Variants in exon 2 of MED12 gene causes uterine leiomyoma's through over-expression of MMP-9 of ECM pathway.

AIMS: To study the impact of Mediator complex subunit 12 (MED12) gene variants on the encoded protein's function and pathogenic relevance for genesis of uterine leiomyoma's (ULs). METHODS: Mutational analysis in exon-2 of MED12 gene was performed by PCR amplification and DNA sequencing in 89 clinically diagnosed ULs tissues. Pathogenicity prediction of variation was performed by computational analysis. The functional effects of missense variation were done by quantity RT-PCR and western blot analysis. RESULT(S): Out of 89 samples, 40 (44.94%) had missense variation in 14 different CDS position of exon-2 of MED12 gene. Out of 40 missense variation, codon 44 had 25 (62.5%) looking as a hotspot region for mutation for ULs, because CDS position c130 and c131present at codon 44 that have necleotide change G>A, T, C at c130 and c131 have necleotide change G>A and C. We also find somenovel somatic mutations oncodon 36 (T > C), 38 (G>T) of exon-2 and 88 (G>C) of intron-2. No mutations were detected in uterine myometrium samples. Our computational analysis suggests that change in Med12c .131 G>A leads to single substitution of amino acid [Glycine (G) to Aspartate (D)] which has a pathogenic and lethal impact and may cause instability of MED12 protein. Further, analysis of extracellular matrix (ECM) component (MMP-2 & 9, COL4A2 and α-SMA) mRNA and protein expression levels in the set of ULs having MED12 mutation showed significantly higher expression of MMP-9 and α-SMA. CONCLUSION(S): The findings of present study suggest that missense variation in codon 44 of MED12 gene lead to the genesis of leiomyoma's through over-expression of MMP-9 of ECM pathway which could be therapeutically targeted for non-surgical management of ULs.

Identification and molecular characterization of two recurrent missense mutations in the RS1 gene in two families with X-linked retinoschisis from North India.

X-linked retinoschisis (XLR) is a rare medical condition that involves in the splitting of neurosensory layers and the impairment of vision in the retina. In majority of the XLR cases, pathogenic variants in Retinoschisin 1 (RS1) gene have been implicated in males with an early age of onset during early childhood. In the present study, we have recruited two North Indian families having multiple affected male members, who were diagnosed with XLR. The entire protein-coding region of RS1 was screened by PCR-Sanger sequencing and two recurrent pathogenic variants (p.I81N and p.R102Q) were unraveled. The in vitro study of these variants demonstrated the aggregation of mutant RS1 within the endoplasmic reticulum. Furthermore, mutant forms of this protein showed significant intracellular retention, which was evident by the absence of retinoschisin protein fractions in the extracellular media. These inferences were also supported by extensive bioinformatics analysis of the mutants, which showed dramatic conformational changes in the local structure of retinoschisin. Thus, our study suggests that the identified pathogenic variants interfere with proper protein folding, leading to anomalous structural changes ultimately resulting in intracellular retention of retinoschisin within the retina.

Whole exome sequencing identifies a novel splice-site mutation in IMPG2 gene causing Stargardt-like juvenile macular dystrophy in a north Indian family.

We report on the genetic analysis of a north Indian family affected with Stargardt-like juvenile macular dystrophy. Considering an autosomal recessive inheritance of macular dystrophy in the recruited family, whole exome sequencing was employed in two affected siblings and their mother. We have identified a novel splice-site variant NC_000003.11(NM_016247.3):c.1239 + 1G > T, co-segregating in the affected siblings, in the Interphotoreceptor Matrix Proteoglycan 2 (IMPG2) gene. The identified variant is present immediately after exon 11, and is predicted to disrupt the wild-type donor splice-site of IMPG2 transcripts. We confirmed the splice-site changes in the IMPG2 transcripts using minigene functional assay. Although a number of studies on IMPG2 have demonstrated its involvement in retinitis pigmentosa and vitelliform macular dystrophy, this is the first report of a splice-site variant in IMPG2 that is responsible for Stargardt-like juvenile macular dystrophy.

TP53 codon 72 polymorphism and the risk of glaucoma in a north Indian cohort: A genetic association study.

BACKGROUND: The TP53 codon 72 Proline-Arginine polymorphism (TP53 P72R) is the most widely studied candidate among those evaluated for a putative association between impaired apoptosis and glaucoma. Considering the earlier findings about enhanced apoptotic potential by the Arg variant of TP53 P72R and the conflicting results about its association with glaucoma, we initiated a hospital-based case-control association study in a north Indian cohort to investigate the association of TP53 P72R with glaucoma. MATERIALS AND METHODS: We examined the status of TP53 P72R in 139 cases of primary open angle glaucoma (POAG) and in 111 cases of primary angle closure glaucoma (PACG) with respect to 218 controls using the polymerase chain reaction-restriction fragment length polymorphism method. Logistic regression analysis including age and gender as covariates was carried out to test the association of the polymorphism with overall glaucoma, POAG, and PACG cases. RESULTS: We observed significant differences between the genotypic distributions of combined glaucoma cases and controls in the recessive model. POAG cases with respect to controls did not exhibit any significant differences in the genotypic distributions. In contrast, the genotypic distributions as per the additive and recessive models in PACG cases were significantly different from those in controls. The two models suggested an increased risk of PACG in the Arg homozygotes of the investigated cohort. CONCLUSIONS: Ours is the first study demonstrating the association of TP53 P72R with the risk of PACG. It emphasizes that apart from narrow anterior chamber angle, impaired apoptotic mechanisms could also be an important contributor toward PACG.

Whole exome sequencing: Uncovering causal genetic variants for ocular diseases.

Identification of causal genetic defects for human diseases took a significant leap when the first generation DNA sequencing technologies enabled biologists extract sequence-based genetic information from living beings. However, these sequencing methods had unavoidable constraints of throughput, scalability, rapidity, and resolution. In this direction, next-generation sequencing (NGS) since the time of its advent has revolutionized the process of gene discovery for both monogenic and multifactorial genetic diseases. Among several variations of NGS, whole exome sequencing (WES) has emerged as a smart strategy that enables identification of disease causing variants present within the coding region of the human genome. The current review focuses primarily on the application of WES in identification of causal variants for ocular diseases. WES has successfully revealed pathogenic variants in a variety of ocular diseases such as retinal degenerations, refractive errors, lens diseases, corneal dystrophies, and developmental ocular defects. It has demonstrated immense potential for molecular diagnosis of genetic ocular diseases. WES has been extensively used in Mendelian and complex cases, familial and sporadic cases, simplex and multiplex cases, and syndromic and non-syndromic cases of ocular diseases. Although many such ocular diseases have been investigated using WES, reports indicate that it has been employed overwhelmingly for heterogeneous retinal degenerations. WES, within a short period of time, has proved to be a cost-effective and promising approach for understanding the genetic basis of ocular diseases.