The scientific community is still surprised by the mobile DNA found in the genomes of all organisms. In human beings, the master mobile DNA is the L1 retrotransposon which mobilizes itself along with Alu , SVA, and processed pseudogenes to new genomic locations. Since we found the first disease-causing insertions of an L1 in two patients with hemophilia A some 23 years ago, much has been learned about L1 and other human retrotransposons through cell culture assays, biochemical analyses, natural insertions into genes, transgenic animals, and bioinformatic analysis. Recently, PCR techniques, high-throughput sequencing and bioinformatic analysis have made it possible to discover essentially all non-reference retrotransposons in any human genome. A recent surprise finding of our lab was that in transgenic animals L1 retrotransposition (Rtn) predominately occurs in early embryonic development as opposed to the germ line. In this proposal, we propose three complementary Specific Aims that build on these findings. First, we study L1 Rtn in human biology by quantifying the somatic Rtn frequency in fetal and adult tissues and compare these frequencies to the frequency estimated for the germ line. We also determine the effects of new L1 insertions on the expression of nearby genes. Second, we have recently modified our technique to find essentially all non-reference, active mouse L1s. We use this procedure to study L1 Rtn in tissues of different mouse strains. Then, after direct determination of the germ line Rtn frequency in different mouse strains, we will compare the somatic and germ line frequencies. We expect that since the mouse has 15-30 times the number of active L1s as the human (1500-3000 vs. 80- 100), Rtn frequency in the mouse will be significantly greater than in humans. Early work has led to exciting new data, indicating that L1 insertions into the mouse genome occur non-randomly in clusters. As in the human work, we will then determine the effect of new mouse L1 insertions on gene expression. Third, we determine the role of Rtn in human disease, studying 1) various cancers whose DNA has been completely sequenced and 2) mutation-negative patients with a well-phenotyped Mendelian disorder, Cornelia de Lange Syndrome. In observing Rtn events in these patients, we will learn the extent to which they have been underestimated in human disease etiology. We predict that retrotransposon effects on human health are significantly greater than currently believed.

Public Health Relevance

L1 retrotransposons are the major mobile DNA of human and mammalian genomes. Here we study the effects of L1 in human health and disease. We search for somatic mutations in human and mice, determine the effects of retrotransposons on gene expression in humans and mice, and analyze both human cancers and Mendelian disorders for causative retrotransposition events.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Genetics of Health and Disease Study Section (GHD)
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Janes, Daniel E
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Johns Hopkins University
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Pereira, Gavin C; Sanchez, Laura; Schaughency, Paul M et al. (2018) Properties of LINE-1 proteins and repeat element expression in the context of amyotrophic lateral sclerosis. Mob DNA 9:35
Kazazian Jr, Haig H; Moran, John V (2017) Mobile DNA in Health and Disease. N Engl J Med 377:361-370
Richardson, Sandra R; Gerdes, Patricia; Gerhardt, Daniel J et al. (2017) Heritable L1 retrotransposition in the mouse primordial germline and early embryo. Genome Res 27:1395-1405
Kazazian Jr, Haig H (2017) Fifty years in human genetics--a career retrospective. FASEB J 31:3712-3718
Doucet, Tara T; Kazazian Jr, Haig H (2016) Long Interspersed Element Sequencing (L1-Seq): A Method to Identify Somatic LINE-1 Insertions in the Human Genome. Methods Mol Biol 1400:79-93
Doucet-O'Hare, Tara T; Sharma, Reema; Rodi?, Nemanja et al. (2016) Somatically Acquired LINE-1 Insertions in Normal Esophagus Undergo Clonal Expansion in Esophageal Squamous Cell Carcinoma. Hum Mutat 37:942-54
Mandal, Prabhat K; Kazazian Jr, Haig H (2016) Purification of L1-Ribonucleoprotein Particles (L1-RNPs) from Cultured Human Cells. Methods Mol Biol 1400:299-310
Goodier, John L; Pereira, Gavin C; Cheung, Ling E et al. (2015) The Broad-Spectrum Antiviral Protein ZAP Restricts Human Retrotransposition. PLoS Genet 11:e1005252
Ewing, Adam D; Gacita, Anthony; Wood, Laura D et al. (2015) Widespread somatic L1 retrotransposition occurs early during gastrointestinal cancer evolution. Genome Res 25:1536-45
Doucet-O'Hare, Tara T; Rodi?, Nemanja; Sharma, Reema et al. (2015) LINE-1 expression and retrotransposition in Barrett's esophagus and esophageal carcinoma. Proc Natl Acad Sci U S A 112:E4894-900

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