The dental abnormalities are the typical features of many ectodermal dysplasias along with congenital malformations of nails, skin, hair, and sweat glands. However, several reports of non-syndromic/isolated tooth agenesis have also been found in the literature. The characteristic features of hypohidrotic ectodermal dysplasia (HED) comprise of hypodontia/oligodontia, along with hypohidrosis/anhidrosis, and hypotrichosis. Pathogenic variants in EDA, EDAR, EDARADD, and TRAF6, cause the phenotypic expression of HED. Genetic alterations in EDA and WNT10A cause particularly non-syndromic/isolated oligodontia. In the current project, we recruited 57 patients of 17 genetic pedigrees (A-Q) from different geographic regions of the world, including Pakistan, Egypt, Saudi Arabia, and Syria. The molecular investigation of different syndromic and non-syndromic dental conditions, including hypodontia, oligodontia, generalized odontodysplasia, and dental crowding was carried out by using exome and Sanger sequencing. We have identified a novel missense variant (c.311G>A; p.Arg104His) in WNT10A in three oligodontia patients of family A, two novel sequence variants (c.207delinsTT, p.Gly70Trpfs*25 and c.1300T>G; p.Try434Gly) in EDAR in three patients of family B and four patients of family C, respectively. To better understand the structural and functional consequences of missense variants in WNT10A and EDAR on the stability of the proteins, we have performed extensive molecular dynamic (MD) simulations. We have also identified three previously reported pathogenic variants (c.1076T>C; p.Met359Thr), (c.1133C>T; p.Thr378Met) and (c.594_595insC; Gly201Argfs*39) in EDA in family D (four patients), E (two patients) and F (one patient), correspondingly. Presently, our data explain the genetic cause of 18 syndromic and non-syndromic tooth agenesis patients in six autosomal recessive and X-linked pedigrees (A-F), which expand the mutational spectrum of these unique clinical manifestations.

Background: Polydactyly is a common genetic limb deformity characterized by the presence of extra fingers or toes. This anomaly may occur in isolation (nonsyndromic) or as part of a syndrome. The disease is broadly divided into preaxial polydactyly (PPD; duplication of thumb), mesoaxial polydactyly (complex polydactyly), and postaxial polydactyly (PAP: duplication of the fifth finger). The extra digits may be present in one or both the limbs. Heterozygous variants in the GLI3, ZRS/SHH, and PITX1 have been associated with autosomal dominant polydactyly, while homozygous variants in the ZNF141, IQCE, GLI1, and FAM92A have been associated with autosomal recessive polydactyly. Pathogenic mutations in the GLI3 gene (glioma‐associated oncogene family zinc finger 3) have been associated with both nonsyndromic and syndromic polydactyly.Methods: Here, we report an extended five generation kindred having 12 affected individuals exhibiting nonsyndromic postaxial polydactyly type A condition. Whole‐exome sequencing followed by variant prioritization, bioinformatic studies, Sanger validation, and segregation analysis was performed.Results: Using exome sequencing in the three affected individuals, we identified a novel heterozygous frameshift variant (c.3567_3568insG; p.Ala1190Glyfs*57) in the transcriptional activator (TA2) domain of the GLI3 encoding gene.Conclusion: To the best of our knowledge, the present study reports on the first familial case of nonsyndromic postaxial polydactyly due to the GLI3 variant in Pakistani population. Our study also demonstrated the important role of GLI3 in causing nonsyndromic postaxial polydactyly.

Charcot–Marie–Tooth disease (CMT) and autosomal recessive spastic ataxia of Charlevoix–Saguenay type (ARSACS) are large heterogeneous groups of sensory, neurological genetic disorders characterized by sensory neuropathies, muscular atrophies, abnormal sensory conduction velocities, and ataxia. CMT2EE (OMIM: 618400) is caused by mutations in MPV17 (OMIM: 137960), CMT4F (OMIM: 614895) is caused by PRX (OMIM: 605725), CMTX1 (OMIM: 302800) is caused by mutations in GJB1 (OMIM: 304040), and ARSACS (OMIM: 270550) is caused by mutations in SACS (OMIM: 604490). In this study, we enrolled four families: DG-01, BD-06, MR-01, and ICP-RD11, with 16 affected individuals, for clinical and molecular diagnoses. One patient from each family was analyzed for whole exome sequencing and Sanger sequencing was done for the rest of the family members. Affected individuals of families BD-06 and MR-01 show complete CMT phenotypes and family ICP-RD11 shows ARSACS type. Family DG-01 shows complete phenotypes for both CMT and ARSACS types. The affected individuals have walking difficulties, ataxia, distal limb weakness, axonal sensorimotor neuropathies, delayed motor development, pes cavus, and speech articulations with minor variations. The WES analysis in an indexed patient of family DG-01 identified two novel variants: c.83G>T (p.Gly28Val) in MPV17 and c.4934G>C (p.Arg1645Pro) in SACS. In family ICP-RD11, a recurrent mutation that causes ARSACS, c.262C>T (p.Arg88Ter) in SACS, was identified. Another novel variant, c.231C>A (p.Arg77Ter) in PRX, which causes CMT4F, was identified in family BD-06. In family MR-01, a hemizygous missense variant c.61G>C (p.Gly21Arg) in GJB1 was identified in the indexed patient. To the best of our knowledge, there are very few reports on MPV17, SACS, PRX, and GJB1 causing CMT and ARSACS phenotypes in the Pakistani population. Our study cohort suggests that whole exome sequencing can be a useful tool in diagnosing complex multigenic and phenotypically overlapping genetic disorders such as Charcot–Marie–Tooth disease (CMT) and spastic ataxia of Charlevoix–Saguenay type.

Background: Jalili syndrome (JS) is a rare cone‐rod dystrophy (CRD) associated with amelogenesis imperfecta (AI). The first clinical presentation of JS patients was published in 1988 by Jalili and Smith. Pathogenic mutations in the Cyclin and CBS Domain Divalent Metal Cation Transport Mediator 4 (CNNM4) magnesium transporter protein have been reported as the leading cause of this anomaly.Methods: In the present study, a clinical and genetic investigation was performed in a consanguineous family of Pakistani origin, showing characteristic features of JS. Sanger sequencing was successfully used to identify the causative variant in CNNM4. Molecular dynamics (MD) simulations were performed to study the effect of amino acid change over CNNM4 protein.Results: Sequence analysis of CNNM4 revealed a novel missense variant (c.1220G>T, p.Arg407Leu) in exon‐1 encoding cystathionine‐β‐synthase (CBS) domain. To comprehend the mutational consequences in the structure, the mutant p.Arg407Leu was modeled together with a previously reported variant (c.1484C>T, p.Thr495Ile) in the same domain. Additionally, docking analysis deciphered the binding mode of the adenosine triphosphate (ATP) cofactor. Furthermore, 60ns MD simulations were carried out on wild type (p.Arg407/p.Thr495) and mutants (p.Arg407Leu/p.Thr495Ile) to understand the structural and energetic changes in protein structure and its dynamic behavior. An evident conformational shift of ATP in the binding site was observed in simulated mutants disrupting the native ATP‐binding mode.Conclusion: The novel identified variant in CNNM4 is the first report from the Pakistani population. Overall, the study is valuable and may give a novel insight into metal transport in visual function and biomineralization.

Background: Dyggve-Melchior-Clausen syndrome (DMC) is a skeletal dysplasia with associated defects of brain development and intelligence. The truncating pathogenic variants in DYM are the most frequent cause of DMC. Smith-McCort (SMC), another skeletal dysplasia, is also caused by non-synonymous DYM variants. Methods and Results: In the current study, we examined a Pakistani consanguineous family with three affected members. Clinical features like spondyloepimetaphyseal dysplasia, indicative of characteristic skeletal abnormalities, and intellectual disability were observed. Our male patients had microcephaly and coarse facial features while the female patient did not represent microcephaly or abnormal facies, which are significant features of DMC patients. Sanger sequencing identified a novel homozygous frameshift insertion (c.95_96insT, p.W33Lfs^*14) in DYM, which likely leads to nonsense-mediated decay (NMD). Conclusion: The novel frameshift change verifies the fact that pathogenic variants in DYM are the most frequent cause of DMC.

Background: Amelogenesis imperfecta (AI) is a highly heterogeneous group of hereditary developmental abnormalities which mainly affects the dental enamel during tooth development in terms of its thickness, structure, and composition. It appears both in syndromic as well as non-syndromic forms. In the affected individuals, the enamel is usually thin, soft, rough, brittle, pitted, chipped, and abraded, having reduced functional ability and aesthetics. It leads to severe complications in the patient, like early tooth loss, severe discomfort, pain, dental caries, chewing difficulties, and discoloration of teeth from yellow to yellowish-brown or creamy type. The study aimed to identify the disease-causing variant in a consanguineous family. Methods: We recruited a consanguineous Pashtun family of Pakistani origin. Exome sequencing analysis was followed by Sanger sequencing to identify the pathogenic variant in this family. Results: Clinical analysis revealed hypomaturation AI having generalized yellow-brown or creamy type of discoloration in affected members. We identified a novel nonsense sequence variant c.1192C > T (p.Gln398*) in exon-12 of SLC24A4 by using exome sequencing. Later, its co-segregation within the family was confirmed by Sanger sequencing. The human gene mutation database (HGMD, 2019) has a record of five pathogenic variants in SLC24A4, causing AI phenotype. Conclusion: This nonsense sequence variant c.1192C > T (p.Gln398*) is the sixth disease-causing variant in SLC24A4, which extends its mutation spectrum and confirms the role of this gene in the morphogenesis of human tooth enamel. The identified variant highlights the critical role of SLC24A4 in causing a rare AI type in humans.