Naseebullah Kakar

Loss-of-function mutations of ILDR1 cause autosomal-recessive hearing impairment DFNB42.

By using homozygosity mapping in a consanguineous Pakistani family, we detected linkage of nonsyndromic hearing loss to a 7.6 Mb region on chromosome 3q13.31-q21.1 within the previously reported DFNB42 locus. Subsequent candidate gene sequencing identified a homozygous nonsense mutation (c.1135G>T [p.Glu379X]) in ILDR1 as the cause of hearing impairment. By analyzing additional consanguineous families with homozygosity at this locus, we detected ILDR1 mutations in the affected individuals of 10 more families from Pakistan and Iran. The identified ILDR1 variants include missense, nonsense, frameshift, and splice-site mutations as well as a start codon mutation in the family that originally defined the DFNB42 locus. ILDR1 encodes the evolutionarily conserved immunoglobulin-like domain containing receptor 1, a putative transmembrane receptor of unknown function. In situ hybridization detected expression of Ildr1, the murine ortholog, early in development in the vestibule and in hair cells and supporting cells of the cochlea. Expression in hair cell- and supporting cell-containing neurosensory organs is conserved in the zebrafish, in which the ildr1 ortholog is prominently expressed in the developing ear and neuromasts of the lateral line. These data identify loss-of-function mutations of ILDR1, a gene with a conserved expression pattern pointing to a conserved function in hearing in vertebrates, as underlying nonsyndromic prelingual sensorineural hearing impairment.

Dopamine transporter deficiency syndrome: phenotypic spectrum from infancy to adulthood.

Dopamine transporter deficiency syndrome is an SLC6A3-related progressive infantile-onset parkinsonism-dystonia that mimics cerebral palsy. Ng et al. describe clinical features and molecular findings in a new cohort of patients. They report infants with classical disease, as well as young adults manifesting as atypical juvenile-onset parkinsonism-dystonia, thereby expanding the disease spectrum.

A novel HSF4 gene mutation (p.R405X) causing autosomal recessive congenital cataracts in a large consanguineous family from Pakistan.

BackgroundHereditary cataracts are most frequently inherited as autosomal dominant traits, but can also be inherited in an autosomal recessive or X-linked fashion. To date, 12 loci for autosomal recessive cataracts have been mapped including a locus on chromosome 16q22 containing the disease-causing gene HSF4 (Genbank accession number NM_001040667). Here, we describe a family from Pakistan with the first nonsense mutation in HSF4 thus expanding the mutational spectrum of this heat shock transcription factor gene.MethodsA large consanguineous Pakistani family with autosomal recessive cataracts was collected from Quetta. Genetic linkage analysis was performed for the common known autosomal recessive cataracts loci and linkage to a locus containing HSF4 (OMIM 602438) was found. All exons and adjacent splice sites of the heat shock transcription factor 4 gene (HSF4) were sequenced. A mutation-specific restriction enzyme digest (HphI) was performed for all family members and unrelated controls.ResultsThe disease phenotype perfectly co-segregated with markers flanking the known cataract gene HSF4, whereas other autosomal recessive loci were excluded. A maximum two-point LOD score with a Zmax = 5.6 at θ = 0 was obtained for D16S421. Direct sequencing of HSF4 revealed the nucleotide exchange c.1213C > T in this family predicting an arginine to stop codon exchange (p.R405X).ConclusionWe identified the first nonsense mutation (p.R405X) in exon 11 of HSF4 in a large consanguineous Pakistani family with autosomal recessive cataract.

Biallelic SZT2 Mutations Cause Infantile Encephalopathy with Epilepsy and Dysmorphic Corpus Callosum

Epileptic encephalopathies are genetically heterogeneous severe disorders in which epileptic activity contributes to neurological deterioration. We studied two unrelated children presenting with a distinctive early-onset epileptic encephalopathy characterized by refractory epilepsy and absent developmental milestones, as well as thick and short corpus callosum and persistent cavum septum pellucidum on brain MRI. Using whole-exome sequencing, we identified biallelic mutations in seizure threshold 2 (SZT2) in both affected children. The causative mutations include a homozygous nonsense mutation and a nonsense mutation together with an exonic splice-site mutation in a compound-heterozygous state. The latter mutation leads to exon skipping and premature termination of translation, as shown by RT-PCR in blood RNA of the affected boy. Thus, all three mutations are predicted to result in nonsense-mediated mRNA decay and/or premature protein truncation and thereby loss of SZT2 function. Although the molecular role of the peroxisomal protein SZT2 in neuronal excitability and brain development remains to be defined, Szt2 has been shown to influence seizure threshold and epileptogenesis in mice, consistent with our findings in humans. We conclude that mutations in SZT2 cause a severe type of autosomal-recessive infantile encephalopathy with intractable seizures and distinct neuroradiological anomalies.

De novo mutations of the gene encoding the histone acetyltransferase KAT6B in two patients with Say-Barber/Biesecker/Young-Simpson syndrome

The Say‐Barber/Biesecker/Young‐Simpson (SBBYS) type of the blepharophimosis–mental retardation syndrome group (Ohdo‐like syndromes) is a multiple congenital malformation syndrome characterized by vertical narrowing and shortening of the palpebral fissures, ptosis, intellectual disability, hypothyroidism, hearing impairment, and dental anomalies. Mutations of the gene encoding the histone‐acetyltransferase KAT6B have been recently identified in individuals affected by SBBYS syndrome. SBBYS syndrome‐causing KAT6B mutations cluster in a ∼1,700 basepair region in the 3′ part of the large exon 18, while mutations located in the 5′ region of the same exon have recently been identified to cause the genitopatellar syndrome (GPS), a clinically distinct although partially overlapping malformation‐intellectual disability syndrome. Here, we present two children with clinical features of SBBYS syndrome and de novo truncating KAT6B mutations, including a boy who was diagnosed at the age of 4 months. Our results confirm the implication of KAT6B mutations in typical SBBYS syndrome and emphasize the importance of genotype–phenotype correlations at the KAT6B locus where mutations truncating the KAT6B protein at the amino‐acid positions ∼1,350–1,920 cause SBBYS syndrome.

A homozygous splice site mutation in TRAPPC9 causes intellectual disability and microcephaly

Autosomal recessive intellectual disability is believed to be particularly prevalent in highly consanguineous populations and genetic isolates and may account for a quarter of all non-syndromic cases. Mutations in more than 50 genes have been reported to be involved in autosomal recessive intellectual disability, including TRAPPC9 (MIM 611966), mutations of which have been identified in six families from different geographical origins. We performed a clinical and molecular genetic study of a consanguineous Pakistani family segregating intellectual disability and microcephaly. SNP-array-based homozygosity mapping revealed suggestive linkage to four genomic regions including one on chromosome 8 that contained TRAPPC9. We detected a homozygous TRAPPC9 splice donor site mutation (c.1024+1G>T) that cosegregated with intellectual disability in the family and led to skipping of exon 3 and exons 3 and 4 in blood-derived patient RNA. We have thus identified a novel splice site mutation leading to exon skipping and premature termination of TRAPPC9 translation. These data further suggest that TRAPPC9 mutations -unlike mutations in the vast majority of the known intellectual disability-associated genes- constitute a more frequent cause of autosomal-recessive cognitive deficits, especially when microcephaly is also present.

Exome sequencing and CRISPR/Cas genome editing identify mutations of ZAK as a cause of limb defects in humans and mice

The CRISPR/Cas technology enables targeted genome editing and the rapid generation of transgenic animal models for the study of human genetic disorders. Here we describe an autosomal recessive human disease in two unrelated families characterized by a split-foot defect, nail abnormalities of the hands, and hearing loss, due to mutations disrupting the SAM domain of the protein kinase ZAK. ZAK is a member of the MAPKKK family with no known role in limb development. We show that Zak is expressed in the developing limbs and that a CRISPR/Cas-mediated knockout of the two Zak isoforms is embryonically lethal in mice. In contrast, a deletion of the SAM domain induces a complex hindlimb defect associated with down-regulation of Trp63, a known split-hand/split-foot malformation disease gene. Our results identify ZAK as a key player in mammalian limb patterning and demonstrate the rapid utility of CRISPR/Cas genome editing to assign causality to human mutations in the mouse in <10 wk.

STIL mutation causes autosomal recessive microcephalic lobar holoprosencephaly

Holoprosencephaly is a clinically and genetically heterogeneous midline brain malformation associated with neurologic manifestations including developmental delay, intellectual disability and seizures. Although mutations in the sonic hedgehog gene SHH and more than 10 other genes are known to cause holoprosencephaly, many patients remain without a molecular diagnosis. Here we show that a homozygous truncating mutation of STIL not only causes severe autosomal recessive microcephaly, but also lobar holoprosencephaly in an extended consanguineous Pakistani family. STIL mutations have previously been linked to centrosomal defects in primary microcephaly at the MCPH7 locus. Our results thus expand the clinical phenotypes associated with biallellic STIL mutations to include holoprosencephaly.

Homozygous missense and nonsense mutations in BMPR1B cause acromesomelic chondrodysplasia-type Grebe

Acromesomelic chondrodysplasias (ACDs) are characterized by disproportionate shortening of the appendicular skeleton, predominantly affecting the middle (forearms and forelegs) and distal segments (hands and feet). Here, we present two consanguineous families with missense (c.157T>C, p.(C53R)) or nonsense (c.657G>A, p.(W219*)) mutations in BMPR1B. Homozygous affected individuals show clinical and radiographic findings consistent with ACD-type Grebe. Functional analysis of the missense mutation C53R revealed that the mutated receptor was partially located at the cell membrane. In contrast to the wild-type receptor, C53R mutation hindered the activation of the receptor by its ligand GDF5, as shown by reporter gene assay. Further, overexpression of the C53R mutation in an in vitro chondrogenesis assay showed no effect on cell differentiation, indicating a loss of function. The nonsense mutation (c.657G>A, p.(W219*)) introduces a premature stop codon, which is predicted to be subject to nonsense-mediated mRNA decay, causing reduced protein translation of the mutant allele. A loss-of-function effect of both mutations causing recessive ACD-type Grebe is further supported by the mild brachydactyly or even non-penetrance of these mutations observed in the heterozygous parents. In contrast, dominant-negative BMPR1B mutations described previously are associated with autosomal-dominant brachydactyly-type A2.

Various Aspects, Patterns and Risk Factors in Breast Cancer Patients of Balochistan

PURPOSE Breast cancer is the commonest malignancy of females throughout the world with one million new cases each year. In Pakistan, the burden of breast cancer disease is high with late stage presentation being a common feature, more than half being stage III or stage IV. The objective of this study was to study various aspects, patterns and risk factors in breast cancer patients of Balochistan. METHOD Present study was performed on 134 patients of breast cancer who were registered in CENAR. The patients were interviewed by providing a questionnaire. Informed consent was taken from all the patients who took part in this study after explanation of the study aims. Body mass index (BMI) was calculated andbiopsy reports were obtained from patients files. All the cases were classified with respect to age, gender, ethnic group (Baloch, Pashtoon, Punjabi, Afghani, Hazara) BMI, cancer type, cancer grade, hormonal status, side of the cancer, fertility and marital status. RESULTS Out of 134 patients, the most common ethnic group was Pashtoon with a total of 42 and the common age group was 41-50 years with a total of 51. Invasive ductal carcinoma (IDC) was the most common type, accounting for in 128 patients (95.5%) followed by invasive lobular carcinoma (ILC). CONCLUSION Pashtoon was the most common ethnic group, IDC was common type and most of the patients had an ER/PR positive hormonal status.