Retrotransposons comprise roughly 1/3 of the human genome. Over evolutionary time, they have played an important role in shaping the genome through a number of mechanisms. We now know that L1 elements are the master retrotransposons of mammalian genomes. They can insert into genes causing disease, lead to deletions and duplications through mispairing and homologous crossing over, transduce flanking sequences during retrotransposition events, and mediate the retrotransposition of non-autonomous retroelements. In this proposal, we study the natural activity of retrotransposons in real time, using a mouse model of retrotransposition from a human L1 transgene. We will answer a number of key biological questions. Do the structural characteristics of retrotranspositions from the transgene match those of endogenous insertions? Is retrotransposition more efficient in male or female germ cells? Does age of the animal affect retrotransposition frequency in male germ cells? Do mechanisms exist that extinguish retrotransposition activity from a transgene after a number of generations? We will then optimize our transgene construct and breed animals containing the optimized constructs to homozygosity in order to increase retrotransposition frequency and produce a practical insertional mutagen in the mouse. We believe that an L1-based mutagenesis system that obviates the treatment of ES cells will be a valuable adjunct to chemical and other mutagenesis systems in determining gene function in mice. In another important aim, we pursue our discovery that SVA elements are L1-mediated, non-autonomous retrotransposons. We determine the important sequences in the SVA element for retrotransposition in the expectation that this knowledge will shed light on the mechanism by which L1s mobilize Alu elements. Thus, this focused proposal will provide answers to key questions of L1 biology and likely lead to a useful mutagenesis system for determination of gene function.
| 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 |
| Hancks, Dustin C; Kazazian Jr, Haig H (2012) Active human retrotransposons: variation and disease. Curr Opin Genet Dev 22:191-203 |
| Goodier, John L; Mandal, Prabhat K; Zhang, Lili et al. (2010) Discrete subcellular partitioning of human retrotransposon RNAs despite a common mechanism of genome insertion. Hum Mol Genet 19:1712-25 |
| Hancks, Dustin C; Kazazian Jr, Haig H (2010) SVA retrotransposons: Evolution and genetic instability. Semin Cancer Biol 20:234-45 |
| Rangwala, Sanjida H; Kazazian Jr, Haig H (2009) The L1 retrotransposition assay: a retrospective and toolkit. Methods 49:219-26 |
| Goodier, John L; Zhang, Lili; Vetter, Melissa R et al. (2007) LINE-1 ORF1 protein localizes in stress granules with other RNA-binding proteins, including components of RNA interference RNA-induced silencing complex. Mol Cell Biol 27:6469-83 |
| Kubo, Shuji; Seleme, Maria Del Carmen; Soifer, Harris S et al. (2006) L1 retrotransposition in nondividing and primary human somatic cells. Proc Natl Acad Sci U S A 103:8036-41 |
| Farley, Alexander H; Luning Prak, Eline T; Kazazian Jr, Haig H (2004) More active human L1 retrotransposons produce longer insertions. Nucleic Acids Res 32:502-10 |
| Deininger, Prescott L; Moran, John V; Batzer, Mark A et al. (2003) Mobile elements and mammalian genome evolution. Curr Opin Genet Dev 13:651-8 |
