Structural variation in Parkinson’s disease: Focusing on the role of Transposable elements in disease predisposition and pathogenesis

Parkinson’s disease (PD) is a neurodegenerative disorder with a complex aetiology including genetic risk factors, environmental exposure and aging. Recent genome wide association studies have been successful at identifying genetic variation that confers a risk for PD, yet despite this it is predicted that the large majority of the genetic attribution to the disease is still unknown. It is also noted that much of the identified risk loci lie within poorly annotated regions of the genome such as those containing repetitive sequences and transposable elements (TE)s, highlighting the importance of further investigation into such regions. Despite many reports that associate TE insertions with PD no study has comprehensively analysed the role of these elements in the disease. The work presented in this thesis sought to ask three main questions; first, are TE overrepresented at PD risk loci using a haplotype block based genome-wide analysis, second are non-reference TE associated with risk of PD using a newly developed TE detection tool and PD WGS data; and third, are TE differentially regulated in the blood or skin of individuals with PD. This work leveraged genetic and expression datasets to comprehensively address the role of TE in PD. Along with identifying that specific TE are overrepresented at PD risk loci we also show that in the blood, specific repetitive elements (satellite) are differentially expression in PD. Most significantly, we characterised known non-reference TE presence/absence polymorphisms in collaboration with the International Parkinson’s Disease Genomic Consortium (IPDGC) in PD whole genome sequencing data (WGS) from the Parkinson’s Progression Markers Initiative (PPMI) cohort using the TE detection tool MELT. We identify that TE insertions are a heritable and common form of genetic variation that lie within potentially important functional domains of the genome. Not only do many non-reference TE map to PD risk loci, but from our initial study we have identified that non-reference TE’s are in moderate linkage disequilibrium with PD risk variants, and thus a candidate causal variant that warrant further study at these loci. In summary, TE insertions are a major source and often overlooked form of genetic variation in the human genome. Collectively the research presented in this thesis suggests that not only could integrating TE variants be a valuable and critical step forward for furthering our understanding of existing risk PD variants, but it could also be valuable for establishing new risk factors

Identification and characterization of polymorphic mobile elements (MEs) in humans

Retrotransposons are mobile elements (MEs) that propagate in a “copy and paste” fashion in the genomes via RNA intermediates. In the human genome, retrotransposons consist of long terminal repeats (LTRs), long interspersed elements (LINEs), short interspersed elements (SINEs), SINE-VNTR- Alus (SVAs), and processed pseudogenes (PPSGs), and they collectively contribute close to 50% of the genome. Some members of these MEs continue to undergo retrotransposition, thereby generating a type of structural variations (SVs) within and between human populations by the presence and absence of ME insertions at specific genomic locations. A large number of such polymorphic MEs have been previously reported and documented, including cases associated with diseases, but with limited sequence characterization and genotype analysis. In this study, we performed extensive computational analysis and compilation of polymorphic MEs from multiple sources. We focused on characterization of complete sequences representing the insertion alleles and pre-integration alleles of ME polymorphic loci, using methods including local sequence assembly based on rich personal genome sequence data for many entries. Further, we performed in silico genotyping and population distribution for these polymorphic MEs for 2600 human subjects representing 28 well recognized populations around the world, as well as phylogenetic analysis of these human subjects using these polymorphic MEs as markers. We identified a total of 4400 polymorphic MEs with full sequence characterization for both the pre-integration and insertion alleles. Among these, 1267 entries represent new insertions not previously documented in the Database of Retrotransposon Insertion Polymorphisms in humans (dbRIP), and 1777 entries represent ME insertions outside the current human reference genome. By individual populations and all samples as whole, all 5 ME types displayed a similar allele distribution pattern with the majority having an allele frequency at 0.5, while differences across ME types are also seen at the very low frequency range. Nevertheless, polymorphic MEs do show substantial geographic differentiation, with numerous continent-specific loci identified. Polymorphic ME-based clustering of human subjects seems to correlate well with what we know about the history and relationship of human populations, indicating the usefulness of polymorphic MEs as markers for studying human evolution. Furthermore, polymorphic MEs were found to participate in both coding and regulatory sequences, signifying their potential contribution to the phenotypic diversity present among human populations and individuals. In conclusion, polymorphic MEs represent a significant source of human genetic diversity with potentials on impacting the structure, function, and evolution of the human genome

Alu elements contain many binding sites for transcription factors and may play a role in regulation of developmental processes

BACKGROUND: The human genome contains over one million Alu repeat elements whose distribution is not uniform. While metabolism-related genes were shown to be enriched with Alu, in structural genes Alu elements are under-represented. Such observations led researchers to suggest that Alu elements were involved in gene regulation and were selected to be present in some genes and absent from others. This hypothesis is gaining strength due to findings that indicate involvement of Alu elements in a variety of functions; for example, Alu sequences were found to contain several functional transcription factor (TF) binding sites (BSs). We performed a search for new putative BSs on Alu elements, using a database of Position Specific Score Matrices (PSSMs). We searched consensus Alu sequences as well as specific Alu elements that appear on the 5 Kbp regions upstream to the transcription start site (TSS) of about 14000 genes. RESULTS: We found that the upstream regions of the TSS are enriched with Alu elements, and the Alu consensus sequences contain dozens of putative BSs for TFs. Hence several TFs have Alu-associated BSs upstream of the TSS of many genes. For several TFs most of the putative BSs reside on Alu; a few of these were previously found and their association with Alu was also reported. In four cases the fact that the identified BSs resided on Alu went unnoticed, and we report this association for the first time. We found dozens of new putative BSs. Interestingly, many of the corresponding TFs are associated with early markers of development, even though the upstream regions of development-related genes are Alu-poor, compared with translational and protein biosynthesis related genes, which are Alu-rich. Finally, we found a correlation between the mouse B1 and human Alu densities within the corresponding upstream regions of orthologous genes. CONCLUSION: We propose that evolution used transposable elements to insert TF binding motifs into promoter regions. We observed enrichment of biosynthesis genes with Alu-associated BSs of developmental TFs. Since development and cell proliferation (of which biosynthesis is an essential component) were proposed to be opposing processes, these TFs possibly play inhibitory roles, suppressing proliferation during differentiation

Transposable elements as hidden neuronal gene regulators in health and disease

In this thesis, we take several approaches to unravel the roles of transposable elements (TEs) and Krüppel-associated box (KRAB) domain zinc finger proteins (KZNFs) in gene regulation. By doing so, we aim to contribute to broadening the knowledge on the involvement of these mobile genetic elements and their co-evolved repressor proteins in health and disease. First, we provide insights into the gene-regulatory potential of TEs in different developing and adult brain regions, and assess if the enhancer activity of TEs is influenced by ageing and age-related neurodegenerative diseases. Next, we dive deeper into specific classes of TEs, the MER52 and SVA elements, and analyse the relationship with the KZNFs binding most abundantly to these elements: ZNF519 and ZNF91. Lastly, we focus on the involvement of structural variation within TEs in disease risk. We highlight the presence of structurally variable SVAs (SV-SVAs) in neurological disease-associated loci, and assess their association to disease-associated SNPs and differential gene expression. Thereby we reveal a novel layer of genetic variation in transposable elements that may contribute to identification of the structural variants that are the actual drivers of disease associations of GWAS loci

The role of mobile elements in recent primate genomes

Mobile elements (MEs), which constitute ~50% of the primate genomes, have contributed to both genome evolution and gene function as demonstrated by ample evidence discovered over the last few decades. The three studies in this thesis aims to provide a better understanding of the evolutionary profile and function of MEs in the primate genomes by taking a computational comparative genomics approach. The first study represents a comprehensive analysis of the differential ME transposition among primates via identification of species-specific MEs (SS-MEs) in eight primate genomes from the families of Hominidae and Cercopithecidae using a comparative genomics approach. In total, 230,855 SS-MEs are identified, which reveal striking differences in retrotransposition level in the eight primate genomes. The second study represents a more focused analysis for the identification of a new type of MEs, which we term “retro-DNA” for non-LTR retrotransposons derived from DNA transposons, in the recent primate genomes. By investigating biallelic DNA transposons that have both the insertion and pre-integration alleles in ten primate genomes, a total of 1,750 retro-DNA elements representing 750 unique insertion events are reported for the first time. The third study provides an analysis of the mechanism underlying the differential SINE transposition in the primate genomes. In this study, Alu profiles are compared and the Alu master copies are identified in six primate genomes in the Hominidae and Cercopithecidae groups. The results show that each lineage of the primates and each species owns a unique Alu profile exclusively defined by the AluY transposition activity, which is determined by the number of Alu master copies and their relative activity. Overall, work in this thesis provides new insights about MEs and their impact on the recent primate genomes by revealing differential ME transposition as an important mechanism in generating genome diversity among primate lineages and species through discovering a new type of MEs and preliminary analysis of the mechanism underlying the differential ME transposition among primates. Furthermore, taking advantage of the recently available primate genomes and transcriptomes data, the work in this thesis demonstrates the great potential of the comparative genomic approach in studying MEs in primate genomes

The structural, functional and evolutionary impact of transposable elements in Eukaryotes

Transposable elements (TEs) are nearly ubiquitous in eukaryotes. The increase in genomic data, as well as progress in genome annotation and molecular biology techniques, have revealed the vast number of ways mobile elements have impacted the evolution of eukaryotes. In addition to being the main cause of difference in haploid genome size, TEs have affected the overall organization of genomes by accumulating preferentially in some genomic regions, by causing structural rearrangements or by modifying the recombination rate. Although the vast majority of insertions is neutral or deleterious, TEs have been an important source of evolutionary novelties and have played a determinant role in the evolution of fundamental biological processes. TEs have been recruited in the regulation of host genes and are implicated in the evolution of regulatory networks. They have also served as a source of protein-coding sequences or even entire genes. The impact of TEs on eukaryotic evolution is only now being fully appreciated and the role they may play in a number of biological processes, such as speciation and adaptation, remains to be deciphered

Retrotransposons as drivers of Mammalian brain evolution

Retrotransposons, a large and diverse class of transposable elements that are still active in humans, represent a remarkable force of genomic innovation underlying mammalian evolution. Among the features distinguishing mammals from all other vertebrates, the presence of a neocor-tex with a peculiar neuronal organization, composition and connectivity is perhaps the one that, by affecting the cognitive abilities of mammals, contributed mostly to their evolutionary success. Among mammals, hominids and especially humans display an extraordinarily expanded cortical volume, an enrichment of the repertoire of neural cell types and more elaborate patterns of neuronal connectivity. Retrotransposon-derived sequences have recently been implicated in multiple layers of gene regulation in the brain, from transcriptional and post-transcriptional control to both local and large-scale three-dimensional chromatin organization. Accordingly, an increasing variety of neurodevelopmental and neurodegenerative conditions are being recognized to be associated with retrotransposon dysregulation. We review here a large body of recent studies lending support to the idea that retrotransposon-dependent evolutionary novelties were crucial for the emergence of mammalian, primate and human peculiarities of brain morphology and function