
and
the
fate
of
affected
mRNAs.
Apart
from
mutations
in
the
core
minor
spliceosome
compo-
nents,
we
also
describe
the
few
reported
human
diseases
caused
by
mutations
at
the
splice
sites
of
U12-type
introns,
though
we
note
that
this
appears
to
be
underexplored
territory
given
the
small
number
of
cases
reported
so
far.
Finally,
we
mention
the
emerging
role
for
the
minor
spliceosome
in
cancer
and
autoimmune
disorders
as
well
as
neurodegenerative
diseases.
2.1.
Diseases
affecting
the
intron
recognition
step
2.1.1.
Isolated
growth
hormone
deficiency
Recessive
mutations
in
the
RNPC3
gene,
encoding
U11/U12-
65K,
one
of
the
seven
minor
spliceosome-specific
proteins,
have
been
associated
with
isolated
growth
hormone
deficiency
(IGHD)
and
associated
pituitary
hypoplasia.
The
65K
protein
is
part
of
a
molecular
bridge
that
connects
the
U11
and
U12
snRNPs
into
a
di-
snRNP
(Fig
1B;
[33]).
Initially,
RNPC3
mutations
were
detected
in
a
single
family
only
[27],
but
additional
cases
with
overlapping
muta-
tions
and
similar
phenotypes
have
been
described
subsequently
[34].
IGHD
is
a
condition
characterized
by
a
shortage
or
absence
of
growth
hormone,
with
the
absence
of
associated
pituitary
hor-
mone
deficiencies.
Genetically,
IGHD
is
a
diverse
disease
and
can
result
from
either
recessive
or
dominant
mutations
in
various
genes
involved
in
pituitary
development
or
function
[35].

Please
cite
this
article
in
press
as:
B.
Verma,
et
al.,
Minor
spliceosome
and
disease,
Semin
Cell
Dev
Biol
(2017),
https://doi.org/10.1016/j.semcdb.2017.09.036
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of
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B.
Verma
et
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/
Seminars
in
Cell
&
Developmental
Biology
xxx
(2017)
xxx–xxx
3
Fig.
1.
(A)
Consensus
sequences
of
human
U12-type
and
U-2
type
introns.
Adapted
from
[11].
(B)
A
simplified
assembly
pathway
of
the
U12-dependent
spliceosome
highlighting
the
intron
recognition
and
catalytic
stages
of
the
assembly.
The
schematic
structure
of
the
U11/U12
intron
recognition
complex
includes
protein
components
specific
to
the
U12-dependent
spliceosome
(65K
(RNPC3),
59K
(PDCD7),
48K
(SNRP48),
35K
(SNRNP35),
31K
(SNRNP31/ZCRB1),
25K
(SNRP25K),
20K
(SNRNP20/ZMAT5),
Urp
(ZRSR2)
and
also
the
SF3b
complex
shared
between
the
two
spliceosomes.
Additionally,
human
diseases
specifically
associated
with
either
assembly
step
of
the
minor
spliceosome
assembly
are
indicated
on
right.
All
reported
IGHD
cases
associated
with
RNPC3
mutations
are
compound
heterozygous
with
a
missense
P474T
mutation
com-
bined
with
either
R502X
or
R205X
nonsense
mutation
[27,34].
At
the
protein
level
the
P474T
and
R502X
mutations
map
to
the
C-
terminal
RNA
recognition
motif
(RRM)
of
the
protein,
while
the
R205X
maps
to
the
proline-rich
region
between
the
two
RRMs
(Fig.
2A).
Although
the
tissue
affected
in
IGHD,
the
pituitary
gland,
cannot
be
sampled
from
the
patients,
RT-PCR
and
RNA-seq
analyses

Please
cite
this
article
in
press
as:
B.
Verma,
et
al.,
Minor
spliceosome
and
disease,
Semin
Cell
Dev
Biol
(2017),
https://doi.org/10.1016/j.semcdb.2017.09.036
ARTICLE IN PRESS
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Biology
xxx
(2017)
xxx–xxx
Table
1
Human
diseases
showing
defects
in
splicing
of
U12-type
introns.
Defect
in
Locus
Disease
Abbreviation
References
Integral
components
of
minor
snRNPs
RNPC3
Isolated
Growth
Hormone
Deficiency
IGHD
[27]
ZRSR2
Myelodysplastic
Syndrome MDS
[29]
RNU12
Early-onset
Cerebellar
ataxia
EOCA
[28]
RNU4ATAC
Microcephalic
osteodysplastic
primordial
dwarfism/Taybi-Linder
Syndrome
MOPD1/TALS
[31,32]
RNU4ATAC
Roifman
syndrome
RFMN
[30]
U12-type
5ss STK11
Peutz-Jegher’s
syndrome
PJS
[49]
TRAPPC2
Spondyloepiphyseal
dysplasia
tarda
SEDT
[50]
Biogenesis/Assembly FUS
Amyotrophic
lateral
sclerosis
ALS
[64]
SMN1
Spinal
muscular
atrophy SMA
[10,57–61]
Fig.
2.
Disease-associated
mutations
in
the
specific
protein
and
snRNA
components
of
the
minor
spliceosome.
A)
Domain
structure
of
the
U11/U12-65K
proteins
with
the
locations
of
IGHD-associated
mutations
indicated.
B)
Secondary
structure
of
U12
snRNA
showing
the
location
of
a
point
mutation
associated
with
EOCA.
Binding
site
for
the
U11/U12-65K
protein
is
also
indicated.
C)
Secondary
structure
of
the
U4atac/U6atac
di-snRNA
complex
showing
mutations
associated
with
MOPDI
(red)
and
RFMN
(cyan).
A
mutation
shared
between
MOPD1/TALS
and
RFNM
patients
is
indicated
in
red
text
surrounded
by
a
cyan
rectangle.
Binding
sites
for
15.5K,
PRPF31,
the
PRPF4/PRPF3/PPIH
complex
proteins
and
the
U11/U12-65K
protein
are
also
shown.
D)
Domain
structure
of
the
ZRSR2
protein
showing
mutations
associated
with
MDS.
Mutations
are
from
the
COSMIC
database
[76].
Mutations
studied
by
Madan
et
al.
[29]
are
shown
in
blue.
was
carried
out
on
P474T/R502X
patient
lymphocytes
[27].
Consis-
tent
with
the
function
of
the
65K
protein
in
the
intron
recognition
complex,
several
types
of
aberrant
splicing
events
associated
with
a
subset
of
U12-type
introns
were
detected,
including
increased
retention
of
U12-type
introns,
activation
of
nearby
cryptic
U2-
type
splice
sites
and
exon
skipping
events.
Biochemical
analysis
of
patient
cells
revealed
a
reduction
in
levels
of
the
65K
protein,
as
well
as
reduced
stability
of
the
U11/U12
di-snRNP
complex.
This
result
is
consistent
with
the
function
of
the
65K
protein
as
a
component
of
themolecular
bridge
between
U11
and
U12
snRNPs
through
interactions
with
the
U11-59K
protein
and
U12
snRNA
[33].
The
65K-U12
snRNA
interaction
is
mediated
by
the
C-terminal
RRM
domain
and,
as
predicted,
the
IGHD
mutations
were
shown
to
either
reduce
(P474T)
or
eliminate
(R502X)
the
binding
of
65K

Please
cite
this
article
in
press
as:
B.
Verma,
et
al.,
Minor
spliceosome
and
disease,
Semin
Cell
Dev
Biol
(2017),
https://doi.org/10.1016/j.semcdb.2017.09.036
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(2017)
xxx–xxx
5
to
the
stem-loop
III
of
U12
snRNA
(Fig.
2B;
[36]).
The
picture
that
is
emerging
from
these
studies
is
that
the
IGHD
mutations
lead
to
impaired
interaction
between
the
65K
protein
and
U12
snRNA,
which
in
turn
causes
destabilization
or
reduced
formation
of
the
U11/U12
di-snRNP.
Additionally,
it
is
likely
that
both
RNPC3
non-
sense
mutations
described
for
IGHD
lead
to
allele-specific
decay
of
the
65K
mRNA
via
nonsense
mediated
decay
(NMD)
pathway,
as
shown
recently
for
the
R502X
allele
[36].
This
is
expected
to
signif-
icantly
reduce
the
levels
of
65K
proteins
encoded
by
the
nonsense
alleles.
However,
we
note
that
additional
mechanisms
may
be
at
play,
as
it
is
not
currently
known
whether
the
65K
protein
is
purely
a
structural
component
of
the
U11/U12
di-snRNP,
or
whether
it
has
additional
functions
in
the
splicing
process.
A
zebrafish
study
indi-
cated
that
the
65K
protein
may
have
other
functions
later
in
the
spliceosome
assembly
[37].
Consistently,
recent
work
reported
an
interaction
between
65K
and
the
3terminal
stem-loop
of
U6atac
snRNA
(Fig.
2C;
[38]),
which
is
similarly
impaired
by
the
IGHD
mutations
[36].
2.1.2.
Cerebellar
ataxia
A
mutation
in
the
noncoding
U12
snRNA
gene
(RNU12)
has
recently
been
associated
with
an
early-onset
cerebellar
ataxia
(EOCA)
[28].
In
general,
ataxias
are
a
diverse
group
of
disorders
characterized
by
defects
in
muscle
coordination
which,
in
the
case
of
cerebellar
ataxias,
results
from
abnormal
development
and/or
degeneration
of
the
cerebellum.
The
patients
in
this
single
study
are
homozygous
for
a
point
mutation
(84C
>
T;
see
Fig.
2B)
with
symptoms
of
hypotonia
at
infancy,
delayed
motor
development,
abnormal
gait,
speech
and
learning
difficulties
[28].
RNAseq
and
RT-PCR
analyses
of
patient
mononuclear
cells
showed
increased
U12-type
intron
retention
and
upregulation
of
the
U12
snRNA.
The
84C
>
T
mutation
is
located
at
the
base
of
U12
snRNA
stem-
loop
III,
and
is
predicted
to
weaken
the
stem-closing
G-C
base-pair
by
converting
it
to
a
weaker
G-U
base-pair
(Fig.
2B).
Mechanistic
understanding
of
how
this
mutation
leads
to
a
defect
in
minor
spliceosome
function
is
currently
lacking.
The
mutation
does
not
directly
overlap
with
any
known
functional
elements,
such
as
sequences
involved
in
known
RNA–RNA
or
RNA–protein
interac-
tions;
however,
the
apical
part
of
this
long
stem-loop
acts
as
the
binding
site
for
the
65K
protein
(Fig
2B;
[33])
and
the
assembly
site
for
the
Sm
protein
ring,
an
integral
part
of
all
snRNPs,
is
located
only
two
nucleotides
away.
Furthermore,
as
the
U12
snRNA
not
only
recognizes
the
branch
point
sequence
during
intron
recogni-
tion,
but
is
also
part
of
the
catalytic
core
of
the
minor
spliceosome,
several
additional
mechanistic
scenarios
are
conceivable.
Further
functional
studies
are
needed
to
uncover
the
mechanistic
basis
of
this
disease.
2.1.3.
Myelodysplastic
syndrome
Myelodysplastic
syndrome
(MDS;
myelodysplasia)
is
a
term
for
a
group
of
disorders
characterized
by
inefficient
hematopoiesis,
with
a
risk
of
progression
to
acute
myeloid
leukemia
(AML).
In
recent
years,
several
whole-exome
sequencing
studies
have
revealed
frequent
somatic
mutations
in
genes
encoding
splicing
factors,
including
U2AF1,
ZRSR2,
SRSF2
and
SF3B1
in
patients
with
MDS
reviewed
by
[9,39].
Although
patients
with
a
combination
of
different
splicing
factor
mutations
have
been
reported,
typically
only
one
splicing
factor
gene
in
each
patient
has
been
mutated.
Fur-
thermore,
the
splicing
factor
mutations
frequently
co-occur
with
mutations
in
genes
encoding
epigenetic
factors,
cell
signaling
reg-
ulators
and
transcriptional
regulators
[39,40].
Here,
we
focus
on
the
Urp/ZRSR2
protein
involved
in
U12-type
3ss
recognition.
The
ZRSR2
protein
shares
both
primary
structure
and
functional
features
with
the
splicing
factor
U2AF1,
that
facilitates
3ss
recogni-
tion
with
the
major
introns.
Both
contain
a
central
U2AF-homology
domain
(UHM)
flanked
on
each
side
by
a
CCCH-type
zinc
finger,
and
an
RS
domain
near
the
C-terminus
(Fig.
2D).
ZRSR2
is
an
integral
component
of
the
U11/U12
di-snRNP
and
functions
in
the
recogni-
tion
of
the
U12-type
3splice
site,
but
it
has
also
been
reported
to
function
during
the
second
catalytic
step
of
U2-type
intron
splicing
[19,25].
MDS-associated
mutations
in
ZRSR2
include
a
wide
spec-
trum
of
nonsense,
missense,
frameshift
and
splice
site
mutations
which
are
uniformly
scattered
across
the
gene
(Fig.
2D).
This
distri-
bution
is
in
striking
contrast
to
MDS
cases
with
mutations
in
U2AF1,
SRSF2
and
SF3B1
genes
that
show
a
narrower
mutation
spectrum,
with
particular
preference
for
missense
mutations
that
are
located
mostly
within
a
small
number
of
hotspot
positions.
Of
the
numerous
somatic
mutations
reported
in
the
ZRSR2
(Fig.
2D),
only
few
have
been
studied
in
detail
[29].
In
the
Madan
et
al.
[29]
RNAseq
study,
bone
marrow
samples
from
eight
ZRSR2-
mutant
MDS
patients,
each
carrying
a
different
mutation,
were
compared
to
both
ZRSR2
wild-type
MDS
patients
and
healthy
indi-
viduals.
The
analysis
revealed
a
global
increase
in
U12-type
intron
retention,
as
well
as
activation
of
cryptic
U2-type
splice
sites
only
in
MDS
patients
carrying
the
mutated
ZRSR2
allele.
Importantly,
the
effect
of
ZRSR2
mutations
and
knockdown
of
ZRSR2
on
U2-type
intron
splicing
were
comparatively
small.
The
affected
U2-type
introns
were
mostly
in
genes
containing
U12-type
introns.
Similar
conclusions
have
also
been
reported
in
plants
[41].
These
findings
are
in
stark
contrast
to
the
earlier
biochemical
experiments
which
suggested
that
an
essential
role
for
ZRSR2
was
in
splicing
of
both
U2-
and
U12-type
introns
[19].
2.2.
Diseases
affecting
the
formation
of
catalytically
active
spliceosome
2.2.1.
MOPD1/TALS
Recessive
mutations
in
the
U4atac
snRNA
gene
(RNU4atac)
associated
with
MOPD1/TALS,
a
rare
autosomal
recessive
disease,
represent
the
first
human
disease
with
mutations
in
a
specific
component
of
the
minor
spliceosome
[31,32,42].
U4atac
snRNA
is
an
essential
component
of
the
U4atac/U6atac.U5
tri-snRNP
(minor
tri-snRNP),
which
joins
the
nascent
spliceosome
after
the
intron
boundaries
have
been
recognized
by
the
U11/U12
di-snRNP
(Fig.
1B).
During
the
assembly
of
the
minor
tri-snRNP,
U4atac
forms
an
extensively
base-paired
duplex
with
the
U6atac
snRNA
(Figs.
1B
and
2C).
This
U4atac/U6atac
di-snRNA
complex
forms
a
platform
for
the
binding
of
tri-snRNP-specific
proteins
that
are
needed
for
the
association
of
the
U5
snRNP
to
the
complex.
MOPD1/TALS
in
its
severe
form
is
characterized
by
intrauterine
and
postnatal
growth
retardation,
developmental
defects
in
sev-
eral
organs,
including
microcephaly,
and
death
typically
within
3
years
after
birth.
However,
milder
forms
of
the
disease
have
been
reported
displaying
more
subtle
growth
retardation,
less
severe
developmental
defects
and/or
survival
to
at
least
to
12
years
of
age
or
even
adulthood
[43,44].
Presently,
individual
single-nucleotide
point
mutations
at
ten
different
positions
within
the
U4atac
snRNA
have
been
associated
with
MOPD1/TALS
(Fig.
2C);
in
addition,
one
patient
with
a
duplication
of
U4atac
nucleotides
16–100
has
been
reported
[44].
All
patients
are
either
homozygous
or
compound
heterozygous
for
the
disease
mutations.
In
contrast,
heterozy-
gous
parents
carrying
only
a
single
disease-causing
mutation
are
phenotypically
normal,
suggesting
that
a
loss
of
a
single
allele
is
well-tolerated
at
the
cellular
level.
The
majority
of
disease-causing
mutations,
particularly
those
leading
to
severe
forms
of
the
disease,
are
found
in
the
U4atac
5stemloop,
with
only
few
pathogenic
mutations
located
else-
where
in
the
U4atac
snRNA
(Fig.
2C).
Earlier
studies
from
the
major
spliceosome
have
indicated
that
this
region
is
recognized
by
a
group
of
proteins
(Fig.
2C),
particularly
the
15.5K
(Snu13)
and
PRPF31
pro-
teins,
that
are
both
necessary
for
the
association
of
U5
snRNP
to

Please
cite
this
article
in
press
as:
B.
Verma,
et
al.,
Minor
spliceosome
and
disease,
Semin
Cell
Dev
Biol
(2017),
https://doi.org/10.1016/j.semcdb.2017.09.036
ARTICLE IN PRESS
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No.
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Pages
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6
B.
Verma
et
al.
/
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in
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&
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Biology
xxx
(2017)
xxx–xxx
the
tri-snRNP
[45].
Consistently,
the
MOPD1/TALS
mutations
in
this
region
reduce
the
15.5K
binding,
which
in
turn
decreases
the
cellu-
lar
levels
of
U4atac/U6atac.U5
tri-snRNP
[46].
Importantly,
there
is
a
correlation
between
disease
severity
and
the
affinity
of
the
15.5K
protein.
Mutations
on
the
tip
of
the
5stemloop
that
reduce
most
the
15.5K
binding,
also
lead
to
severe
forms
of
the
disease.
In
con-
trast,
the
more
distal
mutations
(such
as
U4atac
positions
30,
53
and
55–see
Fig.
2C)
have
little
effect
on
the
binding
of
the
15.5K
pro-
tein
and
are
consequently
associated
with
milder
symptoms,
such
as
longer
life
expectancy
[43,46].
Additionally,
at
least
one
of
the
mutations
(124G
>
A)
reduces
the
U4atac
snRNA
expression
levels
[46].
No
transcriptome
studies
have
yet
been
conducted
for
MOPD1/TALS,
but
qPCR
analyses
have
shown
increased
U12-type
intron
retention
in
patient
cells
that
can
be
rescued
by
expression
of
wild
type
U4atac
snRNA
[31,32].
Significantly,
an
analysis
with
a
reporter
system
indicates
that
>90%
of
the
minor
spliceosome
activity
may
have
been
compromised
[31].
However,
it
is
possible
that
the
reporter
system
used
may
have
exacerbated
the
splic-
ing
defect
as
the
endogenous
transcripts
in
patient
cell
lines
and
iPS
cells
carrying
the
mutation
show
much
milder
splicing
defects
[31,32,46].
2.2.2.
Roifman
syndrome
In
addition
to
MOPD1/TALS,
mutations
in
the
RNU4atac
locus
have
been
associated
with
a
phenotypically
different
Roifman
syndrome
(RFMN)
[30].
Patients
suffering
from
RFMN
are
char-
acterized
by
poor
pre-
and
post-natal
growth,
distinct
facial
dystrophies,
cognitive
delay
and
immunological
defects
which,
with
the
exception
of
growth
defects,
are
distinct
to
Roifman
patients
[30].
Interestingly,
all
RFMN
patients
are
compound
het-
erozygotes,
with
one
mutation
shared
with
MOPD1/TALS
patients
with
severe
forms
of
the
diseases
(Fig.
2C).
The
other
mutation
is
predominantly
located
among
the
highly
conserved
positions
of
the
RNU4atac
stem
II,
except
for
a
single
case
with
a
mutation
within
the
Sm
site.
No
functional
studies
have
been
performed
with
the
RFMN
mutations,
but
it
is
safe
to
assume
that
the
mutations
shared
with
the
MOPD1/TALS
lead
to
a
similar
defect
in
minor
tri-snRNP
assem-
bly
[46].
In
contrast,
the
stem
II
mutations
presumably
lead
to
a
milder
functional
defect
in
minor
splicing,
given
the
overall
less
severe
disease
phenotype.
While
the
molecular
defects
resulting
from
the
stem
II
mutations
remain
to
be
determined,
the
investiga-
tion
from
the
major
spliceosome
assembly
tentatively
suggest
that
the
association
of
the
PRPF4/PRPF3/PPIH
complex
(Fig.
2C)
with
the
stem
II
may
be
compromised
[30,47,48].
In
addition,
RNAseq
analyses
of
the
patient
cells
revealed
a
widespread
retention
of
U12-type
introns.
Significantly,
neither
increased
retention
of
U2-type
introns,
nor
other
types
of
alter-
native
splicing
changes
were
detected
in
the
patient
cells
[30],
suggesting
a
very
specific
splicing
defect
affecting
U12-type
introns
only.
2.3.
Diseases
with
mutations
in
the
U12-type
splice
sites
To
our
knowledge
there
are
only
two
reported
cases
in
lit-
erature
that
specifically
attribute
U12-type
splice
site
mutations
with
human
disease.
These
include
the
autosomal
dominant
dis-
order
Peutz-Jeghers
syndrome
(PJS),
characterized
by
the
presence
of
multiple
intestinal
polyps
and
a
high
risk
for
cancer
[49],
and
Spondyloepiphyseal
dysplasia
tarda
(SEDT),
an
X-linked
recessive
disorder
[50].
Both
are
caused
by
mutations
in
a
U12-type
5ss
that
lead
to
incorrect
splicing
of
the
respective
mRNAs.
Additionally,
the
Motor
endplate
disease
in
mouse
reports
a
mutation
of
a
U2-type
5ss
in
a
downstream
intron,
leading
to
skipping
of
the
upstream
U12-type
intron
[51].
These
three
cases
do
not
only
demonstrate
that
a
single
point
mutation
in
the
highly
conserved
U12-type
5ss
sequence
can
efficiently
disable
the
U12-type
intron
recognition,
but
also
that
splicing
of
a
U12-type
intron
is
linked
to
other
splicing
events
in
the
same
transcript
and
that
in
some
cases
the
outcome
may
be
difficult
to
predict.
PJS,
in
which
a
mutation
at
the
+1
position
of
an
AT–AC-type
U12-type
intron
is
changed
to
G,
is
a
particu-
larly
illuminating
case
[49].
At
the
sequence
level,
it
appears
to
be
a
simple
U12-type
5ss
subtype
change,
but
the
experimentally
determined
outcome
is
in
fact
complex,
showing
activation
of
sev-
eral
non-canonical
3splice
sites
that
illuminate
the
flexibility
of
the
U12-dependent
spliceosome
in
3ss
recognition.
Even
though
the
outcome
in
each
case
is
the
same,
i.e.
introduction
of
a
pre-
mature
STOP-codon
and
mRNA
decay
via
the
NMD
pathway,
this
particular
mutation
demonstrates
that
simple
point
mutations
can
lead
to
unexpected
consequences.
2.4.
Other
spliceosome
mutations
affecting
minor
spliceosome
activity
In
addition
to
diseases
caused
by
mutations
in
the
specific
components
of
the
minor
spliceosome,
there
are
several
addi-
tional
human
diseases
caused
by
congenital
or
somatic
mutations
in
either
the
shared
components
of
the
two
spliceosomes
or
the
assembly
factors
necessary
for
both
major
and
minor
snRNPs.
Examples
of
the
former
include
Retinitis
pigmentosa
(RP)
mutations
in
genes
encoding
the
tri-snRNP
specific
proteins
PRPF3,
PRPF4,
PRPF6,
PRPF8,
PRPF31
and
BRR2
[Reviewed
in
8],
and
MDS
muta-
tions
in
PRPF8
[52],
in
SF3b1,
a
component
of
both
U2
snRNP
and
U11/U12
di-snRNP
[26],
and
in
SRSF2,
a
splicing
activator
poten-
tially
affecting
both
spliceosomes
[53].
Spinal
muscular
atrophy
(SMA)
represents
the
latter
group
and
is
caused
by
mutations
in
the
SMN1
gene,
coding
for
an
essential
factor
for
the
assembly
of
all
Sm-class
snRNPs,
including
components
of
both
major
and
minor
spliceosomes
[54].
With
RP,
the
shared
protein
composition
between
the
minor
and
major
tri-snRNPs
[17]
and
functional
studies
[55,56]
predict
that
the
mutations
should
have
an
equal
effect
on
both
the
major
and
minor
tri-snRNPs.
With
SMA,
however,
several
investigations
using
different
model
systems
have
reported
a
preferential
reduc-
tion
in
the
cellular
levels
of
minor
snRNPs
[10,57–61].
In
a
subset
of
the
studies
this
has
been
reported
to
lead
to
an
increased
retention
U12-type
introns,
suggesting
that
defects
in
the
splicing
of
minor
introns
may
contribute
to
the
SMA
symptoms.
In
a
Drosophila
model
of
SMA
the
effects
on
neuronal
cells
were
attributed
to
a
single
U12-type
intron
containing
gene,
Stasimon,
encoding
a
transmem-
brane
protein
required
for
axonal
growth
and
regulation
of
synaptic
transmission
of
motor
neurons
[60].
However,
some
aspects
of
this
study
have
been
challenged
subsequently
[62].
Finally,
two
independent
studies
have
linked
minor
spliceo-
some
to
amyotrophic
lateral
sclerosis
(ALS).
Ishihara
et
al.
[63]
reported
that
ALS-associated
depletion
of
TDP-43
protein
leads
to
a
co-depletion
of
U12
snRNA
both
in
cultured
cells
and
in
patient
samples,
presumably
due
to
defects
in
snRNP
assembly.
In
another
study
[64],
ALS-associated
mutations
in
the
Fused
in
sarcoma
(FUS)
protein
lead
to
reduced
nuclear
import
of
FUS
and
its
accumulation
in
the
cytoplasm.
FUS
is
a
multifunctional
RNA-binding
protein
that
interacts
with
several
splicing
factors,
including
U11
snRNP
and
is
needed
for
the
splicing
of
at
least
a
subset
of
minor
introns
[64].
Sig-
nificantly,
the
cytoplasmic
accumulation
of
a
particular
FUS
mutant
leads
to
a
concomitant
mislocalization
of
U11
and
U12
snRNPs
in
the
cytoplasm,
which
in
turn
leads
to
defects
in
the
splicing
of
U12-type
introns.

Please
cite
this
article
in
press
as:
B.
Verma,
et
al.,
Minor
spliceosome
and
disease,
Semin
Cell
Dev
Biol
(2017),
https://doi.org/10.1016/j.semcdb.2017.09.036
ARTICLE IN PRESS
G Model
YSCDB-2404;
No.
of
Pages
10
B.
Verma
et
al.
/
Seminars
in
Cell
&
Developmental
Biology
xxx
(2017)
xxx–xxx
7
Fig.
3.
Model
for
molecular
consequences
of
the
minor
spliceosome
diseases.
Mutations
in
the
minor
spliceosome
components
can
lead
to
nuclear
retention
of
the
partially
spliced
mRNA,
or
intron
retention
and
associated
activation
of
cryptic
U2-type
splice
sites
or
exon
skipping
events.
These
in
turn
lead
to
nuclear
or
cytoplasmic
decay,
or
formation
of
the
abnormal
proteins
as
indicated.
3.
Minor
spliceosome
components
as
biomarkers
In
addition
to
being
a
target
for
disease-causing
mutations,
sev-
eral
components
of
the
minor
spliceosome
have
been
reported
as
cancer
biomarkers.
A
recent
study
by
Xu
et
al.
[65]
using
serum
from
patients
with
scleroderma
and
coincident
cancer
reported
autoantibodies
targeted
to
components
of
the
U11/U12
di-snRNP,
particularly
the
U11/U12-65K
protein,
but
also
the
25K,
35K
and
59K
proteins.
Of
these,
the
U11/U12-65K
autoantibodies
appear
to
show
a
reliable
association
with
an
increased
risk
of
cancer
occurring
within
2
years
of
the
onset
of
scleroderma
[66].
Similar
associations
have
been
described
for
U11
snRNA
in
familial
prostate
cancers
[67]
and
U11-59K
protein
in
Acute
Myeloid
Leukemia
[68].
However,
the
role
of
minor
spliceosome-specific
autoantibodies
in
the
pathogenesis
of
scleroderma
and/or
coincident
cancer,
or
the
expression
level
changes
of
other
minor
spliceosome
components
in
other
diseases
is
presently
unclear.
4.
Consequences
of
minor
spliceosome-specific
mutations
Presently,
mutations
in
four
specific
components
of
the
minor
spliceosome
have
been
associated
with
human
diseases
showing
very
different
disease
phenotypes.
RT-PCR
and
RNAseq
analyses
have
demonstrated
that
each
of
the
five
human
diseases
show
the
expected
molecular
level
phenotype,
i.e.
the
increased
levels
of
unspliced
U12-type
introns
in
the
patient
cells.
With
a
subset
of
diseases,
additional
splicing
defects,
particularly
activation
of
cryp-
tic
U2-type
splice
sites
and
exon
skipping
events
have
also
been
described.
Presently,
there
has
not
been
clear
consensus
on
how
to
interpret
the
molecular
consequences
of
minor
spliceosome
muta-
tions.
Here,
we
propose
a
simple
classification
of
disease-causing
mutations
according
to
their
impact
on
the
recognition
of
the
U12-
type
introns.
This
provides
a
simple
framework
for
predicting
the
molecular
consequences
of
the
mutations,
but
may
also
help
to
interpret
the
subsequent
disease
phenotypes.
4.1.
The
fate
of
the
mRNA:
intron
retention
vs
cryptic
splicing
According
to
this
classification
the
mutations
associated
with
both
the
MOPD1/TALS
and
RFMN
affect
the
formation
of
the
U4atac/U6atac.U5
tri-SNRP
(see
2.2.1
and
2.2.2),
but
not
the
initial
intron
recognition
step.
Therefore,
in
these
diseases
the
spliceo-
some
assembly
is
arrested
at
the
pre-spliceosome
stage
where
the
U11/U12
di-snRNP
is
bound
to
the
intron
but
the
subsequent
assembly
steps
are
inhibited.
Consequently,
the
bound
U11/U12
di-snRNP
can
be
expected
to
maintain
exon-definition
interactions
with
the
surrounding
(U2-type)
introns
via
the
RS
domain
contain-
ing
U11-35K
[69]
and
possibly
Urp/ZRSR2
proteins.
Furthermore,
the
U11/U12
di-snRNP
bound
to
the
partially
spliced
mRNA
can
trap
such
transcripts
in
the
nucleus
[70],
where
they
may
be
targeted
by
the
nuclear
quality
control
mechanisms
[71,72].
In
contrast,
the
loss
of
U11/U12
binding
and
the
recognition
of
U12-type
introns
in
IGHD
and
MDS
(and
possibly
EOCA)
leads
to
a
different
outcome.
Due
to
the
loss
of
U11/U12
binding,
transcripts
containing
unspliced
U12-type
introns
are
free
to
be
exported
to
the
cytoplasm.
Concomitantly,
the
loss
of
local
exon
definition
interac-
tions
may
lead
to
increased
activation
of
cryptic
splice
sites
or
exon
skipping
events
near
the
U12-type
introns
[73].
The
outcome
can
be
either
truncated
or
altered
protein,
or
mRNA
decay
via
the
NMD
quality
control
pathway
(Fig.
3).
Is
there
evidence
supporting
this
model?
RNAseq
analyses
from
both
IGHD
and
MDS
patient
cells
provide
extensive
evidence
of
acti-
vation
of
cryptic
U2-type
splice
sites
and
increased
levels
of
exon
skipping
near
U12-type
introns
in
addition
to
the
expected
reten-
tion
of
U12-type
introns
[27,29].
In
contrast,
besides
the
increased
intron
retention
of
U12-type
introns
seen
with
both
MOPD1/TALS
and
RFMN,
there
is
no
evidence
of
the
associated
cryptic
U2-type

Please
cite
this
article
in
press
as:
B.
Verma,
et
al.,
Minor
spliceosome
and
disease,
Semin
Cell
Dev
Biol
(2017),
https://doi.org/10.1016/j.semcdb.2017.09.036
ARTICLE IN PRESS
G Model
YSCDB-2404;
No.
of
Pages
10
8
B.
Verma
et
al.
/
Seminars
in
Cell
&
Developmental
Biology
xxx
(2017)
xxx–xxx
splicing
events
[30–32,46]
as
stated
by
Merico
et
al.
[30]
in
their
detailed
RNAseq
analysis
of
RFMN
patient
cells.
Together,
present
evidence
supports
the
proposal
that
the
consequences
of
the
minor
spliceosome
mutations
are
linked
to
the
type
of
assembly
defect
introduced.
However,
with
both
MOPD1/TALS
and
RFMN
it
is
pos-
sible
that
some
arrested
transcripts
trapped
in
the
nucleus
may
either
lose
the
bound
U11/U12
di-snRNP
or
undergo
the
splicing
process
post-transcriptionally
(see
Fig.
3).
4.2.
Fate
of
the
mRNA
and
disease
severity
It
is
somewhat
puzzling
to
note
that
the
severity
of
congen-
ital
minor
spliceosome
diseases
and
the
level
of
mRNA
splicing
defect
follow
a
counterintuitive
association
as
described
above.
The
two
diseases
showing
only
relatively
mild
intron
retention
defects
(MOPD1/TALS
and
RFMN)
show
pleiotropic,
and
often
severe,
defects
in
many
tissues
[30–32].
In
contrast,
IGHD
and
EOCA
show
large
splicing
defects
that
were
detected
presumably
ubiq-
uitously
−
also
from
cells/tissues
not
associated
with
the
disease
[27,28].
Even
though
the
present
small
dataset
does
not
allow
gen-
eralization,
it
is
tempting
to
speculate
that
the
difference
may
be
linked
to
the
fate
of
the
unspliced
mRNAs.
Specifically,
it
is
possible
that
the
unspliced
U12-type
introns
or
transcripts
con-
taining
cryptic
splicing
events
are
better
tolerated
when
exported
to
cytoplasm
where
they
can
be
targeted
by
the
cytoplasmic
qual-
ity
control
mechanisms.
Transcripts
trapped
in
the
nucleus
may,
in
contrast,
accumulate
in
some
tissues
or
lead
to
other
detrimen-
tal
downstream
effects.
This
hypothesis
receives
tentative
support
from
a
zebrafish
study
where
a
specific
65K
mutation
arrests
the
spliceosome
assembly
at
a
late
stage
and
presumably
traps
such
mRNAs
in
the
nucleus.
Unlike
the
more
restricted
phenotype
seen
with
IGHD
65K
mutations,
the
zebrafish
phenotype
is
severe,
with
developmental
defects
in
multiple
organs
[37].
4.3.
Defects
with
individual
genes
vs
a
global
splicing
defect
One
of
the
outstanding
questions
is
whether
the
pathologi-
cal
consequences
of
the
minor
spliceosome
diseases
are
caused
by
a
global
defect
in
the
splicing
of
U12-type
introns
or
whether
the
various
effects
seen
in
different
tissues
are
caused
by
mis-
splicing
of
a
small
subset
of
genes
containing
U12-type
introns.
Gene
expression
analyses
have
suggested
various
candidate
genes
responsible
for
the
observed
disease
phenotypes.
These
must,
how-
ever,
be
interpreted
with
caution
since
in
all
cases
except
MDS,
the
gene
expression
analyses
have
been
done
using
cells
other
than
the
actual
affected
tissue.
Furthermore,
it
is
not
yet
possible
to
conclude
whether
a
par-
ticular
tissue
or
cell
type
would
be
more
susceptible
to
defects
in
the
U12-dependent
spliceosome.
Of
the
present
congenital
minor
spliceosome
diseases,
the
only
common
target
appears
to
be
the
nervous
system.
Specifically,
microcephaly
has
been
reported
for
three
of
the
diseases
(severe
for
MOPD1/TALS,
mild
for
RFMN
and
IGHD),
which
with
MOPD1/TALS
is
associated
with
struc-
tural
abnormalities
of
brain.
Additionally,
RFMN
and
EOCA
patients
display
cognitive
delay
and
cerebellar
ataxia,
respectively.
The
observed
tissue-specific
effects
may
be
linked
to
the
associations
of
the
minor
spliceosome
activity
with
cell
proliferation
[37,74]
or
cel-
lular
response
to
stress
[75].
Consistently,
recent
work
has
reported
that
components
of
the
U11/U12
di-snRNP,
including
the
65K,
48K
and
Urp/ZRSR2
proteins
are
downregulated
during
neuron
terminal
differentiation
[70].
5.
Future
perspective
The
recent
discoveries
of
human
diseases
affecting
the
minor
spliceosome
have
significantly
advanced
our
understanding
of
the
significance
and
function
of
the
U12-dependent
spliceosome.
How-
ever,
many
aspects
of
the
disease
process,
including
the
basis
of
the
tissue
specificity
and
the
detailed
mechanism
of
the
disease
process
are
yet
to
be
determined
for
most
diseases,
requiring
animal
models
of
each
disease.
These
will
also
help
to
address
the
more
fundamen-
tal
question
of
the
existence
of
two
separate
spliceosomes.
Acknowledgements
This
work
was
supported
by
the
Academy
of
Finland
(Grant
140087
to
MJF
and
Grant
278798
to
BV)
and
Sigrid
Jusélius
Foun-
dation
(MJF).
AJN
was
supported
by
the
Integrative
Life
Science
doctoral
program
at
the
University
of
Helsinki.
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Please
cite
this
article
in
press
as:
B.
Verma,
et
al.,
Minor
spliceosome
and
disease,
Semin
Cell
Dev
Biol
(2017),
https://doi.org/10.1016/j.semcdb.2017.09.036
ARTICLE IN PRESS
G Model
YSCDB-2404;
No.
of
Pages
10
B.
Verma
et
al.
/
Seminars
in
Cell
&
Developmental
Biology
xxx
(2017)
xxx–xxx
9
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Minor
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disease,
Semin
Cell
Dev
Biol
(2017),
https://doi.org/10.1016/j.semcdb.2017.09.036
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YSCDB-2404;
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