Inherited retinal diseases (IRDs) are a diverse set of visual disorders that collectively represent a major cause of early-onset blindness. With the reduction in sequencing costs in recent years, whole-genome sequencing (WGS) is being used more frequently, particularly when targeted gene panels and whole-exome sequencing (WES) fail to detect pathogenic mutations in patients. In this study, we performed mutation screens using WGS for a cohort of 311 IRD patients whose mutations were undetermined. A total of nine putative pathogenic mutations in six IRD patients were identified, including six novel mutations. Among them, four were deep intronic mutations that affected mRNA splicing, while the other five affected protein-coding sequences. Our results suggested that the rate of resolution of unsolved cases via targeted gene panels and WES can be further enhanced with WGS; however, the overall improvement may be limited.

CCND1 encodes for Cyclin D1 protein and single nucleotide polymorphisms (SNPs) can modulate its activity. In the current study, the impact of CCND1 SNPs on structure and/or function of cyclin D1 protein using in silico tools was investigated. Our analysis revealed only one splice site SNP (c.1988+5G<A) can effect CCND1 function. Subsequently, 78 out of 169 missense variants were predicted as pathogenic by Polyphen2, SIFT, PROVEAN, SNP & GO and PANTHER, and 4/78 missense SNPs were further evaluated because these four SNPs were found to be reside in highly conserved region of cyclin D1. However, they did not show any major impact on tertiary structure and domain of cyclin D1 but overall R15S and A190S has displayed a significant diseased phenotype and an altered molecular mechanism predicted by MutPred, FATHMM, SNPeffect, SNAP2, and PredictSNP. Consistently, A190S, R179L and R15S may also cause a decrease in stability of cyclin D1 anticipated by I-Mutant, HOPE and SNPeffect. Furthermore, the Kaplan-Meier plotter has explained that high expression of CCND1 is associated with less survival rate of breast cancer patients. Altogether, our study suggests that c.1988+5G<A, R15S, R179L and A190S SNPs could directly or indirectly destabilize cyclin D1.

Background: Loss of function mutations in the spermine synthase gene (SMS) have been reported to cause a rare X-linked intellectual disability known as Snyder-Robinson Syndrome (SRS). Besides intellectual disability, SRS is also characterized by reduced bone density, osteoporosis and facial dysmorphism. SRS phenotypes evolve with age from childhood to adulthood. Methods: Whole exome sequencing was performed to know the causative gene/pathogenic variant. Later we confirmed the pathogenic variant through Sanger sequencing. Furthermore, we also performed the mutational analysis through HOPE SERVER and SWISS-MODEL. Also, radiographs were also obtained for affected individual to confirm the disease features. Results: In this article, we report the first Pakistani family consisting of three patients with SRS and a novel missense pathogenic variant in the SMS gene (c.905 C > T p.(Ser302Leu)). In addition to the typical phenotypes, one patient presented with early-onset seizures. Clinical features, genetic and in-silico analysis linked the affected patients of the family with Snyder-Robinson and suggest that this novel mutation affects the spermine synthase activity. Conclusion: A novel missense variant in the SMS, c.905C > T p. (Ser302Leu), causing Snyder- Robinson Syndrome (SRS) is reported in three members of Pakistani Family.

Background: Patients with focal segmental glomerulosclerosis (FSGS) characterized by steroid-resistant nephrotic syndrome (SRNS) are prone to progress to ESRD. Mechanism for the FSGS patients’ response to steroid treatment is still unknown and currently, it is impossible to predict the steroid resistance before treatment of patients with FSGS. Methods: To identify biomarkers and potential therapeutic targets of FSGS patients with SRNS, patients diagnosed as kidney biopsy-proven FSGS and nephrotic syndrome (NS) were prospectively enrolled. They were divided into 2 groups, steroid-sensitive NS and SRNS based on their treatment response. Cortical regions were selected from biopsied renal tissues, and glomeruli were isolated under an inverted microscope. RNA was prepared from the isolated glomeruli and further used for microarray analysis. Followed by multiple analyses, the top 6 highest and lowest, and a selected panel of differentially expressed genes obtained and their related pathways were validated via real-time PCR, western blot, and measurement of reactive oxygen species (ROS). Results: In SRNS group, we discovered that the most significant up-regulated pathway was primarily related to cellular amino acid and derivative metabolic process. Meanwhile, the most significant down-regulated pathway was primarily involved in anatomical structure morphogenesis. Moreover, we found NADPH oxidase 4 (NOX4), one of the key regulators of renal ROS, at a much higher level in SRNS both at transcriptomic and proteomic levels. We also found the levels of ROS, p-p38 MAPK and matrix metalloproteinase (MMP)-2, which were all regulated by NOX4, were also higher in glomeruli isolated from SRNS patients. At last, we detected stimulated by retinoic acid gene 6 homolog (STRA6), a cell surface receptor formerly known as a gene preventing podocytes from over-proliferative lesion induced by HIV infection and was up-regulated by retinoic acid, expressed at a much higher level in SRNS kidneys. Conclusion: We found 2 potential mechanisms underline the SRNS, NOX4/ROS/P38 MAPK/MMP-2 pathway and STRA6. Our findings provided new insights into the steroid resistance.

Background: Congenital hypogonadotropic hypogonadism (CHH) is a heterogeneous disorder characterized by delayed or loss of puberty and infertility due to functional deficiency in the hypothalamic gonadotropin-releasing hormone (GnRH). CHH can be classified into 2 subtypes on the basis of olfaction: Kallmann syndrome and normosmic CHH (nCHH). The spectrum of genetic variants causing CHH is continually expanding. Here, we recruited a consanguineous Pakistani family having 2 male and 2 female infertile patients diagnosed with idiopathic nCHH. Aims: The aim of this study was to investigate the genetic cause of nCHH in the family. Methods: Clinical and physical analyses were performed for the patients. Genetic analysis was carried out using whole exome and Sanger sequencing. Results: Clinical and physical investigations confirmed low levels of gonadotropins and failure of secondary sexual development in the patients. Genetic analysis identified a novel nonsense mutation (chr4: g.68619942G>A, c.112C>T, p.Arg38*) in the gonadotropin-releasing hormone receptor gene (GNRHR) recessively co-segregating with nCHH in this family. All the patients are homozygous and their parents are heterozygous carriers, while normal siblings are heterozygous carriers or wild-type for this mutation, indicating that the identified mutation is pathogenic for nCHH in the family. Conclusion: We report the first homozygous nonsense mutation in the GNRHR gene (chr4: g. 68619942G>A, c.112C>T, p. Arg38*) that is associated with familial nCHH. Hence, our study displayed a good correlation of the genotype and phenotype of nCHH patients.