Mutations in SRCAP, Encoding SNF2-Related CREBBP
Activator Protein, Cause Floating-Harbor Syndrome
Rebecca L. Hood,1,2 Matthew A. Lines,3 Sarah M. Nikkel,3,4 Jeremy Schwartzentruber,5
Chandree Beaulieu,6 Ma1gorzata J.M. Nowaczyk,7 Judith Allanson,3 Chong Ae Kim,8
Dagmar Wieczorek,9 Jukka S. Moilanen,10 Didier Lacombe,11 Gabriele Gillessen-Kaesbach,12
Margo L. Whiteford,13 Caio Robledo D.C. Quaio,8 Israel Gomy,8 Debora R. Bertola,8 Beate Albrecht,9
Konrad Platzer,12 George McGillivray,14 Ruobing Zou,2 D. Ross McLeod,15 Albert E. Chudley,16,17
Bernard N. Chodirker,16,17 Janet Marcadier,6 FORGE Canada Consortium,18 Jacek Majewski,5,19
Dennis E. Bulman,2,* Susan M. White,14,20 and Kym M. Boycott3,4,6,*
Floating-Harbor syndrome (FHS) is a rare condition characterized by short stature, delayed osseous maturation, expressive-language deficits,
and a distinctive facial appearance. Occurrence is generally sporadic, although parent-to-child transmission has been reported on
occasion. Employing whole-exome sequencing, we identified heterozygous truncating mutations in SRCAP in five unrelated individuals
with sporadic FHS. Sanger sequencing identified mutations in SRCAP in eight more affected persons. Mutations were de novo in all six
instances in which parental DNA was available. SRCAP is an SNF2-related chromatin-remodeling factor that serves as a coactivator for
CREB-binding protein (CREBBP, better known as CBP, the major cause of Rubinstein-Taybi syndrome [RTS]). Five SRCAP mutations, two
of which are recurrent, were identified; all are tightly clustered within a small (111 codon) region of the final exon. These mutations are
predicted to abolish three C-terminal AT-hook DNA-binding motifs while leaving the CBP-binding and ATPase domains intact. Our findings
show that SRCAP mutations are the major cause of FHS and offer an explanation for the clinical overlap between FHS and RTS.
Floating-Harbor syndrome (FHS [MIM 136140]) is a rare
condition characterized by short stature, delayed osseous
maturation, language deficits, and a distinctive facial
appearance. The dysmorphic features typical of this
disorder include a triangular face, short philtrum, wide
mouth with a thin vermilion border of the upper lip, and
long nose with a narrow bridge, broad base, full tip, and
low-hanging columella.1–4 Some degree of intellectual or
learning disability is present in most individuals, and
language (both receptive and expressive) is typically
more severely affected. The name ‘‘Floating Harbor’’ is
a portmanteau of Boston Floating Hospital and Harbor
General Hospital (Torrance, CA), the two institutions
from which the initial case reports originated.1,2 Of the
50 or so cases of FHS in the literature, the majority are
sporadic, although four reported instances of parent-tochild
transmission suggest that this is an autosomal-dominant
disorder in at least some instances.4–7 Some authors
have highlighted the clinical overlap between FHS and
Rubinstein-Taybi syndrome (RTS [MIM 180849]), which
shares several key features (short stature, a long nose
with low-hanging columella, a thin vermilion border of
the upper lip, and anomalous thumbs).3,7 Despite the
recognition of FHS as a distinct clinical entity more than
25 years ago, no causative mutations have been identified
to date.
To identify the genetic basis of FHS, we assembled
a cohort of 13 unrelated probands, three of whom were
previously reported.4 The clinical details of these individuals
are presented in Table 1 and Figure 1. To identify
FHS-causing mutations, we performed exome capture
and high-throughput sequencing of five unrelated affected
persons (probands 1–5). Approval of the study design was
obtained from the institutional research ethics board
(Children’s Hospital of Eastern Ontario), and free and
informed consent was obtained from each study subject
(or parent, if appropriate) prior to enrollment. We performed
exome target enrichment by using the Agilent
SureSelect 50 Mb All Exon Kit, and sequencing (Illumina
HiSeq) generated 35–40 Gbp of 100 bp paired-end reads
per sample. Reads were preprocessed (trimmed) and aligned
to hg19 (seeWeb Resources for list of tools).We used an inhouse
annotation pipeline to identify coding and splicesite
variants that met a minimum quality threshold (i.e.,
R20% of reads supported the variant). Next, we filtered
the variants to exclude common polymorphisms (>1%
minor-allele frequency) represented in dbSNP131, in the
1000 Genomes pilot release, or in 270 exomes sequenced
for individuals with unrelated disorders at our center.
Presuming FHS to be an autosomal-dominant condition,
we identified genes containing a single rare variant in each
of several probands in a combinatorial fashion. Table 2 lists
the numbers of potential candidate genes containing rare
variants in any n probands as n is increased. Of five
sequenced individuals with classic FHS, we noted that all
contained heterozygous truncating variants clustered in
the final (34th) exon of a single gene, SRCAP (encoding
SNF2-related CREBBP activator protein). To confirm SRCAP
as the gene mutated in FHS, we identified SRCAP exon 34
mutations with Sanger sequencing in a validation cohort
of eight more unrelated probands (Table 1 and Figure 2;
Figure S1, available online). All of these mutations are truncating
(nonsense or frameshift) alleles, tightly clustered
between codons 2,407 and 2,517; none are represented in
dbSNP131, 1000 Genomes, or the National Heart, Lung,
and Blood Institute (NHLBI) Exome Variant Server (see
Web Resources). Two mutations in particular, c.7330C>T
(NM_006662.2) (p.Arg2444* [NP_006653]) in six individuals
and c.7303C>T (p.Arg2435*) in four individuals,
accounted for the large majority of mutations. FHS-causing
mutations were shown to be de novo in all six instances in
which parental DNA samples were available.
SRCAP encodes a switch/sucrose nonfermentable (SWI/
SNF)-type chromatin-remodeling ATPase identified in
a two-hybrid screen for interacting partners of CREBbinding
protein (CREBBP, hereafter called CBP).8 In reporter
assays, SRCAP is a potent coactivator for CREB and
CBP-mediated transcription.8,9 In transgenic Drosophila,
exogenous SRCAP colocalizes with transcriptionally active
chromatin and augments CBP’s presence at these sites.10
Affinity-purified SRCAP precipitates as a large complex that
catalyzes ATP-dependent substitution of the variant
histone H2A.Z into nucleosomes.11 This activity has been
confirmed by knockdown experiments with natural
promoters, and it is correlated with in vivo target-gene
expression.12 Separately, SRCAP has also been shown to
transduce signals belonging to the nuclear (steroid)
hormone receptor and Notch pathways, indicating that it
has diverse roles in gene expression.10,13
In keeping with its multiple coactivator roles, SRCAP
contains several discrete functional domains.8–10 These
domains include an SNF2-like ATPase, an N-terminal HSA
(Helicase-SANT-associated) domain, and three C-terminal
AT-hook DNA-binding motifs; the CBP interaction domain
of SRCAP is located centrally (Figure 2). Given the structure
of SRCAP, the nonrandom clustering of truncating mutations
seen in our study participants is strongly suggestive
of a dominant-negative disease mechanism due to loss of
one or more critical domain(s), for instance the three
C-terminal AT-hook motifs. Several more arguments support
this. First, in reporter assays, the major transactivation
function of SRCAP is located in a 655 residue C-terminal
fragment abolished by FHS-causing truncations.9 Second,
expression of a construct solely consisting of the CBP interaction
domain of SRCAP strongly inhibits CREB-mediated
transactivation in a dominant-negative fashion.9 Third,
the Database of Genomic Variants (see Web Resources)
contains two HapMap control individuals who bear a
208 kb deletion copy-number variation (#2,209) containing
SRCAP and nine adjacent genes and who have no reported
phenotype.
In general, the phenotype of persons with SRCAP mutations
is concordant with earlier clinical descriptions of FHS
(Table 1 and Figure 1), and nearly all individuals have
short stature and expressive-language impairment. Despite
the remarkable similarity among mutations seen in our
study subjects, cognitive outcomes ranging from ‘‘normal’’
to ‘‘significant intellectual disability’’ were reported. It is
unclear whether genetic modifier(s) and/or currently
unidentified environmental factors are responsible. Many
of our study subjects had additional systemic malformations,
particularly genitourinary (eight individuals) and
cardiac (three individuals) malformations. Again, genotype-
phenotype correlations explaining these features are
lacking. Given that FHS is a dominant condition exhibiting
a high de novo mutation rate, one might expect
a paternal age effect to be present, and indeed the mean
paternal age of the affected individuals was advanced
(36.9 years; range: 29–44 years).
Importantly, our findings suggest a basis for the longrecognized
phenotypic overlap between FHS and RTS,
the latter of which is caused by alterations in CBP or its
homolog, p300.14–16 Because alterations in both CBP and
SRCAP are expected to produce widespread target-gene
dysregulation, further studies are needed before we can
determine which transcriptional targets are primarily
responsible for each phenotype and whether any of these
might be valid therapeutic targets. The availability of
a molecular test for FHS will greatly improve the reliability
of a formerly challenging clinical diagnosis. Historically,
a diagnosis of FHS has relied upon the presence of typical
facial features because many other key findings (e.g., short
stature and language impairment) are nonspecific. The
advent of molecular diagnosis for this condition will allow
us to gain a better appreciation of the true prevalence and
phenotypic spectrum of FHS.
Supplemental Data
Supplemental Data include one figure and can be found with this
article online at http://www.cell.com/AJHG/.
Acknowledgments
The authors would first like to thank the study participants
and their families, without whose participation and cooperation
this work would not have been possible. This work was funded
by the government of Canada through Genome Canada, the
Canadian Institutes of Health Research (CIHR), and the Ontario
Genomics Institute (OGI-049). Additional funding was provided
by Genome Que´bec and Genome British Columbia. K.M.B. is supported
by a Clinical Investigatorship Award from the CIHR Institute
of Genetics. This work was selected for study by the FORGE
Canada Steering Committee, consisting of K. Boycott (University
of Ottawa), J. Friedman (University of British Columbia),
J. Michaud (University of Montreal), F. Bernier (University of
Calgary), M. Brudno (University of Toronto), B. Fernandez (Memorial
University), B. Knoppers (McGill University), M. Samuels
(Universite´ de Montreal), and S. Scherer (University of Toronto).
Received: November 22, 2011
Revised: December 5, 2011
Accepted: December 7, 2011
Published online: January 19, 2012
Saturday, May 5, 2012
Summary of Canadian Discovery of the FHS Gene
Dr. Kym Boycott’s lab in Ontario, Canada recently
discovered “SRCAP” as the gene which causes Floating Harbor Syndrome
(FHS), a condition characterized by short stature, delayed bone age,
speech delay and distinct facial features. While
we are still learning a lot about SRCAP and its role in the body, Dr.
Boycott’s team found that mutations which cause FHS are clustered in on
specific part (exon 34) of the SRCAP gene. This suggests that this is a
very important part of the gene for its function.
While everyone has two copies of the SRCAP gene- one inherited from
their mother and one inherited from their father- individuals with FHS
have a change or “mutation” on one of their copies of SRCAP, making it
unable to work properly. Having a mutation in
one copy of the SRCAP leads to the features seen in individuals with
FHS. To date, all SRCAP mutations reported have been “de novo” or “of
the new.” This terms refers to the fact that the mutations were not
seen in the parents of the individual with FHS,
and therefore, the gene mutation was not inherited from a parent.
Instead, de novo mutations occur spontaneously, or by chance, at the
time of conception of the individual with FHS. These SRCAP mutations
are not caused by environmental insults (what parents
ate, or did) but rather just happen by chance. Because of this, it is
very unlikely that parents of a child with FHS will have another child
with FHS. However, when individuals with FHS have children, because
they already carry the SRCAP mutation, they have
a 50% or 1 in 2 chance of having a child who also has with FHS. There
are genetic testing options to test pregnancies to see if they carry a
FHS mutation.
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