Regulation And Function Of Retroelements

Eunice Kennedy Shriver National Institute Of Child Health & Human Development

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Abstract

Adaptation to stress Cells are regularly challenged by environmental stress for which rapid responses are critical to their survival. To cope with adverse conditions, cells activate transient programs of transcription that alter expression of hundreds to thousands of genes. Specific and prewired transcription responses have evolved due to frequent exposure to a common set of external stresses. However, it is not clear how cells cope when confronted with environmental shock, defined here as novel stresses or conditions that are unusually prolonged or extreme for which existing responses are inadequate to support survival. Adaptation to environmental shock requires genetic alterations that would typically occur in regulatory regions. One potential solution is the hypothesis that transposable elements promote adaptation to stress. TEs are genetically diverse mobile sequences that have proliferated extensively throughout eukaryotic genomes. These mobile elements respond to stress and through integration alter genome structure. In her seminal studies of maize, Barbara McClintock found TE mobility allows them to enter and take over control of genes. TEs carry regulatory elements that can control genes by altering their transcription and splicing. The high copy numbers of TEs, together with their regulatory sequences cause them to have a large role in shaping the transcriptome. The ability of TEs to impact transcription adds support to McClintocks proposal that TEs provide a means to overcome the threat of environmental shock by reorganizing the genome. Despite many reports that TEs are induced by stress, and examples of individual integration events that result in growth phenotypes, the tenet that TE mobility provides a program that overcomes environmental stress has not been directly tested. TEs of model organisms provide a unique opportunity to study the biological impact of active transposition in genetically tractable systems. Integration of the LTR retrotransposon Tf1 of Schizosaccharomyces pombe has been studied by determining as many as 1 million integration events with high throughput sequencing. The overwhelming majority of de novo insertions cluster in the nucleosome-depleted region of RNA polymerase pol II transcribed promoters. Importantly, the insertions are directed to approximately 1,000 promoters with an enrichment for stress response genes. A stress response enhancer imbedded in Tf1 causes integration to induce the expression of adjacent promoters. The prominent clustering of integration in promoters and the influence of the Tf1 enhancer on adjacent genes suggests the intriguing possibility that Tf1 may be wired to provide a genetic system for efficient adaptation to environmental stress. In this work we found TE expression and mobility in S. pombe was greatly increased when cells were exposed to unusual forms of stress such as heavy metals, caffeine, and the plasticizer phthalate. By subjecting cells with integration to CoCl2 we found the TE integration provided the major path to resistance. A substantial group of insertions that provided resistance were linked to TOR regulation and metal response genes. Studies of strains containing a single Tf1 insertion adjacent to TOR and metal response genes demonstrated that one integration event is sufficient to cause resistance to CoCl2. These single integration events altered the expression of the adjacent genes suggesting the Tf1 causes adaptation to stress by altering expression of genes capable of mitigating CoCl2 toxicity. We are extending these studies by addressing the impact of LTR-LTR recombination, the common recombination of the 5 and 3 LTRs of a retrotransposon that results in removing the element and leaving behind a solo LTR. The majority of LTR-retroelements undergo this process leaving behind the footprints of previous integration events. We are testing the possibility that this LTR-LTR recombination is a mechanism that resets the adaptation provided by an integration once the stress condition subsides. Retrotransposon insertions associated with risk of neurologic and psychiatric diseases Neurologic and psychiatric disorders affect 25% of the world population. Due to the complexity of the mammalian nervous system, the genetic and cellular etiology of these diseases remains largely unclear. Progress in genetic methodology has provided the potential to identify mechanisms that underlie these diseases. One approach that has successfully identified important disease loci are the genome-wide association studies (GWAS). However, in the cases of neurologic and major psychiatric disorders, GWAS have identified large numbers of loci each associated with small increases in risk. Importantly, there is extensive overlap of the loci that contribute to major psychiatric disorders indicating that related molecular mechanisms may underlie distinct clinical phenotypes. The SNPs identified by GWAS to have highest disease association (TASs; trait associated SNPs) are genetic tags that identify a genomic region that contain the causal mutation(s) leading to the disease risk. Limits on the design of GWAS typically prevents these studies from identifying causal gene alleles. Determining causal variants remains the most challenging and rate limiting, but also the most important step in defining the genetic architecture of diseases. The vast majority of GWAS TASs lie in intergenic or intronic regions and therefore do not alter coding sequence. For these SNPs to be causal they would likely have regulatory effects on transcription. Structural variants such as rearrangements, copy number variants, and TE insertions comprise a substantial and disproportionately large fraction of the genetic variants found to alter gene expression. In human, the dominant families of TEs are Long INterspersed Element-1 (LINE-1 or L1) and Alu Short Interspersed Elements (SINEs), which are mobilized by L1. TEs are particularly facile at altering gene expression because they have evolved various sequences that act on enhancers. Given that TEs make up approximately 45% of the human genome, it is not surprising that their regulatory features are abundant sources of tissue specific promoter activity. Relatively recent TE insertions can proliferate in the population and become common alleles. The 1000 Genomes Project described genetic variation of diverse human populations by sequencing whole genomes of 2,504 individuals. This extensive survey of genetic variation detected 17,000 polymorphic insertions of TEs. These polymorphic TEs have the potential to alter gene expression and affect common disease risk. Some TEs have been implicated at disease loci detected by GWAS. Given the difficulty in identifying genetic variants responsible for neurologic and psychiatric disorders and the regulatory capacity of TEs, we tested whether polymorphic TEs are potential causative variants of these diseases. We analyzed 593 GWAS of neurologic and psychiatric diseases which in total reported 753 TASs. From the 17,000 polymorphic TEs we found 76 were in linkage disequilibrium (LD) with TASs, indicating the TEs were among the variants with potential to be causative. We extended our analysis by evaluating each candidate TE for a role in altering expression of proximal genes. In one approach we determined whether polymorphic TEs could disrupt regulatory sequences as annotated with the epigenomic data of the NIH Roadmap Epigenomics Consortium. Ten of the TE candidates were located in regions of chromatin with regulatory function active in neurologic tissues. We also tested whether the polymorphic TEs were significantly associated with altered expression of proximal genes. By analyzing multi-tissue expression data from GTEx we found 31 of the TASs linked to TEs were expression quantitative trait loci (eQTLs) for adja