Date of Award:

5-1-1996

Document Type:

Dissertation

Degree Name:

Doctor of Philosophy (PhD)

Department:

Biology

Committee Chair(s)

Dennis L. Welker

Committee

Dennis L. Welker

Committee

Gregory J. Podgorski

Committee

Thomas A. Grover

Committee

Noelle E. Cockett

Committee

Reed P. Warren

Abstract

Transposable elements are DNA segments that possess the capability to move to new chromosomal or extrachromosomal locations. Tdd-4 is a newly isolated transposable element from Dictyostelium discoideum that was captured while studying maintenance properties of Dictyostelium plasmids. Approximately 20 copies are present in the D. discoideum AX4 genome. Two classes of elements were identified, which include a 3.8 kilobase full-length version and a 3.4 kilobase deleted version. Restriction fragment length polymorphism analysis demonstrates that Tdd-4 utilizes a cut-and-paste mode of transposition. Sequence analysis reveals that the 145 base pair inverted terminal repeats contain the 5'-TG...CA-3' conserved terminal dinucleotides found in prokaryotic transposons and retroviral/retrotransposon DNA sequences. Tdd-4 open reading frames are pieced together by removal of six introns. Introns 1-5 conform to the GT-AG rule, whereas intron 6 appears to be an AT-AA intron. Also, intron 6 undergoes an alternative 5' splicing reaction. The alternatively spliced region encodes 15 tandem SPXX repeats. These tetrapeptide repeats are postulated to form a DNA-binding motif. For other transposons that produce two proteins from the same gene, the full-length protein serves as the transposase and the truncated protein plays a role in the regulation of transposition. By analogy, the full-length Tdd-4 protein is the putative transposase and the truncated Tdd-4 protein is the putative transposition inhibitor. Protein database searches revealed that Tdd-4 encoded proteins contain regions similar to retroviral/retrotransposon integrases. Based on the deduced amino acid sequence, the structure of the putative Tdd-4 transposase appears to be the same as retroviral integrases, possessing an N-terminal HHCC domain, a central DDE motif, and a C-terminal DNA-binding domain composed of the SPXX motif. Forced expression of the Tdd-4 transposase and Tdd-4 inhibitor cDNAs did not demonstrate a biological activity in a variety of assays. DNA fragments that flank eight separate genomic Tdd-4 elements were sequenced and analyzed. Tdd-4 integration appears to elicit a 4-5 base pair target site duplication. DNA fragments that flank Tdd-4 elements preferentially hybridize to moderately repetitive regions in the D. discoideum AX4 genome.

Download

Share

COinS

Copyright for this work is retained by the student. If you have any questions regarding the inclusion of this work in the Digital Commons, please email us at .