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Human Molecular Genetics Advance Access published online on September 23, 2005
Human Molecular Genetics, doi:10.1093/hmg/ddi354
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© The Author 2005. Published by Oxford University Press. All rights reserved
Received June 16, 2005
Revised September 17, 2005
Accepted September 17, 2005

Article

Ribonucleoprotein particle formation is necessary but not sufficient for LINE-1 retrotransposition

Deanna A. Kulpa 1 and John V. Moran 2*

1 Departments of Human Genetics, University of Michigan Medical School, Ann Arbor, MI 48109-0618
2 Departments of Human Genetics, University of Michigan Medical School, Ann Arbor, MI 48109-0618; Departments of Internal Medicine, University of Michigan Medical School, 1241 E. Catherine, 4909 Buhl, Ann Arbor, MI 48109-0618

* To whom correspondence should be addressed.
John V. Moran, E-mail: moranj{at}umich.edu


  Abstract

LINE-1 elements (L1s) are abundant non-LTR retrotransposons that mobilize through an RNA intermediate by target site primed reverse transcription (TPRT). The L1-encoded proteins (ORF1p and ORF2p) preferentially associate with their encoding transcript to form a ribonucleoprotein particle (RNP), which is a proposed retrotransposition intermediate. Here, we have used epitope tagging to discriminate the proteins encoded by engineered L1s from those encoded by endogenously expressed L1s. We demonstrate that an L1 containing an epitope tag at the carboxyl terminus of ORF1p remains retrotransposition-competent, and that tagged ORF1p and its encoding RNA localize to cytoplasmic RNPs. We also identified two classes of ORF1p mutants, one that severely decreased RNP formation and blocked retrotransposition, and another that allows RNP formation but reduces retrotransposition by 100-fold. Thus, these data indicate that RNP formation is important but not sufficient for L1 retrotransposition, and suggest that ORF1p also may function at downstream steps in the L1 retrotransposition pathway.


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