Current updates on genetic spectrum of usher syndrome.

Usher syndrome (USH) is a genetic disorder that is characterized by sensorineural hearing loss (HL) and visual abnormality, i.e., loss of night vision and side (peripheral) vision. Usher syndrome is categorized into four subtypes (USH1, USH2, USH3, USH4) on the basis of phenotypic spectrum. Profound hearing loss (HL), vestibular are flexia and language disturbance are typically associated with Usher type 1, while USH2 is linked with moderate to severe level of congenital HL. USH3 has late onset of deafness in life (referred to as "postlingual"), inconstant vestibular abnormality and onset of retinitis pigmentosa (RP) typically in 2nd decade of life. Patients with USH4 have no vestibular impairment and have late onset of retinitis pigmentosa (RP) and sensorineural hearing loss. Until now, 15 genetic loci have been reported to be linked with all types of USH. Among reported USH loci, nine are related to be involved in USH1, three in USH2, two in USH3 and one locus in USH4, respectively. Current review has described different types of Usher syndrome and their molecular genetics, and role of usher proteins in sensory organs. Moreover, we also suggested certain candidate genes for uncharacterized loci that may help the molecular geneticist to reach their target easily. Conclusion: The current catalogue of USH genetic data may assist in genetic counseling, genetic diagnosis, and genotype-phenotype correlation.

Identification of GLI1 and KIAA0825 Variants in Two Families with Postaxial Polydactyly.

Polydactyly is a rare autosomal dominant or recessive appendicular patterning defect of the hands and feet, phenotypically characterized by the duplication of digits. Postaxial polydactyly (PAP) is the most common form and includes two main types: PAP type A (PAPA) and PAP type B (PAPB). Type A involves a well-established extra digit articulated with the fifth or sixth metacarpal, while type B presents a rudimentary or poorly developed superfluous digit. Pathogenic variants in several genes have been identified in isolated and syndromic forms of polydactyly. The current study presents two Pakistani families with autosomal recessive PAPA with intra- and inter-familial phenotype variability. Whole-exome sequencing and Sanger analysis revealed a novel missense variant in KIAA0825 (c.3572C>T: p.Pro1191Leu) in family A and a known nonsense variant in GLI1 (c.337C>T: p.Arg113*) in family B. In silico studies of mutant KIAA0825 and GLI1 proteins revealed considerable structural and interactional modifications that suggest an abnormal function of the proteins leading to the disease phenotype. The present study broadens the mutational spectrum of KIAA0825 and demonstrates the second case of a previously identified GLI1 variant with variable phenotypes. These findings facilitate genetic counseling in Pakistani families with a polydactyly-related phenotype.

A GHRHR founder mutation causes isolated growth hormone deficiency type IV in a consanguineous Pakistani family.

Background: Isolated growth hormone deficiency (IGHD) is caused by a severe shortage or absence of growth hormone (GH), which results in aberrant growth and development. Patients with IGHD type IV (IGHD4) have a short stature, reduced serum GH levels, and delayed bone age. Objectives: To identify the causative mutation of IGHD in a consanguineous family comprising four affected patients with IGHD4 (MIM#618157) and explore its functional impact in silico. Methods: Clinical and radiological studies were performed to determine the phenotypic spectrum and hormonal profile of the disease, while whole-exome sequencing (WES) and Sanger sequencing were performed to identify the disease-causing mutation. In-silico studies involved protein structural modeling and docking, and molecular dynamic simulation analyses using computational tools. Finally, data from the Qatar Genome Program (QGP) were screened for the presence of the founder variant in the Qatari population. Results: All affected individuals presented with a short stature without gross skeletal anomalies and significantly reduced serum GH levels. Genetic mapping revealed a homozygous nonsense mutation [NM_000823:c.G214T:p.(Glu72*)] in the third exon of the growth-hormone-releasing hormone receptor gene GHRHR (MIM#139191) that was segregated in all patients. The substituted amber codon (UAG) seems to truncate the protein by deleting the C-terminus GPCR domain, thus markedly disturbing the GHRHR receptor and its interaction with the growth hormone-releasing hormone. Conclusion: These data support that a p.Glu72* founder mutation in GHRHR perturbs growth hormone signaling and causes IGHD type IV. In-silico and biochemical analyses support the pathogenic effect of this nonsense mutation, while our comprehensive phenotype and hormonal profiling has established the genotype-phenotype correlation. Based on the current study, early detection of GHRHR may help in better therapeutic intervention.

The molecular genetics of human appendicular skeleton.

Disorders that result from de-arrangement of growth, development and/or differentiation of the appendages (limbs and digit) are collectively called as inherited abnormalities of human appendicular skeleton. The bones of appendicular skeleton have central role in locomotion and movement. The different types of appendicular skeletal abnormalities are well described in the report of "Nosology and Classification of Genetic skeletal disorders: 2019 Revision". In the current article, we intend to present the embryology, developmental pathways, disorders and the molecular genetics of the appendicular skeletal malformations. We mainly focused on the polydactyly, syndactyly, brachydactyly, split-hand-foot malformation and clubfoot disorders. To our knowledge, only nine genes of polydactyly, five genes of split-hand-foot malformation, nine genes for syndactyly, eight genes for brachydactyly and only single gene for clubfoot have been identified to be involved in disease pathophysiology. The current molecular genetic data will help life sciences researchers working on the rare skeletal disorders. Moreover, the aim of present systematic review is to gather the published knowledge on molecular genetics of appendicular skeleton, which would help in genetic counseling and molecular diagnosis.

In Silico Analysis Identified Putative Pathogenic Missense nsSNPs in Human SLITRK1 Gene.

Human DNA contains several variations, which can affect the structure and normal functioning of a protein. These variations could be single nucleotide polymorphisms (SNPs) or insertion-deletions (InDels). SNPs, as opposed to InDels, are more commonly present in DNA and may cause genetic disorders. In the current study, several bioinformatic tools were used to prioritize the pathogenic variants in the SLITRK1 gene. Out of all of the variants, 16 were commonly predicted to be pathogenic by these tools. All the variants had very low frequency, i.e., <0.0001 in the global population. The secondary structure of all filtered variants was predicted, but no structural change was observed at the site of variation in any variant. Protein stability analysis of these variants was then performed, which determined a decrease in protein stability of 10 of the variants. Amino acid conservation analysis revealed that all the amino acids were highly conserved, indicating their structural and functional importance. Protein 3D structure of wildtype SLITRK1 and all of its variants was predicted using I-TASSER, and the effect of variation on 3D structure of the protein was observed using the Missense3D tool, which presented the probable structural loss in three variants, i.e., Asn529Lys, Leu496Pro and Leu94Phe. The wildtype SLITRK1 protein and these three variants were independently docked with their close interactor protein PTPRD, and remarkable differences were observed in the docking sites of normal and variants, which will ultimately affect the functional activity of the SLITRK1 protein. Previous studies have shown that mutations in SLITRK1 are involved in Tourette syndrome. The present study may assist a molecular geneticist in interpreting the variant pathogenicity in research as well as diagnostic setup.

A novel protein truncating mutation in L2HGDH causes L-2-hydroxyglutaric aciduria in a consanguineous Pakistani family.

BACKGROUND: L-2-hydroxyglutaric aciduria (L2HGA) is a rare neurometabolic disorder that occurs due to accumulation of L-2-hydroxyglutaric acid in the cerebrospinal fluid (CSF), plasma and urine. The clinical manifestation of L2HGA includes intellectual disability, cerebellar ataxia, epilepsy, speech problems and macrocephaly. METHODS: In the present study, we ascertained a multigenerational consanguineous Pakistani family with 5 affected individuals. Clinical studies were performed through biochemical tests and brain CT scan. Locus mapping was carried out through genome-wide SNP genotyping, whole exome sequencing and Sanger sequencing. For in silico studies protein structural modeling and docking was done using I-TASSER, Cluspro and AutoDock VINA tools. RESULTS: Affected individuals presented with cognitive impairment, gait disturbance, speech difficulties and psychomotor delay. Radiologic analysis of a male patient revealed leukoaraiosis with hypoattenuation of cerebral white matter, suggestive of hypomyelination. Homozygosity mapping in this family revealed a linkage region on chromosome 14 between markers rs2039791 and rs781354. Subsequent whole exome analysis identified a novel frameshift mutation NM_024884.3:c.180delG, p.(Ala62Profs*24) in the second exon of L2HGDH. Sanger sequencing confirmed segregation of this mutation with the disease phenotype. The identification of the most N-terminal loss of function mutation published thus far further expands the mutational spectrum of L2HGDH.

Alopecia-mental retardation syndrome: Molecular genetics of a rare neuro-dermal disorder.

Alopecia-mental retardation syndrome (APMR) is a rare autosomal recessive neuro-dermal disorder. It is characterized by heterogeneous phenotypic features, that is, absence of hair on the scalp, eyelashes, and eyebrows and mild to severe intellectual disability. So far, approximately 14 families (i.e., Iranian, Pakistani, and Swiss) with APMR have been reported in the scientific literature. Its precise prevalence is still unknown, but according to a predictive estimate, it prevails with the ratio of 1 in 1,000,000 persons worldwide. Until now, only four loci (two characterized and two uncharacterized) have been reported to be involved in APMR. The pathogenic variants in alpha-2-HS-glycoprotein [AHSG; APMR1 (MIM#203650)] and lanosterol synthase [LSS; APMR4 (MIM#618840)] are the characterized genetic factors associated with APMR. Among them, AHSG was reported in a consanguineous Iranian family and LSS gene in a Swiss origin family, while the remaining two uncharacterized loci, that is, APMR2 and APMR3, are reported in the Pakistani population. The current mini-report discusses the molecular genetics and mutational spectrum of APMR syndrome, its differential diagnosis from related disorders, and prediction of plausible candidate genes in two uncharacterized loci.

The molecular genetics of UV-Sensitive syndrome: A rare dermal anomaly.

Ultraviolet-sensitive syndrome is a rare skin disorder characterised by heterogeneous phenotypic spectrum of skin freckling, telangiectasia and acute sunburn. It usually has an autosomal recessive pattern. So far, only 18 patients from nine different families (Japanese, French, Israeli, Iranian and Pakistani) have been reported in scientific literature. Its precise prevalence is still unknown, but, according to an estimate, its prevalence ratio is 1:100,000 worldwide. Until now, only three genes have been reported to be involved in the syndrome; the Excision Repair Cross-Complementing, Group 6, the Excision Repair Cross-Complementing, Group 8 and the UV-Stimulated Scaffold Protein A (UVSSA). Among these genes, the last one is reported to be more prevalent among different ethnicities, including Pakistani. Physiologically, most of the syndrome genes are involved in the transcription-coupled nucleotide excision pathway. In order to reduce the disease severity, the patients are advised to use medicated skin moisturisers or sun-blocks, sunglasses and gloves, while going out in the sun to avoid sun exposure. The current narrative review was planned to discuss the molecular genetics and the mutational spectrum of the syndrome, and to describe the differential diagnosis of various related disorders in order to facilitate clinical researchers.

Investigating the thermodynamic and kinetics properties of acid phosphatase extracted and purified from seedlings of Chenopodium murale.

Herein acid phosphatase isoenzyme was extracted from the C. murale seedlings. The purification was accomplished by chromatographic techniques and passing through DEAE-cellulose and Sephadex G-100 column. The specific activity of acid phosphatase 5.75 U/mg of protein was obtained with 66 purification fold 15.8% yield and molecular mass was 29 kDa with very faint bands corresponding to 18 kDa and 14 kDa. The maximal activity at pH 5.0 and 50 °C best illustrated by first order kinetics. When temperature was raised (55 °C to 75 °C), the deactivation rate constant was increased from 0.001 to 0.014 min-1, while half-life was decreased from 693 to 49 min-1. The results of activity collected at different temperature were then used to estimate, activation energy of hydrolysis reaction (Ea = 47.59 kJmol-1). A high Z-value (18.86 °C min-1) was obtained indicating a less sensitivity towards temperatures. The residual activity examinations were carried out from 55 °C to 75 °C and assessing the Deactivation Energy (Ed 116.39 kJmol-1), Enthalpy change (ΔH° 113.55kJmol-1), Entropy change (ΔS° 110.33kJmol-1) and change in Gibbs free energy (ΔG° 10.02 kJmol-1). Taken together, thermodynamic parameters confirm the high stability of enzyme and show potential commercial applicability.

Investigating the Impact of Various Parameters On the Activity of Acid Phosphatases from Seedlings of Coronopus didymus.

The thermal stability of purified acid phosphatase from the germinating seedlings of Coronopus didymus (Jangli halon) was investigated by studying the impact of various thermodynamic parameters [t1/2, Ed, ΔH° (enthalpy change), ΔG° (free energy change), and ΔS° (entropy change)] of heat treatment in the temperature range of 55-75 °C. The thermal denaturation of acid phosphatase, assessed by loss in activity, was evidently followed by first-order kinetics, which varies with time and yield during the process of denaturation. The half-life of the enzyme was 693 min at 55 °C. The Ed (activation energy of denaturation) was calculated by the Arrhenius plot (30 kcal mol-1), and the Z-value was 17.3 °C. The various thermodynamic parameters studied were as follows: ΔH°, the change in enthalpy of inactivation, was 121.93 kJ mol-1 at 55 °C; ΔG°, the change in free energy of inactivation, was 110.65 kJ mol-1 at 55 °C; and ΔS°, the change in entropy of inactivation, was 34.39 J mol-1 k-1 at 55 °C. This suggests that acid phosphatase activity is thermostable to long heat treatment up to 60 °C.

Identification of a novel protein truncating mutation p.Asp98* in XPC associated with xeroderma pigmentosum in a consanguineous Pakistani family.

BACKGROUND: Xeroderma pigmentosum (XP) is a rare genetic disorder, which is characterized by hyper-sensitivity to solar ultraviolet (UV) radiation. Clinical consequences of sun exposure are skin lesions and an increased risk of developing skin cancer. Genetic studies have identified eight genes associated with xeroderma pigmentosum. The proteins encoded by these genes are mainly involved in DNA repair mechanisms. METHODS: Molecular genetic characterization of patients with xeroderma pigmentosum involved positional cloning methods such as homozygosity mapping and subsequent candidate gene analysis. Mutation screening was performed through Sanger DNA sequencing. RESULTS AND DISCUSSION: In this case study, we report a novel protein truncating mutation in XPC associated with autosomal recessive xeroderma pigmentosum in a consanguineous Pakistani family. Genetic mapping revealed a novel single base insertion of a thymine nucleotide NM_004628.4: c.291dupT (c.291_292insT) in the second exon of XPC. The identified mutation leads to a premature stop codon (TGA) at amino acid position 98 (p.Asp98*) and thus presumably results in a truncated protein. The Xeroderma pigmentosum, complementation group C (XPC) is located on 3p25.1 and encodes a protein involved in nucleotide excision repair. The identified mutation presumably truncates all functional domains of the XPC protein, which likely results in the loss of protein function. CONCLUSION: The study expands the knowledge of the mutational spectrum of XPC and is valuable for genetic counseling of affected individuals and their families.

Electrochemical and spectroscopic investigations of protonated ferrocene-DNA intercalation.

The interaction of protonated ferrocene (PF) with chicken blood DNA (CB-DNA) has been investigated in vitro by cyclic voltammetry (CV) and UV-Vis spectroscopy as well as viscosity measurements under stomach pH and body temperature. The peak potentials shift in CV, hyperchromism in UV absorption titration, an increase in the viscosity of DNA and the results of the effect of ionic strength on the binding constant strongly support the intercalation of PF into the DNA double helix. The diffusion coefficients of PF in the presence and absence of DNA were 9.54 x 10(-11) and 1.34 x 10(-10) m2/s, respectively. The binding constant of the PF-DNA complex and the number of binding sites on a DNA molecule were calculated as being 3.07 x 10(2) M(-1) and 2.96, with the help of the Scatchard equation. An expression by Carter et al. was used for determining the binding site size (0.17 bp). The binding constant was also determined by UV absorption titration.