The role of transposable elements in neurological disorders

Transposable elements (TEs) are active in neuronal cells raising the question whether TE insertions contribute to risk of neuropsychiatric disease. While genome wide association studies (GWAS) serve as a tool to discover genetic loci associated with neuropsychiatric diseases, unfortunately GWAS do not directly detect structural variants such as TEs.  To examine the role of TEs in psychiatric and neurologic disease we evaluated 17,000 polymorphic TEs and find 76 are in linkage disequilibrium with disease haplotypes (P < 10−6) defined by GWAS.  From these 76 polymorphic TEs we identify causal candidates based on having insertions in genomic regions of regulatory chromatin and on having associations with altered gene expression in brain tissues. We show that lead candidate insertions have regulatory effects on gene expression in human neural stem cells altering the activity of a minimal promoter. Taken together, we identify 10 polymorphic TE insertions that are potential candidates on par with other variants for having a causal role in neurologic and psychiatric disorders (Ahn HW, Worman ZF, Lechsinska A, Payer LM, Wang T, Malik N, Li W, Burns KH, Nath A, Levin HL. Retrotransposon insertions associated with risk of neurologic and psychiatric diseases. EMBO Rep. 2022 Nov 11:202255197).

Identification of an integrase-independent pathway of retrotransposition

Retroviruses and Long terminal repeat (LTR) retrotransposons rely on integrase (IN) to insert their copy DNA (cDNA) into the genome of host cells. Nevertheless, in the absence of IN, retroelements can retain significant levels of insertion activity. We have characterized the IN-independent pathway of Tf1 and found insertion sites had homology to the primers of reverse transcription which form single stranded DNAs at the termini of the cDNA. In the absence of IN activity a similar bias was observed with HIV-1. Our studies showed the Tf1 insertions result from single strand annealing (SSA), a non-canonical form of homologous recombination mediated by Rad52. By expanding our analysis of insertions to include repeat sequences we found most formed tandem elements by inserting at pre-existing copies of a related transposable element. Surprisingly, we found wild type Tf1 uses the IN-independent pathway as an alternative mode of insertion (Li F, Lee M, Esnault C, Wendover K, Guo Y, Atkins P, Zaratiegui M, Levin HL. Identification of an integrase-independent pathway of retrotransposition. Sci Adv. 2022. PMID: 35767609).

Dense Transposon Integration Reveals Essential Cleavage and Polyadenylation Factors Promote Heterochromatin Formation

Heterochromatin functions as a scaffold for factors responsible for gene silencing and chromosome segregation. Heterochromatin can be assembled by multiple pathways, including RNAi and RNA surveillance. We identified factors that form heterochromatin using dense profiles of transposable element integration in Schizosaccharomyces pombe. The candidates include a large number of essential proteins such as four canonical mRNA cleavage and polyadenylation factors. We find that Iss1, a subunit of the poly(A) polymerase module, plays a role in forming heterochromatin in centromere repeats that is independent of RNAi. Genome-wide maps reveal that Iss1 accumulates at genes regulated by RNA surveillance. Iss1 interacts with RNA surveillance factors Mmi1 and Rrp6, and importantly, Iss1 contributes to RNA elimination that forms heterochromatin at meiosis genes. Our profile of transposable element integration supports the model that a network of mRNA cleavage and polyadenylation factors coordinates RNA surveillance, including the mechanism that forms heterochromatin at meiotic genes (Lee SY, Hung, S, Esnault C, Pathak R, Johnson KR, Kankole O, Yamashita A, Zhang H, Levin HL, (2020), Dense transposon integration reveals essential cleavage and polyadenylation factors promote heterochromatin formation, Cell Reports, 25;30(8)2686-2698).

Transposable element insertions in fission yeast drive adaptation to environmental stress

Cells are regularly exposed to a range of naturally occurring stress that can restrict growth or cause lethality. In response, cells activate expression networks with hundreds of genes that together increase resistance to common environmental insults. However, stress response networks can be insufficient to ensure survival, which raises the question of whether cells possess genetic programs that can promote adaptation to novel forms of stress. We found transposable element (TE) mobility in Schizosaccharomyces pombe was greatly increased when cells were exposed to unusual forms of stress such as heavy metals, caffeine, and the plasticizer phthalate. By subjecting TE-tagged cells to CoCl2, we found the TE integration provided the major path to resistance. Groups of insertions that provided resistance were linked to TOR regulation and metal response genes. We extended our study of adaptation by analyzing TE positions in 57 genetically distinct wild strains. The genomic positions of 1048 polymorphic LTRs were strongly associated with a range of stress response genes, indicating TE integration promotes adaptation in natural conditions. These data provide strong support for the idea, first proposed by Barbara McClintock, that TEs provide a system to modify the genome in response to stress (Esnault C, Lee M, Ham C, Levin HL, 2019, Transposable element insertion in fission yeast drives adaptation to environmental stress, Genome Research, 29:85-95).

LEDGF/p75 interacts with mRNA splicing factors and targets HIV-1 integration to highly spliced genes

The host chromatin-binding factor LEDGF/p75 interacts with HIV-1 integrase and directs integration to active transcription units. To understand how LEDGF/p75 recognizes transcription units, we sequenced 1 million HIV-1 integration sites isolated from cultured HEK293T cells. Analysis of integration sites showed that cancer genes were preferentially targeted, raising concerns about using lentivirus vectors for gene therapy. Additional analysis led to the discovery that introns and alternative splicing contributed significantly to integration site selection. These correlations were independent of transcription levels, size of transcription units, and length of the introns. Multivariate analysis with five parameters previously found to predict integration sites showed that intron density is the strongest predictor of integration density in transcription units. Analysis of previously published HIV-1 integration site data showed that integration density in transcription units in mouse embryonic fibroblasts also correlated strongly with intron number, and this correlation was absent in cells lacking LEDGF. Affinity purification showed that LEDGF/p75 is associated with a number of splicing factors, and RNA sequencing (RNA-seq) analysis of HEK293T cells lacking LEDGF/p75 or the LEDGF/p75 integrase-binding domain (IBD) showed that LEDGF/p75 contributes to splicing patterns in half of the transcription units that have alternative isoforms. Thus, LEDGF/p75 interacts with splicing factors, contributes to exon choice, and directs HIV-1 integration to transcription units that are highly spliced (Singh, P., Plumb, M., Ferris, A., Iben, J., Wu, X., Fadel, H., Esnault, C., Poeschla, E., Hughes, S., Kvaratskhelia, M., and Levin, H. L., 2015, LEDGF/p75 interacts with mRNA splicing factors and targets HIV-1 integration to highly spliced genes, Genes & Development, 29:2287-97).

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