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Biology of Human LINE-1 Retrotransposons

Research Summary

Dr. García Pérez studies how the activity of common human transposable elements (LINE-1 retrotransposons) affects our heritable genome. Current research in his lab uses pluripotent cells to learn how the host controls the activity of LINE-1 elements.

Transposable elements (TEs) are mobile pieces of DNA that can move ("jump") in our genome, thereby influencing a myriad of cellular processes. Long interspersed element-1 (LINE-1) retrotransposons comprise approximately 20 percent of the mammalian genome, and LINE-1 retrotransposition events can create genetic diversity through a variety of mechanisms. It is becoming increasingly evident that the activity of TEs—from acting as simple insertional mutagens to inducing other complex genomic alterations (e.g., mis-splicing and deletions of genomic DNA)—is a major force driving human genome evolution. However, despite the large contribution of TEs to human genome structure and their impact on human diseases, little is known about how the host regulates TEs and their genomic and epigenomic effects.

Most LINE-1 retrotransposition events in humans occur during early embryogenesis, when the genome is in a pluripotent state. Thus, it stands to reason that restriction mechanisms must exist in those cells to avoid high rates of mutagenesis in the germline genome. Previous data from our laboratory and others have demonstrated that human LINE-1 elements are naturally expressed in cells that mimic early embryogenesis. Our lab therefore uses an array of pluripotent cells to determine how the activity of LINE-1 is regulated by the host (with a specific focus on epigenetic mechanisms) and how the activity of LINE-1 elements affects the genome of pluripotent cells.

Epigenetic Control of LINE-1 Retrotransposition in Humans
In research I performed as a postdoctoral fellow (John V. Moran laboratory, University of Michigan) and in my lab in Spain, we have described the expression and mobilization of active LINE-1 elements in human embryonic stem cells (hESCs), supporting the hypothesis that heritable LINE-1 insertions accumulate during early embryogenesis. More recently, my lab generated and characterized a small panel of induced pluripotent stem cells (iPSCs). Because of the striking similarities between iPSCs and hESCs, we analyzed LINE-1 expression in iPSCs and parental somatic cells. Our data revealed that upon reprogramming, LINE-1 expression and retrotransposition increase in iPSCs relative to the parental cell line (a study in collaboration with the laboratories of John V. Moran and Warner Greene [Gladstone Institute]). My lab recently reported that unknown epigenetic mechanisms likely act to control the expression of LINE-1 elements in hESCs. Surprisingly, although LINE-1 elements are randomly distributed in the human genome, my lab demonstrated that expressed LINE-1 elements in hESCs are mainly located within human genes.

These data provide evidence that pluripotent cells control the expression of LINE-1 elements by epigenetic mechanisms other than DNA methylation. Thus, ongoing research in my lab involves reprogramming somatic cells to iPSCs to characterize epigenetic mechanisms that control LINE-1 expression in pluripotent cells. Because iPSCs reactivate the expression of LINE-1 in a relatively short period of time, we will combine genetics and genomics to identify mechanisms responsible for the epigenetic control of LINE-1 expression in pluripotent versus differentiated cells.

Retrotransposition is a complex process that requires host-encoded factors to succeed; conversely, other host-encoded factors likely act to regulate and restrict LINE-1 retrotransposition. However, most host factors that interact with LINE-1 elements during trafficking and insertion within a cell remain to be discovered. Last year, in collaboration with the lab of John V. Moran, we described that LINE-1 insertions in pluripotent cells are epigenetically silenced by histone modifications. Remarkably, LINE-1 silencing occurs in pluripotent cells and not in differentiated cells derived from them, indicating that such silencing may act as a restriction mechanism to reduce high rates of mutagenesis in our heritable genome. In addition, we demonstrated that there are two genetically separable pathways to (1) initiate LINE-1 silencing and (2) maintain the silenced status of a new LINE-1 insertion. Current research in my lab (in collaboration with John V. Moran's lab) aims to characterize this new cellular pathway that mediates the epigenetic control of LINE-1 elements in pluripotent cells.

More recently, my lab has obtained further data indicating that LINE-1-induced chromatin changes in pluripotent cells can spread to flanking genomic regions (i.e., acting as an epimutagen). However, nothing is known of the consequences that silencing a new LINE-1 insertion may have on pluripotent genomes. Thus, my lab is analyzing epigenetic alterations induced by a new LINE-1 insertion in pluripotent cells.

These studies will shed light on epigenetic mechanisms that control the fate and expression of mammalian retrotransposons in pluripotent cells, and will determine the extent of "epimutagenesis" associated with new LINE-1 mobilization events in our pluripotent genome.

Somatic LINE-1 Retrotransposition
It was long assumed that LINE-1 activity would be most prominent in places where new insertions would be transmitted to newborns, for example, in germ cells or during early human embryogenesis. However, that view changed in 2005 with the description of somatic LINE-1 activity in the rodent brain by the labs of John V. Moran and Fred H Gage (Salk Institute). Later, and in collaboration with the Gage lab, we demonstrated that human neuronal progenitor cells (NPCs) exhibit LINE-1 expression and support high levels of LINE-1 mobilization. Besides the potential contribution of LINE-1 activity to brain plasticity/behavior, these studies suggest an ongoing somatic LINE-1 retrotransposition load. However, we do not know which other somatic human cell types may be affected by the activity of LINE-1 elements.

Currently, my lab is taking advantage of the differentiation potential of hESCs to learn which other human somatic cells may accommodate LINE-1 retrotransposition. My lab is developing an inducible LINE-1 retrotransposition system, with the goal of differentiating cells to a specific lineage (ectodermal, endodermal, or mesodermal) and then activating retrotransposition. In addition, our long-term goal is to generate a human in vivo LINE-1 mobilization assay, exploiting the teratogenic properties of hESCs and iPSCs in immunocompromised mice. In sum, we will inspect human LINE-1 mobilization in different germ cell layers in vivo.

LINE-1 Retrotransposition and Human Diseases
We are just starting to discover the contribution of LINE-1 activity in several human diseases (cancer, Rett syndrome, and others). While I was a postdoctoral fellow in the laboratory of John V. Moran, we demonstrated that an alternative pathway of LINE-1 insertion could occur with high frequency in DNA-repair-deficient cell lines. This alternative pathway of LINE-1 retrotransposition, termed endonuclease-independent (ENi) retrotransposition, is likely an ancient mechanism to repair DNA and has elucidated intriguing similarities between telomerase and LINEs. Notably, ENi retrotransposition can lead to LINE-1 insertions with uncommon structures, often involving major alterations at the insertion site.

Current research in my lab is examining whether other DNA-repair deficiencies may be associated with deregulation of the ENi pathway. Because most new LINE-1 insertions accumulate during early embryogenesis, we are using a reprogramming strategy to generate iPSCs from human cells known to have deficiencies in DNA repair pathways (e.g., non-homologous end joining and homologous recombination pathways) to analyze the impact of LINE-1 retrotransposition in defined human diseases. Because most LINE-1 insertions occur during development, we hypothesize that using DNA-repair-deficient pluripotent cells is both informative and physiologically relevant to decipher the potential impact of LINE-1 activity on a subset of human genetic diseases.

Grants from the Spanish Ministry of Health, Junta de Andalucía government, and a Marie Curie IRG Action provided partial support for these projects.

As of January 17, 2012

Scientist Profile

International Early Career Scientist
Pfizer-University of Granada-Junta de AndalucÌa Center for Genomics and Oncological Research
Developmental Biology, Genetics