GENE-39397; No.

of pages: 6; 4C:
Gene xxx (2014) xxxxxx

Contents lists available at ScienceDirect

Gene
journal homepage: www.elsevier.com/locate/gene

A complex microcephaly syndrome in a Pakistani family associated

with a novel missense mutation in RBBP8 and a heterozygous
deletion in NRXN1
Zehra Agha a,b,1, Zafar Iqbal b,1, Maleeha Azam a, Maimoona Siddique c, Marjolein H. Willemsen b,
Tjitske Kleefstra b, Christiane Zweier d, Nicole de Leeuw b, Raheel Qamar a,e, Hans van Bokhoven b,f,
a
Department of Biosciences, Faculty of Science, COMSATS Institute of Information Technology, Islamabad, Pakistan
b
Department of Human Genetics, Nijmegen Centre for Molecular Life Sciences, Radboud university medical center, Nijmegen, The Netherlands
c
Shifa International Hospital, Islamabad, Pakistan
d
Institute of Human Genetics, Friedrich-Alexander-University Erlangen-Nuremberg, Erlangen, Germany
e
Al-Nafees Medical College & Hospital, Isra University, Islamabad, Pakistan
f
Department of Cognitive Neuroscience, Donders Institute for Brain, Cognition and Behavior, Nijmegen, The Netherlands

a r t i c l e i n f o a b s t r a c t

Article history: We report on a consanguineous Pakistani family with a severe congenital microcephaly syndrome resembling
Accepted 8 January 2014 the Seckel syndrome and Jawad syndrome. The affected individuals in this family were born to consanguineous
Available online xxx parents of whom the mother presented with mild intellectual disability (ID), epilepsy and diabetes mellitus.
The two living affected brothers presented with microcephaly, white matter disease of the brain, hyponychia,
Keywords:
dysmorphic facial features with synophrys, epilepsy, diabetes mellitus and ID. Genotyping with a 250 K SNP
Human congenital microcephaly syndrome
Seckel syndrome
array in both affected brothers revealed an 18 MB homozygous region on chromosome 18p11.21-q12.1
Jawad syndrome encompassing the SCKL2 locus of the Seckel and Jawad syndromes. Sequencing of the RBBP8 gene, underlying
RBBP8 the Seckel and Jawad syndromes, identied the novel mutation c.919A N G, p.Arg307Gly, segregating in a reces-
Psychiatric disorders sive manner in the family. In addition, in the two affected brothers and their mother we have also found a het-
NRXN1 erozygous 607 kb deletion, encompassing exons 1319 of NRXN1. Bidirectional sequencing of the coding exons
Intellectual disability of NRXN1 did not reveal any other mutation on the other allele. It thus appears that the phenotype of the mildly
affected mother can be explained by the NRXN1 deletion, whereas the more severe and complex microcephalic
phenotype of the two affected brothers is due to the simultaneous deletion in NRXN1 and the homozygous
missense mutation affecting RBBP8.
2014 Elsevier B.V. All rights reserved.

1. Introduction
The etiology of intellectual disability (ID) is highly heterogeneous.
Mutations in over 450 genes have been implicated in ID, which only rep-
resents a minor part of all ID genes that have been predicted (Inlow and
Restifo, 2004; Schuurs-Hoeijmakers et al., 2011; van Bokhoven, 2011).
The sheer number of ID genes presents a challenge for identication of
Abbreviations: ATR, ataxiatelangiectasia and RAD3-related; BMI, Body mass index;
CBC, complete blood count; CNV, copy number variation; DDR, DNA-damage response;
DSB, Double-strand break; EGF, epidermal growth factor; ID, intellectual disability; Mb,
megabase; MIM, Mendelian Inheritance in Man; MRI, magnetic resonance imaging;
MRN, MRE11-RAD50-NBS1; NRXN1, neurexin 1; OFC, occipitalfrontal head circumfer-
ence; qPCR, quantitative polymerase chain reaction; RBBP8, retinoblastoma-binding
protein 8; SCKL, Seckel syndrome; SD, standard deviation; SNP, single nucleotide polymor-
phism; WMD, white matter disease.
Corresponding author at: Department of Human Genetics 855, Nijmegen Centre for
Molecular Life Sciences, Radboud University Medical Center, P.O. Box 9101, 6500 HB
Nijmegen, The Netherlands. Tel.: +31 24 3616696; fax: +31 24 3668752.
E-mail address: Hans.vanbokhoven@radboudumc.nl (H. van Bokhoven).
1
The rst two authors have contributed equally to this work.
the primary genetic defect in individual families and isolated cases. Also
in their clinical presentation, ID disorders show extensive phenotypic
variability. A plethora of associated features can be seen in most individ-
uals presenting with ID. Facial dysmorphism and neurological features,
such as autism, epilepsy, attention decit hyperactivity disorder and be-
havioral anomalies are particularly common. These associated features
can be highly characteristic for a specic syndrome caused by mutations
in a particular gene. However, a large degree of clinical variability is often
seen for mutations in a specic gene, which can hamper clinical diagnosis
in isolated patients and small families.
Microcephaly is seen in a variety of syndromes comprising ID. The
head circumference in microcephaly is reduced three standard devia-
tions below the age- and sex-related means and reects the reduction
in underlying brain volume (Woods et al., 2005). Microcephaly associat-
ed syndromes differ from each other on the basis of other clinical
manifestations (Woods et al., 1992, 2005). The Seckel syndrome (MIM
210600), Filippi syndrome (MIM 272440) and Jawad syndrome (MIM
251255) are three separate syndromes in which microcephaly, ID,
short stature and digital malformations are common hallmarks.

0378-1119/$ see front matter 2014 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.gene.2014.01.027

Please cite this article as: Agha, Z., et al., A complex microcephaly syndrome in a Pakistani family associated with a novel missense mutation in
RBBP8 and a heterozygous deletion in NRXN1, Gene (2014), http://dx.doi.org/10.1016/j.gene.2014.01.027
2 Z. Agha et al. / Gene xxx (2014) xxxxxx

Qvist et al. recently reported RBBP8 (MIM 604124), as the causative
gene for both the Jawad syndrome and Seckel syndrome (Qvist et al.,
2011). This gene is located on chromosome 18q11.2 and encodes the
ubiquitously expressed retinoblastoma-binding protein 8, a nuclear
protein that is involved in the regulation of cell proliferation. It is a con-
served DNA repair protein that facilitates DNA end re-sectioning in the
double-strand break (DSB) repair pathway (D'Andrea, 2010).
Another important ID associated gene is NRXN1 (neurexin 1, MIM
600565), which is one of the largest human genes (1.1 Mb). It codes
for a neuronal cell adhesion molecule, and is located on chromosome
2p16.3 (Boucard et al., 2005). Recent data suggest that heterozygous
mutations in NRXN1 represent a susceptibility factor for a broad spec-
trum of neurological and neuropsychological disorders for example,
schizophrenia (Ching et al., 2010; Kirov et al., 2008; Rujescu et al.,
2009), ID disorders (McIntosh et al., 2006; Zahir et al., 2008; Zweier
et al., 2009), autism, (Gregor et al., 2011) and various other neuropsy-
chiatric disorders (Ching et al., 2010). A number of studies reveal reces-
sive genetic defects in NRXN1 associated with severe ID and dysmorphic
features resembling the PittHopkins like syndrome in some cases
(Duong et al., 2012; Harrison et al., 2011; Zweier et al., 2009).
In the current study, we have identied a homozygous mutation in
RBBP8, which co-segregates with microcephaly-associated ID syndrome
in the Pakistani family (MRQ12). In addition, we have also identied a
heterozygous deletion encompassing the NRXN1 in this family, which
is present in two affected sibs with a complex phenotype, as well as
the mother with a mild phenotype.
2. Methods
2.1. Clinical analysis
The proband of MRQ12 was ascertained at the National Training
Centre for Special Persons, Islamabad, Pakistan, where he was
referred for evaluation of ID and microcephaly. Informed written
consent was obtained from participants to this study or their
guardians prior to blood sampling. The study has been approved by
the Ethics Review Committee of COMSATS Institute of Information
Technology, Islamabad, Pakistan and Radboud University, Nijmegen,
The Netherlands.
Family MRQ12 is a consanguineous Pakistani family (Fig. 1) from the
northeastern region of Pakistan. At the age of 72 years, the mother pre-
sented with mild learning disabilities and epilepsy. She also suffered
from insulin dependent diabetes mellitus since the age of 22 years.
The father had normal intellectual functioning throughout his life. At
the age of 76 years he had symptoms of Parkinson's disease but with
no family history of the disease. The mother gave birth to six children,
and one pregnancy was spontaneously terminated at four months due
to growth arrest of the male fetus. Four of the six children were intellec-
tually normal without any congenital abnormality. One of her sons
(IV:2: Fig. 1) died at the age of 18 years from leukemia but he did not
have ID nor any psychiatric problems. The two youngest sons (IV:5
and IV:6; Fig. 1) suffered from microcephaly-associated ID syndrome.
The patient IV:5 (proband: Fig. 2a) was born after 40 weeks of ges-
tation of an uneventful pregnancy. Microcephaly was noted at birth
but detailed clinical data of birth weight and height of the proband
and the other affected brother were not available. The proband started
walking at the age of 11 months and speaking at the age of four years.
At three months of age his intellectual weakness was noted due to de-
layed milestones, at age 10 years he had symptoms of progressive
hyponychia (Fig. 2b). Upon clinical evaluation at the age of 40 years
the proband (Fig. 2a) had relatively short stature and obesity (height
162 cm, weight 76 kg) while his BMI was 27.8 (average BMI for
Pakistani adults is 18.8 3.6). He had symmetrical microcephaly with
a head circumference of 46 cm b 2 SD for occipitalfrontal head
circumference (OFC). He also had dysmorphic facial features with
synophrys, a prominent irregular shaped long nose and a short philtrum.
He also had deep set eyes and bilateral convergent strabismus. His ears
were rotated posteriorly and he had a highly pitched and shrill voice.
On his ngers and toes complete hyponychia was observed, and he
suffered from hyperhidrosis and could not count or write. His gait was

Fig. 1. Pedigree of MRQ12. The lled squares are affected males and the unlled circles and squares are unaffected females and males, respectively. The circle lled in gray indicates the
different and mild phenotype. The small sized square with diagonal line represents prenatal death of the fetus. Symbols with a diagonal line represent deceased individuals. The arrow
indicates the proband. The symbols R+/M, RM/M represent the RBBP8 unaffected carriers and homozygous mutants, respectively. Ndel/+ and N+/+reect the NRXN1 heterozygous deletion
and normal copy number of NRXN1, respectively.

Please cite this article as: Agha, Z., et al., A complex microcephaly syndrome in a Pakistani family associated with a novel missense mutation in
RBBP8 and a heterozygous deletion in NRXN1, Gene (2014), http://dx.doi.org/10.1016/j.gene.2014.01.027
 Z. Agha et al. / Gene xxx (2014) xxxxxx 3

Fig. 2. Photographs showing the clinical features of the patients. (a) Frontal picture of 40-year old proband. The gure shows the synophrys and clearly depicts the deeply set eyes of the
proband. The proband shows microcephaly, squinting of the eyes and a prominent nose on an elongated face. The ears of the patient are projected backward. (b) Picture of the hand of the
40-year old proband showing complete hyponychia of the nger nails. (c) Frontal picture of the 39-year old brother of the proband.

normal and he could visualize things normally. Speech was normal, but
his comprehension was poor. Cardiologic evaluation, metabolic screens,
complete blood count (CBC) and liver function tests were all normal,
while his brain MRI revealed accentuated white matter of the cerebrum
and the cerebellum (Fig. 3), suggestive of an underlying white matter
disease (WMD) of the brain. Otherwise the ventricle and cortical sulci
were normal with no focal mass lesion or midline shift and no abnormal
collection or hemorrhage.
Patient IV:6, the proband's 39 years old affected brother (Fig. 2c),
had a similar phenotype as his brother, including obesity and short
stature (height 156 cm, and weight 69 kg) while his BMI was 25.1 (aver-
age BMI at adult age for Pakistani population is 18.8 3.6). He had mi-
crocephaly with a head circumference of 47 cm b 2 SD OFC. He had
synophrys and his eyes were deeply set but he did not have a squint.
His nose was large, prominent and irregularly shaped, while his ears
were rotated posteriorly. His voice was high pitched and shrill, he had
complete hyponychia of ngers and toes, had hyperhidrosis since an
early age and also had seizures. His EEG was not recorded hence the ep-
ileptic seizures cannot be diagnosed. Intellectually he was weaker than
his brother, he could speak only a few words and he was very aggressive.
2.2. Genetic analysis
Genomic DNA was extracted from the peripheral blood of the partic-
ipating family members (III:1, III:2, IV:1, IV:5 and IV:6), using a standard
phenolchloroform extraction method (Sambrook and Russell, 2006).
Genotyping of the affected members (IV:5 and IV:6), their parents
(III:1 and III:2) and a healthy sib (IV:1) of the family MRQ12 was per-
formed using Affymetrix 250 K single nucleotide polymorphisms
(SNP) microarray (Affymetrix Inc. Santa Clara, CA) in order to obtain
the copy number variation (CNV) data while the HomozygosityMapper
(http://www.homozygositymapper.org/) was used to nd common
homozygous regions among affected family members. The 250 K data
also revealed heterozygous deletion in the affected members of MRQ12.
Genomic real-time qPCR analysis was performed for family mem-
bers III:1, III:2, IV:1, IV:5 and IV:6 on a 7500 Fast Real-Time PCR System
(Applied Biosystems, Foster City, CA, US) by using Power SYBR Green
PCR Master Mix (Applied Biosystems, Foster City, CA, US) according to
the manufacturer's instructions. The melting curves of all PCR products
showed a single PCR product, while the controls were negative. Copy
numbers were measured relative to the reference gene cystic brosis
transmembrane conductance regulator (CFTR).
The coding exons (219) of RBBP8, and (224) of NRXN1 of the
proband and family members were screened by bidirectional sequencing
using ABI-PRISM 3730 (Life Technologies, Foster City, CA) as described
previously (Gregor et al., 2011).
3. Results
Genotyping of III:1, III:2, IV:1, IV:5 and IV:6 family members of
MRQ12 by 250 K SNP array revealed four homozygous regions i.e. Chr.6
(3.15 Mb); Chr.7 (5.51 Mb); Chr.10 (3.63 Mb) and Chr.18 (17.71 Mb) in
two affected IV:5 and IV:6 while these regions were not homozygous in
their healthy sister IV:1 nor in the parents III:1 and III:2. The other living
healthy siblings (IV:4 and IV:7) were unavailable for testing. The region at
chromosome 18p21.2q12.2 with anking SNPs rs8060649 and
rs16954273 contained the RBBP8, which has been previously reported
to be mutated in both the Seckel and Jawad syndromes (Qvist et al.,
2011). In order to screen for a mutation in the patients of family
MRQ12, we sequenced all the coding exons of RBBP8 and found a homo-
zygous missense change (c.919A N G, p.Arg307Gly) in exon 11 (Fig. 4).
The substitution co-segregated with the phenotype in a recessive manner

Fig. 3. Magnetic resonance imaging (MRI) of the family members. (a) Normal brain, (b, c) Turbo FLAIR region of the brain of MRQ12's proband showing diffuse white matter. The red ar-
rows in the gures are pointing toward the area of the brain where abnormal white matter deposition can be seen. (For interpretation of the references to color in this gure legend, the
reader is referred to the web version of this article.)

Please cite this article as: Agha, Z., et al., A complex microcephaly syndrome in a Pakistani family associated with a novel missense mutation in
RBBP8 and a heterozygous deletion in NRXN1, Gene (2014), http://dx.doi.org/10.1016/j.gene.2014.01.027
4 Z. Agha et al. / Gene xxx (2014) xxxxxx
Fig. 4. Sequencing chromatograms. The upper chromatogram represents the heterozygous
change c.919A N G in the unaffected father while the lower chromatogram shows the ho-
mozygous change c.919A N G in the affected proband.
(Fig. 1) in the two sons (IV:5 and IV:6) with the severe phenotype, while
the mother (III:2), father (III:1) and healthy daughter (IV:1) were hetero-
zygous for the substitution. This RBBP8 variant was not seen in N13,000
alleles in the Exome Variant Server database (http://evs.gs.washington.
edu/EVS/) [August 2013].
The 250 K array CNV data also revealed a 610 kb deletion in
2p16.3 (Chr2:50.2150.82 Mb UCSC Human Genome Browser build
19; Fig. 5), resulting in a heterozygous deletion of exon 13 to exon 19
of NRXN1, which was conrmed by qPCR. The deletion was inherited
by the severely affected sons (IV:5 and IV:6) in a dominant manner
from the mother (III:2), who was suffering from mild ID and epilepsy,
while both the father (III:1) and daughter (IV:1) had normal copy num-
ber of NRXN1. No mutation on the other allele of NRXN1 was detected
from the paternal chromosome.
Please cite this article as: Agha, Z., et al., A complex microcephaly syndrome in a Pakistani family associated with a novel missense mutation in
RBBP8 and a heterozygous deletion in NRXN1, Gene (2014), http://dx.doi.org/10.1016/j.gene.2014.01.027
4. Discussion
RBBP8 point mutations have previously been reported in the Seckel
syndrome and Jawad syndrome (Qvist et al., 2011). The Seckel syn-
drome belongs to the group of genome instability disorders, collectively
referred to as DNA-damage response (DDR) and repair defective
syndromes (O'Driscoll et al., 2003). While, cancer predisposition is
often associated with such syndromes, only a few cancers have been re-
ported for the Seckel syndrome patients. Instead, the Seckel syndrome
pathogenesis is primarily based on marked growth and neurological
impairments (O'Driscoll et al., 2003). The Jawad syndrome is another
autosomal recessive congenital microcephaly syndrome that shares
the SCKL2 locus with the Seckel syndrome (Hassan et al., 2008).
The Seckel syndrome also known as dwarsm syndrome has various
dysmorphic facial features such as bird shaped appearance that is a
diagnostic feature of this syndrome. The syndrome is usually character-
ized by intrauterine growth retardation and postnatal microcephaly
and dwarsm associated with ID (Borglum et al., 2001; Filippi, 1985;
Goodship et al., 2000; Kilinc et al., 2003; Sharif and Donnai, 2004),
while the Jawad syndrome is characterized by microcephaly, growth
retardation, caf au lait white pigmentation on the skin, congenital
anonychia and digital malformation (Hassan et al., 2008).
For the Seckel syndrome ve different forms have been identied,
i.e. SCKL1, characterized by a mutation that creates an alternative splice
site in the ATR (ataxia telangiectasia and Rad3 related; MIM 601215)
(Borglum et al., 2001), SCKL2 (MIM 606744) mapping to 18p11.31
q11.2 (O'Driscoll et al., 2003), and SCKL3 (MIM 608664), mapped
to 14q23q24 (Goodship et al., 2000). SCKL4 (MIM 613676) maps to
13q12.2 and is caused by recessive mutations in the CENPJ (centromeric
protein J, MIM 609279) (Grifth et al., 2008), and SCKL5 (MIM 613823)
is caused by mutations in the CEP152 (centrosomal protein 152-KD,
MIM 613529), which maps to 15q21 (Kalay et al., 2011).
The affected males in family MRQ12 shared the bird shaped appear-
ance of the Seckel syndrome and microcephaly with both the Jawad and
Seckel syndromes, while congenital anonychia is shared by the Jawad
syndrome and the MRQ12 family. ID and microcephaly are common
features shared between the Seckel syndrome, Jawad syndrome and
the present family (Table 1). Microcephaly is thought to result from
reduced proliferative potential in the developing nervous system,
most likely due to increased cell death of the neuronal stem cells or pro-
genitor cells in the rapidly expanding fetal brain (O'Driscoll and Jeggo,
2008).
RBBP8 has been observed to work in DSB repair processing as licens-
ing of DNA-end resection that requires cell-cycle dependent phosphor-
ylation of RBBP8, and in the absence of this protein, DSB processing is
impaired and the activation of ATR, which is a DNA damage responsive
protein, is blocked (Qvist et al., 2011). Based on these observations,
RBBP8 was screened by Qvist et al. in two families, one with the Seckel
syndrome and the other with the Jawad syndrome. Both were previous-
ly linked to the SCKL2 locus at 18p11.31q11.2 (Qvist et al., 2011). These
analyses revealed two independent homozygous mutations in RBBP8, an
intronic mutation in the Seckel syndrome leading to an alternatively
spliced transcript and the appearance of a premature stop codon
predicting a loss-of-function allele due to nonsense-mediated RNA
decay and/or the generation of a C-terminally truncated form of RBBP8.
While in the Jawad syndrome there was a two base pair deletion in
exon 11, causing a frame-shift and the appearance of a premature stop
codon, either way, a complete loss of RBBP8 activity would be predicted
affecting DSB resection and ATR activation (Qvist et al., 2011). The ATR
protein orchestrates cellular responses to DNA damage and replication
stress. Complete loss of ATR function thus leads to chromosomal instabil-
ity and cell death (Fang et al., 2004).
In our study, we have identied a novel homozygous mutation in the
RBBP8 gene(c.919ANG) that predicts an amino acid substitution
p.Arg307Gly. The arginine at this position is highly conserved. The
change of a large positively charged arginine by a small and highly
 Z. Agha et al. / Gene xxx (2014) xxxxxx 5

Fig. 5. NRXN1 deletion region of family MRQ12. (a) Original data of CNV analysis derived from 250 k Affymetrix SNP array of proband, showing heterozygous deletion in the indicated
region (chr2:502, 10508, 17 Mb UCSC Human Genome Browser version 19). (b) Figure showing the NRXN1 exons from 124 with its two isoforms, alpha-isoform and beta-isoform,
it represents the 607 kb microdeletion in family MRQ12 covering exons 13 to 19 coding for both isoforms i.e. part of alpha and promoter of beta.

exible glycine residue is signicant and generally predicts the creation
of instability due to the increased exibility. Arg307 is not located in a
known functional domain and there is no crystal structure available
for RBBP protein family members to make further predictions about
the functional consequences of this mutation.
Inactivating mutations of RBBP8 are expected to impair the capacity
of cells to respond optimally to endogenously arising DNA damage. This
would lower the apoptotic threshold of the cells and cause reduction in
their proliferative potential, which can cause retardation in the growth
of the patient (Qvist et al., 2011). Our results are in agreement with
the ndings of Qvist et al. (2011) as our mutation is also located in the
C-terminal part of the RBBP8 protein, which could result in the produc-
tion of an abnormal protein. Since RBBP8 acts together with the MRN
(MRE11-RAD50-NBS1) complex to promote DNA end resection and
the generation of single-stranded DNA, this function could be impaired,
and this is critically important for the homologous recombination repair
(Yuan and Chen, 2009). Based on these factors we also propose that the
mutation we have identied in MRQ12 is responsible for the severe
phenotype of the two sibs as this region bears an MRN interaction
domain (Sartori et al., 2007) that is crucial for DNA-end resection.
Although a second MRN interaction point has been found in the
N-terminal part of the protein (Yuan and Chen, 2009), the C-terminal
region is essential for RBBP8-mediated activation of MRN-associated
nuclease activity. Therefore it is probable that this mutation can render
the MRN-RBBP8 complex ineffective and cause the impaired production
of single stranded DNA. This results in the inactivation of ATR that ulti-
mately cause defective apoptosis activity in patients due to hypersensi-
tivity to DNA damage.
In addition to the RBBP8 mutation we identied an intragenic 610 kb
deletion comprising exons 1319 of NRXN1, including a promoter
which is used to generate -neurexin of NRXN1. A number of studies
have shown that neurexins have an essential role in the development
of synapse, not in its initial adhesion but in the recruitment of molecular
components and its maturation (Zhang et al., 2005). There are three
neurexin genes in mammals, which have the capacity to generate a
stunning variety of distinct transcripts by using alternate promoters,
splice sites and exons (Rowen et al., 2002; Tabuchi and Sudhof, 2002).
For NRXN1, most transcripts use the upstream promoter and encode
alpha-neurexin isoforms; fewer transcripts are produced from the
downstream promoter yielding the beta-neurexin isoforms. Alpha-
neurexins contain epidermal growth factor-like (EGF-like) sequences
and laminin G domains, and they interact with neurexophilins. Beta-
neurexins lack EGF-like sequences and contain fewer laminin G do-
mains than alpha-neurexins (Kirov et al., 2008). Interestingly, for each
of the three Neurexin genes, knockout of the -neurexins, leaving the
-neurexins intact, leads to perinatal lethality due to the loss of presyn-
aptic Ca2+ channel function (Missler et al., 2003). Different studies have
shown that -neurexins have a unique function that is not provided

Table 1
Comparison of clinical and morphometric ndings in the Seckel (SCKL2), Jawad and MRQ12 patients.

Seckel (SCKL2) Jawad MRQ12

Appearance of symptoms Infancy Infancy Infancy

Birth weight (kg)
Height (SD)
Head circumference (SD)
Facial characteristics
Global developmental delay
Skin abnormalities
Digital malformation
Voice
Low (1 to 1.5) Data not available Data not available
Reduced (3.5 to 5.5) Normal Reduced
Reduced (4.7 to 5.0) Reduced (5.0 to 7.0) Reduced
Narrow, not receding, forehead and Sharply receding foreheads, prominent Narrow, bird shaped, deeply set eyes,
prominent noses, small craniums noses, small craniums squint, prominent nose, hypertelorism,
low ears
Mild Moderate to severe Moderatesevere
Caf au lait spots Caf au lait-like spots of white appearance Hyperhydrosis
Phalangeal joint swellings, clinodactyly Phalangeal joint swellings, clinodactyly, Anonychia congenita
polydactyly, syndactyly, total absence of nails
Normal Normal Shrilled, high pitched

Please cite this article as: Agha, Z., et al., A complex microcephaly syndrome in a Pakistani family associated with a novel missense mutation in
RBBP8 and a heterozygous deletion in NRXN1, Gene (2014), http://dx.doi.org/10.1016/j.gene.2014.01.027
6 Z. Agha et al. / Gene xxx (2014) xxxxxx
by -neurexins. Moreover, a patient with mild ID and autistic features
reported by Zahir et al. (2008) had a heterozygous deletion that specif-
ically affected NRXN1-, leaving NRXN1- intact. The heterozygous loss
of exon 13 to exon 19 of -neurexin, as well as deletion of the promoter
for -neurexin is likely to be causative for the mild ID of the mother. The
NRXN1 deletion may also explain her epilepsy, since heterozygous
exon-disrupting deletions of NRXN1 have been shown to represent a ge-
netic risk factor common in idiopathic generalized epilepsy (Moller
et al., 2013). Moreover, epilepsy is seen in 43% of NRXN1 deletion
carriers (Bena et al., 2013). From the current work the etiology of diabe-
tes mellitus in the mother is not clear. Interestingly in a recent study a
family was reported in which a heterozygous deletion comprising the
upstream promoter and rst intron of NRXN1 was seen in a patient
with diabetes mellitus (Duong et al., 2012). In addition, it is of note
that the neurexins are expressed in the -cells of the pancreas, therefore
any aberrations in the protein could potentially lead to a decrease in
function of the pancreas (Suckow et al., 2008). Together with our obser-
vations, these data warrant further investigation of the involvement of
NRXN1 in diabetes. Finally, it has been shown that a NRXN1 polymor-
phism was associated with white matter abnormalities in autism
and schizophrenia (Voineskos et al., 2011), suggesting that the NRXN1
deletion might also contribute to the white matter anomalies that
were observed in the two affected males.
5. Conclusion
In conclusion, we propose that the severe Seckel/Jawad syndrome-
like phenotype of the two brothers in MRQ12 is attributed to the homo-
zygous RBBP8 missense mutation combined with a heterozygous exonic
deletion of NRXN1. Their mother is comparatively less affected and has
mild learning disability and epilepsy that can be explained by a NRXN1
deletion. This deletion could be a contributing factor to the mild ID and
diabetes mellitus of the mother. Clinical variability is typical for heterozy-
gous NRXN1 deletions, suggesting modulation by other genetic factors. It
is also conceivable that modier loci are linked to genes such as RBBP8
that have a role in neurodevelopment.
Conict of interest
The authors state that they have no conict of interest.
Acknowledgments
We are grateful to all the family members for their participation in
this study. We thank Dr. Hanka Venselaar for helpful comments about
the predicted effect of the amino acid change. This study was funded by
the European Union's Seventh Framework Program under grant agree-
ment number 241995, project GENCODYS and grant agreement number
223143, project TECHGENE. Zehra Agha was supported by the IRSIP pro-
gram of the Higher Education Commission (HEC), Islamabad, Pakistan. CZ
was funded by a grant from the Deutsche Forschungsgemeinschaft
(Zw184/1-1). This study was also supported by funds to RQ from the
HEC National Research Program for Universities under grant no 2155. ZI
was supported by HEC.
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