Identification of a de novo LRP1 mutation in a Saudi family with Tetralogy of Fallot.

BACKGROUND: Tetralogy of Fallot (TOF) is a rare, complex congenital heart defect caused by genetic and environmental interactions that results in abnormal heart development during the early stages of pregnancy. Genetic basis of TOF in Saudi populations is not yet studied. Therefore, the objective of this study is to screen for the molecular defects causing TOF in Saudi patients. METHODS: A family with non-syndromic TOF was recruited from the Western region of Saudi Arabia. Whole exome sequencing (WES) was performed on the proband and her parents. The identified candidate variant was verified by sanger sequencing. Also, different computational biology tools were used to figure out how candidate variants affect the structure and function of candidate protein involved in TOF. RESULTS: A novel heterozygous de novo mutation in LRP1 (p. G3311D) gene was identified in the index case. Also, this variant was absent in the in-house exome sequencing data of 80 healthy Saudi individuals. This variant was predicted to be likely pathogenic, as it negatively affects the biophysical chemical properties and stability of the protein. Furthermore, functional biology data from knock out mouse models confirms that molecular defects in LRP1 gene leads to cardiac defects and lethality. This variant was not previously reported in both Arab and global population genetic databases. CONCLUSION: The findings in this study postulate that the LRP1 variant has a role in TOF pathogenesis and facilitate accurate diagnosis as well as the understanding of underlying molecular mechanisms and pathophysiology of the disease.

Retinal pigment epithelium extracellular vesicles are potent inducers of age-related macular degeneration disease phenotype in the outer retina.

Age-related macular degeneration (AMD) is a leading cause of blindness. Vision loss is caused by the retinal pigment epithelium (RPE) and photoreceptors atrophy and/or retinal and choroidal angiogenesis. Here we use AMD patient-specific RPE cells with the Complement Factor H Y402H high-risk polymorphism to perform a comprehensive analysis of extracellular vesicles (EVs), their cargo and role in disease pathology. We show that AMD RPE is characterised by enhanced polarised EV secretion. Multi-omics analyses demonstrate that AMD RPE EVs carry RNA, proteins and lipids, which mediate key AMD features including oxidative stress, cytoskeletal dysfunction, angiogenesis and drusen accumulation. Moreover, AMD RPE EVs induce amyloid fibril formation, revealing their role in drusen formation. We demonstrate that exposure of control RPE to AMD RPE apical EVs leads to the acquisition of AMD features such as stress vacuoles, cytoskeletal destabilization and abnormalities in the morphology of the nucleus. Retinal organoid treatment with apical AMD RPE EVs leads to disrupted neuroepithelium and the appearance of cytoprotective alpha B crystallin immunopositive cells, with some co-expressing retinal progenitor cell markers Pax6/Vsx2, suggesting injury-induced regenerative pathways activation. These findings indicate that AMD RPE EVs are potent inducers of AMD phenotype in the neighbouring RPE and retinal cells.

pRB-Depleted Pluripotent Stem Cell Retinal Organoids Recapitulate Cell State Transitions of Retinoblastoma Development and Suggest an Important Role for pRB in Retinal Cell Differentiation.

Retinoblastoma (Rb) is a childhood cancer of the developing retina, accounting for up to 17% of all tumors in infancy. To gain insights into the transcriptional events of cell state transitions during Rb development, we established 2 disease models via retinal organoid differentiation of a pRB (retinoblastoma protein)-depleted human embryonic stem cell line (RB1-null hESCs) and a pRB patient-specific induced pluripotent (iPSC) line harboring a RB1 biallelic mutation (c.2082delC). Both models were characterized by pRB depletion and accumulation of retinal progenitor cells at the expense of amacrine, horizontal and retinal ganglion cells, which suggests an important role for pRB in differentiation of these cell lineages. Importantly, a significant increase in the fraction of proliferating cone precursors (RXRγ+Ki67+) was observed in both pRB-depleted organoid models, which were defined as Rb-like clusters by single-cell RNA-Seq analysis. The pRB-depleted retinal organoids displayed similar features to Rb tumors, including mitochondrial cristae aberrations and rosette-like structures, and were able to undergo cell growth in an anchorage-independent manner, indicative of cell transformation in vitro. In both models, the Rb cones expressed retinal ganglion and horizontal cell markers, a novel finding, which could help to better characterize these tumors with possible therapeutic implications. Application of Melphalan, Topotecan, and TW-37 led to a significant reduction in the fraction of Rb proliferating cone precursors, validating the suitability of these in vitro models for testing novel therapeutics for Rb.

Activation of autophagy reverses progressive and deleterious protein aggregation in PRPF31 patient-induced pluripotent stem cell-derived retinal pigment epithelium cells.

INTRODUCTION: Mutations in pre-mRNA processing factor 31 (PRPF31), a core protein of the spliceosomal tri-snRNP complex, cause autosomal-dominant retinitis pigmentosa (adRP). It has remained an enigma why mutations in ubiquitously expressed tri-snRNP proteins result in retina-specific disorders, and so far, the underlying mechanism of splicing factors-related RP is poorly understood. METHODS: We used the induced pluripotent stem cell (iPSC) technology to generate retinal organoids and RPE models from four patients with severe and very severe PRPF31-adRP, unaffected individuals and a CRISPR/Cas9 isogenic control. RESULTS: To fully assess the impacts of PRPF31 mutations, quantitative proteomics analyses of retinal organoids and RPE cells were carried out showing RNA splicing, autophagy and lysosome, unfolded protein response (UPR) and visual cycle-related pathways to be significantly affected. Strikingly, the patient-derived RPE and retinal cells were characterised by the presence of large amounts of cytoplasmic aggregates containing the mutant PRPF31 and misfolded, ubiquitin-conjugated proteins including key visual cycle and other RP-linked tri-snRNP proteins, which accumulated progressively with time. The mutant PRPF31 variant was not incorporated into splicing complexes, but reduction of PRPF31 wild-type levels led to tri-snRNP assembly defects in Cajal bodies of PRPF31 patient retinal cells, altered morphology of nuclear speckles and reduced formation of active spliceosomes giving rise to global splicing dysregulation. Moreover, the impaired waste disposal mechanisms further exacerbated aggregate formation, and targeting these by activating the autophagy pathway using Rapamycin reduced cytoplasmic aggregates, leading to improved cell survival. CONCLUSIONS: Our data demonstrate that it is the progressive aggregate accumulation that overburdens the waste disposal machinery rather than direct PRPF31-initiated mis-splicing, and thus relieving the RPE cells from insoluble cytoplasmic aggregates presents a novel therapeutic strategy that can be combined with gene therapy studies to fully restore RPE and retinal cell function in PRPF31-adRP patients.

Genome-Wide Association Study-Guided Exome Rare Variant Burden Analysis Identifies IL1R1 and CD3E as Potential Autoimmunity Risk Genes for Celiac Disease.

Celiac disease (CeD) is a multifactorial autoimmune enteropathy characterized by the overactivation of the immune system in response to dietary gluten. The molecular etiology of CeD is still not well-understood. Therefore, this study aims to identify potential candidate genes involved in CeD pathogenesis by applying multilayered system biology approaches. Initially, we identified rare coding variants shared between the affected siblings in two rare Arab CeD families by whole-exome sequencing (WES). Then we used the STRING database to construct a protein network of rare variants and genome-wide association study (GWAS) loci to explore their molecular interactions in CeD. Furthermore, the hub genes identified based on network topology parameters were subjected to a series of computational validation analyses like pathway enrichment, gene expression, knockout mouse model, and variant pathogenicity predictions. Our findings have shown the absence of rare variants showing classical Mendelian inheritance in both families. However, interactome analysis of rare WES variants and GWAS loci has identified a total of 11 hub genes. The multidimensional computational analysis of hub genes has prioritized IL1R1 for family A and CD3E for family B as potential genes. These genes were connected to CeD pathogenesis pathways of T-cell selection, cytokine signaling, and adaptive immune response. Future multi-omics studies may uncover the roles of IL1R1 and CD3E in gluten sensitivity. The present investigation lays forth a novel approach integrating next-generation sequencing (NGS) of familial cases, GWAS, and computational analysis for solving the complex genetic architecture of CeD.

Exploring the role of Islam on the lived experience of patients with Long QT Syndrome in Saudi Arabia.

Genetic services are rapidly growing in the Arab world leading to increasing number of patients being diagnosed with genetic disorders. Islam is the only/major religion of the local population in these countries. Muslim patients integrate religion in virtually every aspect of their lives, and it is vital to understand the role of Islam on their coping and decision-making in the context of genetic counseling. This will help provide patients with the most appropriate services aligned to their religious beliefs and will improve outcomes. Increasing numbers of patients are being diagnosed with Long QT syndrome in Saudi Arabia. Using semi-structured interviews, this study explored the role of Islam on the lived experience of 13 Saudi participants diagnosed with autosomal dominant Long QT syndrome (3/13) or who are carriers of Jervell and Lange-Nielsen syndrome (10/13). The interviews investigated how they made sense of living with the condition in light of their religion/spirituality. The data were analyzed using interpretative phenomenological analysis and produced four superordinate themes: 1) Common belief and idiosyncratic interpretation; 2) Using religion to justify positive reframing of current illnesses; 3) Interplay between belief in medicine and in religion; and 4) Complex impact of diagnosis on religiosity. The results show that the participants' idiosyncratic interpretations of the religious principles, not the principles themselves, had an important influence on their coping, medical decision-making, perceptions regarding the cause of their disease, and compliance with medical advice. A novel insight of the current study is that the personal understanding and interpretation of medical information played the greatest role in the decision-making process, and not the religious beliefs. Thus, it is important for health professionals to give patients' information in a manner that is clear and detailed in order for them to facilitate an informed decision, and to ensure that they fully understand the implications.

Novel MYO1D Missense Variant Identified Through Whole Exome Sequencing and Computational Biology Analysis Expands the Spectrum of Causal Genes of Laterality Defects.

Laterality defects (LDs) or asymmetrically positioned organs are a group of rare developmental disorders caused by environmental and/or genetic factors. However, the exact molecular pathophysiology of LD is not yet fully characterised. In this context, studying Arab population presents an ideal opportunity to discover the novel molecular basis of diseases owing to the high rate of consanguinity and genetic disorders. Therefore, in the present study, we studied the molecular basis of LD in Arab patients, using next-generation sequencing method. We discovered an extremely rare novel missense variant in MYO1D gene (Pro765Ser) presenting with visceral heterotaxy and left isomerism with polysplenia syndrome. The proband in this index family has inherited this homozygous variant from her heterozygous parents following the autosomal recessive pattern. This is the first report to show MYO1D genetic variant causing left-right axis defects in humans, besides previous known evidence from zebrafish, frog and Drosophila models. Moreover, our multilevel bioinformatics-based structural (protein variant structural modelling, divergence, and stability) analysis has suggested that Ser765 causes minor structural drifts and stability changes, potentially affecting the biophysical and functional properties of MYO1D protein like calmodulin binding and microfilament motor activities. Functional bioinformatics analysis has shown that MYO1D is ubiquitously expressed across several human tissues and is reported to induce severe phenotypes in knockout mouse models. In conclusion, our findings show the expanded genetic spectrum of LD, which could potentially pave way for the novel drug target identification and development of personalised medicine for high-risk families.

Pre-mRNA Processing Factors and Retinitis Pigmentosa: RNA Splicing and Beyond.

Retinitis pigmentosa (RP) is the most common inherited retinal disease characterized by progressive degeneration of photoreceptors and/or retinal pigment epithelium that eventually results in blindness. Mutations in pre-mRNA processing factors (PRPF3, 4, 6, 8, 31, SNRNP200, and RP9) have been linked to 15-20% of autosomal dominant RP (adRP) cases. Current evidence indicates that PRPF mutations cause retinal specific global spliceosome dysregulation, leading to mis-splicing of numerous genes that are involved in a variety of retina-specific functions and/or general biological processes, including phototransduction, retinol metabolism, photoreceptor disk morphogenesis, retinal cell polarity, ciliogenesis, cytoskeleton and tight junction organization, waste disposal, inflammation, and apoptosis. Importantly, additional PRPF functions beyond RNA splicing have been documented recently, suggesting a more complex mechanism underlying PRPF-RPs driven disease pathogenesis. The current review focuses on the key RP-PRPF genes, depicting the current understanding of their roles in RNA splicing, impact of their mutations on retinal cell's transcriptome and phenome, discussed in the context of model species including yeast, zebrafish, and mice. Importantly, information on PRPF functions beyond RNA splicing are discussed, aiming at a holistic investigation of PRPF-RP pathogenesis. Finally, work performed in human patient-specific lab models and developing gene and cell-based replacement therapies for the treatment of PRPF-RPs are thoroughly discussed to allow the reader to get a deeper understanding of the disease mechanisms, which we believe will facilitate the establishment of novel and better therapeutic strategies for PRPF-RP patients.

Saudi Familial Hypercholesterolemia Patients With Rare LDLR Stop Gain Variant Showed Variable Clinical Phenotype and Resistance to Multiple Drug Regimen.

Familial hypercholesterolemia (FH), a well-known lipid disease caused by inherited genetic defects in cholesterol uptake and metabolism is underdiagnosed in many countries including Saudi Arabia. The present study aims to identify the molecular basis of severe clinical manifestations of FH patients from unrelated Saudi consanguineous families. Two Saudi families with multiple FH patients fulfilling the combined FH diagnostic criteria of Simon Broome Register, and the Dutch Lipid Clinic Network (DLCN) were recruited. LipidSeq, a targeted resequencing panel for monogenic dyslipidemias, was used to identify causative pathogenic mutation in these two families and in 92 unrelated FH cases. Twelve FH patients from two unrelated families were sharing a very rare, pathogenic and founder LDLR stop gain mutation i.e., c.2027delG (p.Gly676Alafs*33) in both the homozygous or heterozygous states, but not in unrelated patients. Based on the variant zygosity, a marked phenotypic heterogeneity in terms of LDL-C levels, clinical presentations and resistance to anti-lipid treatment regimen (ACE inhibitors, ß-blockers, ezetimibe, statins) of the FH patients was observed. This loss-of-function mutation is predicted to alter the free energy dynamics of the transcribed RNA, leading to its instability. Protein structural mapping has predicted that this non-sense mutation eliminates key functional domains in LDLR, which are essential for the receptor recycling and LDL particle binding. In conclusion, by combining genetics and structural bioinformatics approaches, this study identified and characterized a very rare FH causative LDLR pathogenic variant determining both clinical presentation and resistance to anti-lipid drug treatment.

TagSNP approach for HLA risk allele genotyping of Saudi celiac disease patients: effectiveness and pitfalls.

BACKGROUND: Celiac disease (CD) is a genetically complex autoimmune disease which is triggered by dietary gluten. Human leukocyte antigen (HLA) class II genes are known to act as high-risk markers for CD, where >95% of CD patients carry (HLA), DQ2 and/or DQ8 alleles. Therefore, the present study was conducted to investigate the distribution of HLA haplotypes among Saudi CD patients and healthy controls by using the tag single nucleotide polymorphisms (SNP). METHODS: HLA-tag SNPs showing strong linkage value (r2>0.99) were used to predict the HLA DQ2 and DQ8 genotypes in 101 Saudi CD patients and in 103 healthy controls by using real-time polymerase chain reaction technique. Genotype calls were further validated by Sanger sequencing method. RESULTS: A total of 63.7% of CD cases and of 60.2% of controls were predicted to carry HLA-DQ2 and DQ8 heterodimers, either in the homozygous or heterozygous states. The prevalence of DQ8 in our CD patients was predicted to be higher than the patients from other ethnic populations (35.6%). More than 32% of the CD patients were found to be non-carriers of HLA risk haplotypes as predicted by the tag SNPs. CONCLUSION: The present study highlights that the Caucasian specific HLA-tag SNPs would be of limited value to accurately predict CD specific HLA haplotypes in Saudi population, when compared with the Caucasian groups. Prediction of risk haplotypes by tag SNPs in ethnic groups is a good alternate approach as long as the tag SNPs were identified from the local population genetic variant databases.

Identification of a Rare Exon 19 Skipping Mutation in ALMS1 Gene in Alström Syndrome Patients From Two Unrelated Saudi Families.

Background: Alström syndrome (AS) is a very rare childhood disorder characterized by cardiomyopathy, progressive hearing loss and blindness. Inherited genetic variants of ALMS1 gene are the known molecular cause of this disease. The objective of this study was to characterize the genetic basis and understand the genotype-phenotype relationship in Saudi AS patients. Methods: Clinical phenotyping and whole-exome sequencing (WES) analysis were performed on six AS patients belonging to two unrelated consanguineous Saudi families. Sanger sequencing was performed to determine the mode of inheritance of ALMS1 variant in first-degree family relatives and also to ensure its rare prevalence in 100 healthy population controls. Results: We identified that Alström patients from both the families were sharing a very rare ALMS1, 3'-splice site acceptor (c.11873-2 A>T) variant, which skips entire exon-19 and shortens the protein by 80 amino acids. This disease variant was inherited by AS patients in autosomal recessive mode and is not yet reported in any population-specific genetic databases. AS patients carrying this mutation showed heterogeneity in clinical presentations. Computational analysis of the mutant centroid structure of ALMS1 mRNA revealed that exon-19 skipping enlarges the hairpin loop and decreases the free energy, eventually affecting its folding pattern, stability, and function. Hence, we propose c.11873-2A as an AS causative potential founder mutation in Saudi Arabia because it is found in two families lacking a common lineage. Conclusions: We conclude that WES analysis potentially helps in clinical phenotyping, early diagnosis, and better clinical management of Alström patients showing variable clinical expressivity.

IGFBPs mediate IGF-1's functions in retinal lamination and photoreceptor development during pluripotent stem cell differentiation to retinal organoids.

Development of the retina is regulated by growth factors, such as insulin-like growth factors 1 and 2 (IGF-1/2), which coordinate proliferation, differentiation, and maturation of the neuroepithelial precursors cells. In the circulation, IGF-1/2 are transported by the insulin growth factor binding proteins (IGFBPs) family members. IGFBPs can impact positively and negatively on IGF-1, by making it available or sequestering IGF-1 to or from its receptor. In this study, we investigated the expression of IGFBPs and their role in the generation of human retinal organoids from human pluripotent stem cells, showing a dynamic expression pattern suggestive of different IGFBPs being used in a stage-specific manner to mediate IGF-1 functions. Our data show that IGF-1 addition to culture media facilitated the generation of retinal organoids displaying the typical laminated structure and photoreceptor maturation. The organoids cultured in the absence of IGF-1, lacked the typical laminated structure at the early stages of differentiation and contained significantly less photoreceptors and more retinal ganglion cells at the later stages of differentiation, confirming the positive effects of IGF-1 on retinal lamination and photoreceptor development. The organoids cultured with the IGFBP inhibitor (NBI-31772) and IGF-1 showed lack of retinal lamination at the early stages of differentiation, an increased propensity to generate horizontal cells at mid-stages of differentiation and reduced photoreceptor development at the later stages of differentiation. Together these data suggest that IGFBPs enable IGF-1's role in retinal lamination and photoreceptor development in a stage-specific manner.

Exploring celiac disease candidate pathways by global gene expression profiling and gene network cluster analysis.

Celiac disease (CeD) is a gastrointestinal autoimmune disorder, whose specific molecular basis is not yet fully interpreted. Therefore, in this study, we compared the global gene expression profile of duodenum tissues from CeD patients, both at the time of disease diagnosis and after two years of the gluten-free diet. A series of advanced systems biology approaches like differential gene expression, protein-protein interactions, gene network-cluster analysis were deployed to annotate the candidate pathways relevant to CeD pathogenesis. The duodenum tissues from CeD patients revealed the differential expression of 106 up- and 193 down-regulated genes. The pathway enrichment of differentially expressed genes (DEGs) highlights the involvement of biological pathways related to loss of cell division regulation (cell cycle, p53 signalling pathway), immune system processes (NOD-like receptor signalling pathway, Th1, and Th2 cell differentiation, IL-17 signalling pathway) and impaired metabolism and absorption (mineral and vitamin absorptions and drug metabolism) in celiac disease. The molecular dysfunctions of these 3 biological events tend to increase the number of intraepithelial lymphocytes (IELs) and villous atrophy of the duodenal mucosa promoting the development of CeD. For the first time, this study highlights the involvement of aberrant cell division, immune system, absorption, and metabolism pathways in CeD pathophysiology and presents potential novel therapeutic opportunities.

Identification of Causative Variants Contributing to Nonsyndromic Orofacial Clefts Using Whole-Exome Sequencing in a Saudi Family.

Objectives: Nonsyndromic orofacial clefts (NSOFCs) are the most common craniofacial malformations observed across the globe. They are classified into three types: (a) cleft palate, (b) cleft lip, and (c) cleft lip and palate. To identify the potential candidate genes contributing to polygenic diseases such as NSOFC, linkage analyses, genome-wide association studies, and genomic rearrangements can be used. Genomic analyses, based on massively parallel next-generation sequencing technologies, play a vital role in deciphering the genetic bases of NSOFCs. Materials and Methods: In this study, whole exome sequencing was employed to detect genes that likely contributed to the NSOFC phenotype in a consanguineous Saudi family. Results: The exome analysis revealed NRP1 (rs35320960) as one potential candidate gene that is involved in bone differentiation. The RPL27A gene (rs199996172), which plays a crucial role in ribosome biogenesis, also passed all filters to serve as a candidate gene for NSOFC in this family. Rare variants are situated within the 5' UTR of these two genes. Conclusion: The study suggests that rare variants in NRP1 and RPL27A may be associated with NSOFC disease etiology.

Myocardial infarction biomarker discovery with integrated gene expression, pathways and biological networks analysis.

Myocardial infarction (MI) is the most prevalent coronary heart disease caused by the complex molecular interactions between multiple genes and environment. Here, we aim to identify potential biomarkers for the disease development and for prognosis of MI. We have used gene expression dataset (GSE66360) generated from 51 healthy controls and 49 patients experiencing acute MI and analyzed the differentially expressed genes (DEGs), protein-protein interactions (PPI), gene network-clusters to annotate the candidate pathways relevant to MI pathogenesis. Bioinformatic analysis revealed 810 DEGs. Their functional annotations have captured several MI targeting biological processes and pathways like immune response, inflammation and platelets degranulation. PPI network identify seventeen hub and bottleneck genes, whose involvement in MI was further confirmed by DisGeNET database. OpenTarget Platform reveal unique bottleneck genes as potential target for MI. Our findings identify several potential biomarkers associated with early stage MI providing a new insight into molecular mechanism underlying the disease.

A Novel Homozygous Frameshift Variant in DYM Causing Dyggve-Melchior-Clausen Syndrome in Pakistani Patients.

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.

Complement modulation reverses pathology in Y402H-retinal pigment epithelium cell model of age-related macular degeneration by restoring lysosomal function.

Age-related macular degeneration (AMD) is a multifactorial disease, which is characterized by loss of central vision, affecting one in three people by the age of 75. The Y402H polymorphism in the complement factor H (CFH) gene significantly increases the risk of AMD. We show that Y402H-AMD-patient-specific retinal pigment epithelium (RPE) cells are characterized by a significant reduction in the number of melanosomes, an increased number of swollen lysosome-like-vesicles with fragile membranes, Cathepsin D leakage into drusen-like deposits and reduced lysosomal function. The turnover of C3 is increased significantly in high-risk RPE cells, resulting in higher internalization and deposition of the terminal complement complex C5b-9 at the lysosomes. Inhibition of C3 processing via the compstatin analogue Cp40 reverses the disease phenotypes by relieving the lysosomes of their overburden and restoring their function. These findings suggest that modulation of the complement system represents a useful therapeutic approach for AMD patients associated with complement dysregulation.

Whole exome sequencing of a Saudi family and systems biology analysis identifies CPED1 as a putative causative gene to Celiac Disease.

Celiac disease (CD) is a gastrointestinal disorder whose genetic basis is not fully understood. Therefore, we studied a Saudi family with two CD affected siblings to discover the causal genetic defect. Through whole exome sequencing (WES), we identified that both siblings have inherited an extremely rare and deleterious CPED1 genetic variant (c.241 A > G; p.Thr81Ala) segregating as autosomal recessive mutation, suggesting its putative causal role in the CD. Saudi population specific minor allele frequency (MAF) analysis has confirmed its extremely rare prevalence in homozygous condition (MAF is 0.0004). The Sanger sequencing analysis confirmed the absence of this homozygous variant in 100 sporadic Saudi CD cases. Genotype-Tissue Expression (GTEx) data has revealed that CPED1 is abundantly expressed in gastrointestinal mucosa. By using a combination of systems biology approaches like protein 3D modeling, stability analysis and nucleotide sequence conservation analysis, we have further established that this variant is deleterious to the structural and functional aspects of CPED1 protein. To the best of our knowledge, this variant has not been previously reported in CD or any other gastrointestinal disease. The cell culture and animal model studies could provide further insight into the exact role of CPED1 p.Thr81Ala variant in the pathophysiology of CD. In conclusion, by using WES and systems biology analysis, present study for the first-time reports CPED1 as a potential causative gene for CD in a Saudi family with potential implications to both disease diagnosis and genetic counseling.

Whole exome sequencing identifies rare biallelic ALMS1 missense and stop gain mutations in familial Alström syndrome patients.

Alström syndrome (AS, OMIM ID 203800) is a rare childhood multiorgan disorder, which is widely studied in non-Arab ethnic patients. The clinical and molecular basis of AS and the mode of disease inheritance in consanguineous Arab populations is not well investigated. Therefore, to identify the molecular basis of AS in familial forms, the present study performed whole exome sequencing of 5 AS patients belonging to 2 different Bedouin families from Saudi Arabia. The present study identified the AS causative rare biallelic mutations in ALMS gene:T376S in exon 5 and S909* in exon 8 for family A and an R2721* in exon 10 (R2721*) for family B. ALMS1 targeted genetic sequencing of healthy population controls and family members has confirmed its extremely rare frequency and autosomal recessive mode of inheritance. The truncating mutations S909* and R2721* could cause the loss of CC domains and ALMS motif on C-terminal end of the protein and creates unstable protein, which eventually undergoes intracellular degradation. The premature protein truncating mutations described in our study may eventually provide further insight into the functional domains of the ALMS1 protein and contribute to the understanding of the phenotypic spectrum of AS. Whole exome sequencing based molecular diagnosis is expected to rule out ambiguity surrounding clinical diagnosis of suspected AS cases.