Functional genomic analysis of novel microdeletions and microduplications associated with intellectual disability

Intellectual disability (ID) is a diagnosis given to persons who have life-long cognitive and adaptive impairments that begin early in life. ID affects about 1-3% of the population. Extremely small chromosome losses and gains, called microdeletions and microduplications respectively (or collectively Copy Number Variants, CNVs), are the cause of ID in ~15% of cases and their identification has helped to pinpoint genomic regions that contain ID-genes. The objective of my PhD research was to search for ID candidate genes in subjects with ID, focusing on the functional genomic analysis of genes from CNVs and in the rest of the genome. I studied individuals with unique de novo pathogenic CNVs at chromosomal position 2p15-16.1 or with familial CNVs at chromosomal position 1q21.1. I used a multi-faceted approach that included the study of candidate genes’ 1) expression, 2) sequence variants, 3) knock down consequence in C. elegans and 4) imprinting potential. My results showed that the best candidate genes from the 2p15-16.1 CNV are XPO1, USP34 and REL because their expression is reduced in individuals with deletions. In case of the 1q21.1 CNV, I identified two candidate genes (CHD1L and PRKAB2) from the CNV that had altered expression and cellular function. I also identified a pathogenic sequence change in ATF6 in individuals with a familial 1q21.1 duplication. ATF6 is located outside the 1q21.1 CNV and is part of the Endoplasmic Reticulum (ER) stress response pathway which may contribute to the phenotypic variability in this family. Finally, I identified 3 CNVs in children with ID that overlap putative imprinted regions. The results of my study therefore led to the identification of genes which could contribute to ID as their function is altered in patients with the CNV or their characteristics suggest that they can be sensitive to copy number changes. This work contributes to an improved understanding of how CNVs and additional genetic changes in the rest of the genome can lead to ID.Medicine, Faculty ofPathology and Laboratory Medicine, Department ofGraduat

miRNA and miRNA target genes in copy number variations occurring in individuals with intellectual disability

Background: MicroRNAs (miRNAs) are a family of short, non-coding RNAs modulating expression of human protein coding genes (miRNA target genes). Their dysfunction is associated with many human diseases, including neurodevelopmental disorders. It has been recently shown that genomic copy number variations (CNVs) can cause aberrant expression of integral miRNAs and their target genes, and contribute to intellectual disability (ID). Results To better understand the CNV-miRNA relationship in ID, we investigated the prevalence and function of miRNAs and miRNA target genes in five groups of CNVs. Three groups of CNVs were from 213 probands with ID (24 de novo CNVs, 46 familial and 216 common CNVs), one group of CNVs was from a cohort of 32 cognitively normal subjects (67 CNVs) and one group of CNVs represented 40 ID related syndromic regions listed in DECIPHER (30 CNVs) which served as positive controls for CNVs causing or predisposing to ID. Our results show that 1). The number of miRNAs is significantly higher in de novo or DECIPHER CNVs than in familial or common CNV subgroups (P < 0.01). 2). miRNAs with brain related functions are more prevalent in de novo CNV groups compared to common CNV groups. 3). More miRNA target genes are found in de novo, familial and DECIPHER CNVs than in the common CNV subgroup (P < 0.05). 4). The MAPK signaling cascade is found to be enriched among the miRNA target genes from de novo and DECIPHER CNV subgroups. Conclusions Our findings reveal an increase in miRNA and miRNA target gene content in de novo versus common CNVs in subjects with ID. Their expression profile and participation in pathways support a possible role of miRNA copy number change in cognition and/or CNV-mediated developmental delay. Systematic analysis of expression/function of miRNAs in addition to coding genes integral to CNVs could uncover new causes of ID.Medical Genetics, Department ofMedicine, Faculty ofPathology and Laboratory Medicine, Department ofOther UBCReviewedFacult

Identifying candidate genes for 2p15p16.1 microdeletion syndrome using clinical, genomic, and functional analysis

The 2p15p16.1 microdeletion syndrome has a core phenotype consisting of intellectual disability, microcephaly, hypotonia, delayed growth, common craniofacial features, and digital anomalies. So far, more than 20 cases of 2p15p16.1 microdeletion syndrome have been reported in the literature; however, the size of the deletions and their breakpoints vary, making it difficult to identify the candidate genes. Recent reports pointed to 4 genes (XPO1, USP34, BCL11A, and REL) that were included, alone or in combination, in the smallest deletions causing the syndrome. Here, we describe 8 new patients with the 2p15p16.1 deletion and review all published cases to date. We demonstrate functional deficits for the above 4 candidate genes using patients’ lymphoblast cell lines (LCLs) and knockdown of their orthologs in zebrafish. All genes were dosage sensitive on the basis of reduced protein expression in LCLs. In addition, deletion of XPO1, a nuclear exporter, cosegregated with nuclear accumulation of one of its cargo molecules (rpS5) in patients’ LCLs. Other pathways associated with these genes (e.g., NF-κB and Wnt signaling as well as the DNA damage response) were not impaired in patients’ LCLs. Knockdown of xpo1a, rel, bcl11aa, and bcl11ab resulted in abnormal zebrafish embryonic development including microcephaly, dysmorphic body, hindered growth, and small fins as well as structural brain abnormalities. Our multifaceted analysis strongly implicates XPO1, REL, and BCL11A as candidate genes for 2p15p16.1 microdeletion syndrome