The goal of this project is to develop mariner transposable elements as genetic tools for arthropods, primarily insects, of medical importance. The impact of arthropod vectors of disease around the world is enormous, and novel approaches to their control are needed. These mariner transposons are potentially useful for gene tagging for cloning, for enhancer trapping, and most importantly as transformation vectors. There are currently no efficient transformation vectors available for any non- drosophilid arthropod, so ideas of developing genetic sexing or sterile male techniques are untestable. Imaginative ideas for developing vector- incompetent strains of mosquitoes are untenable without transformation systems, and mariners might even spread such desirable traits. Finally, basic research on the molecular genetics of insects is handicapped by the lack of these tools. Distantly related mariner elements were previously known from the fruit fly Drosophila mauritiana and the cecropia moth Hyalophora cecropia, and are of the short inverted terminal repeat type. The D. mauritiana element is capable of transforming D. melanogaster. Related mariner elements were discovered in other insects using PCR with degenerate primers designed to regions of amino acid conservation between the transposase genes of the fruit fly and cecropia moth elements. They are present in representative of most insect orders, including Anopheles gambiae, plus a centipede and a mite. From sequences of these PCR fragments there is evidence for recent horizontal transfer of two particular elements across orders of insects. The consensus amino acid sequence representing each of these horizontally transferred mariners is almost certain to be an active transposase that has already demonstrated functionality in the cellular environments of diverse insects. These consensus-encoding elements will be identified in species or generated by in vitro mutagenesis using PCR. In either case,""""""""marooned"""""""" constructs that have the consensus transposase gene flanked by its natural regulatory regions, but without inverted terminal repeats, will be created by PCR. These will be tested for transposase activity in Drosophila melanogaster first, by assaying ability to mobilize in trans a dominant marker gene flanked by appropriate mariner termini. Active constructs will then be tested for activity in target species of medical importance, such as various mosquitoes. In these species other dominant markers being identified by workers on particular species will be needed, and methods for mechanical introduction of DNA into embryos will have to be developed. Constructs that are active in any particular species are then immediately available as transformation vectors. In addition, if two or more distinct and potentially independent mariner transformation systems can be developed, they might be employed for gene tagging and enhancer trapping systems.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI033586-02
Application #
2068636
Study Section
Tropical Medicine and Parasitology Study Section (TMP)
Project Start
1994-01-01
Project End
1996-12-31
Budget Start
1995-01-01
Budget End
1995-12-31
Support Year
2
Fiscal Year
1995
Total Cost
Indirect Cost
Name
University of Illinois Urbana-Champaign
Department
Zoology
Type
Schools of Arts and Sciences
DUNS #
041544081
City
Champaign
State
IL
Country
United States
Zip Code
61820
Barry, Elizabeth G; Witherspoon, David J; Lampe, David J (2004) A bacterial genetic screen identifies functional coding sequences of the insect mariner transposable element Famar1 amplified from the genome of the earwig, Forficula auricularia. Genetics 166:823-33
Lampe, David J; Witherspoon, David J; Soto-Adames, Felipe N et al. (2003) Recent horizontal transfer of mellifera subfamily mariner transposons into insect lineages representing four different orders shows that selection acts only during horizontal transfer. Mol Biol Evol 20:554-62
Lampe, D J; Walden, K K; Robertson, H M (2001) Loss of transposase-DNA interaction may underlie the divergence of mariner family transposable elements and the ability of more than one mariner to occupy the same genome. Mol Biol Evol 18:954-61
Zhang, J K; Pritchett, M A; Lampe, D J et al. (2000) In vivo transposon mutagenesis of the methanogenic archaeon Methanosarcina acetivorans C2A using a modified version of the insect mariner-family transposable element Himar1. Proc Natl Acad Sci U S A 97:9665-70
Lampe, D J; Akerley, B J; Rubin, E J et al. (1999) Hyperactive transposase mutants of the Himar1 mariner transposon. Proc Natl Acad Sci U S A 96:11428-33
Rubin, E J; Akerley, B J; Novik, V N et al. (1999) In vivo transposition of mariner-based elements in enteric bacteria and mycobacteria. Proc Natl Acad Sci U S A 96:1645-50
Lampe, D J; Grant, T E; Robertson, H M (1998) Factors affecting transposition of the Himar1 mariner transposon in vitro. Genetics 149:179-87
Robertson, H M; Lampe, D J (1995) Recent horizontal transfer of a mariner transposable element among and between Diptera and Neuroptera. Mol Biol Evol 12:850-62
Robertson, H M; Lampe, D J (1995) Distribution of transposable elements in arthropods. Annu Rev Entomol 40:333-57