Genetics and Gene Regulation Program

Address

405B Stellar Chance Labs
422 Curie Blvd
Philadelphia PA 19104-6100

Office tel.: 215 746-5768
Lab tel.: 215 746-5769
Fax: 215 573-6317
E-mail: luning@mail.med.upenn.edu


Education

Princeton University, A.B. (Molecular Biology),1988
 
University of Pennsylvania, Ph.D. (Immunology), 1996
 
University of Pennsylvania, M.D., 1996

Research Interests

• mobile DNA, rearrangement of immunoglobulin genes, regulation of L1 retrotransposition

Key words: transposon, mobile DNA, retrotransposition, RNA interference, gene delivery vector, antibody, V(D)J recombination, LINE1, L1, receptor editing, immunoglobulin, autoimmunity, lupus, repertoire analysis, CDR3 spectratyping

Search PubMed for articles

Description of Research

DNA can transpose or rearrange in somatic cells. We are interested in mechanisms by which DNA mobility is regulated in mammalian cells. The two types of mobile DNA that we study are antibody genes and L1 retrotransposons.

Antibodies are assembled by DNA recombination from gene segments in B cells. Receptor editing is a process by which B cells can modify pre-existing (already rearranged) antibodies by undergoing further DNA rearrangement. Receptor editing can rescue self-reactive B cells from clonal deletion. Revision of antibody genes may also occur in peripheral lymphoid organs, where it can rescue clones with deleterious somatic mutations. Receptor editing can create cells with multiple receptors (allelic inclusion).

We are interested in exploring the function and significance of edited B cells in the context of systemic autoimmunity. Using anti-DNA knock-in mice, we are exploring how antibody heavy and light chain editing contribute to the B cell repertoire. We have developed a series of molecular assays to characterize editing products and antibody repertoire in B cell populations and are using these assays to determine if receptor editing is a cause or a consequence of autoimmunity. We are also extending these studies to human subjects with autoimmune diseases such as systemic lupus erythematosus, type I diabetes and Sjogren’s syndrome.

L1 retrotransposons (LINE1 elements, L1s) are mobile DNA elements that replicate via an RNA intermediate. The human genome contains over 500,000 L1 insertions of which only approximately 80 can move. Highly active L1s could be harnessed for a variety of biomedical applications including gene delivery, insertional mutagenesis and cell lineage analysis. We use a transgenic mouse model of L1 retrotransposition with an active human L1 element tagged with green fluorescent protein (GFP). Our long-term aim is to study how L1 mobility is regulated in vivo. Several mechanisms have been postulated to limit L1 mobility including methylation, RNA interference (RNAi), alterations in DNA repair enzymes in the non-homologous end joining pathway and apoptosis. We are also pursuing whether states such as transformation, neoplasia, injury (by DNA damaging agents) or even normal aging could result in altered L1 activity.

Tracking a transposon in vivo
An active L1 retrotransposon (mobile element) was tagged with enhanced green fluorescent protein (EGFP). EGFP is only expressed when the tagged L1 element "jumps." The figure shows expression in freshly isolated testis of a mouse with an embryonic L1 insertion event. Some of the tubules in the testis contain the L1 insertion (and thus are green) while others lack the insertion. Genetic analysis of the offspring confirms that this mouse was mosaic for the L1 insertion. The image was created using a Leica MZFLIII stereofluorescent microscope outfitted with a 100 Watt mercury bulb and chroma filters (exciter HQ470/40; emitter 515nm LP) and imaged using a MagnaFire CCD camera and Adobe Photoshop.

Recent Publications

Colleen M. Doyle, Jiong Han, Martin Weigert and Eline T. Luning Prak: Consequences of receptor editing at the lambda locus: multireactivity and light chain secretion. Proc. Natl. Acad. Sci. USA in press 2006.

Sekiguchi, D., Yunk, L., Gary, D., Charan, D., Srivastava, B., Allman, D., Weigert, M. and E. Luning Prak: Development and Selection of Edited B cells in B6.56R Mice. J. Immunol 176(11): 6879-6887, June 2006.

Farkash, E.A., Kao, G.D., Horman, S.R. and E. T. Luning Prak.: Gamma radiation increases endonuclease-dependent L1 retrotransposition in a cultured cell assay. Nucleic Acids Research Vol. 34: 1196-1204, 2006.

Shane Horman, Petr Svoboda and Eline T. Luning Prak: Review: The potential regulation of L1 mobility by RNA interference. Journal of Biomedicine and Biotechnology in press 2006.

Lab

Rotation Projects

Please come visit the lab to discuss rotation projects in any of the areas discussed above. Projects will be tailored to the background and interests of the student.

Lab personnel:

Lenka Yunk, graduate student
Anil Panigrahi, graduate student
Sara Smith, graduate student
Jennifer Sutter, M.D., research fellow
Yang Zhu Du, Ph.D., research specialist
Noah Goodman, research specialist
Peffin Lee, research specialist
Andrew Fesnak, research specialist
Deep Charan, Penn undergraduate
Tushar Khanna, Penn undergraduate