Date of Award:

5-1-1994

Document Type:

Thesis

Degree Name:

Master of Science (MS)

Department:

Biology

Department name when degree awarded

Biology (Molecular Biology)

Committee Chair(s)

Gregory J. Podgorski

Committee

Gregory J. Podgorski

Committee

Elizabeth Hood

Committee

Dennis Welker

Committee

Joseph K. K. Li

Abstract

The Dictyostelium discoideum cyclic nucleotide phosphodiesterase (PDE) is synthesized during growth and at different developmental stages and is controlled by three stage-specific promoters. The aggregation-specific promoter is induced by extracellular 3', 5' -cyclic adenosine monophosphate (cAMP) and is developmentally regulated. Regulatory elements of the aggregation-specific promoter were analyzed by serial deletions and by subcloning promoter segments. The full-length promoter was found to consist of disperse and redundant developmental regulatory elements. A region -556 to -450 upstream of the transcription start site is critical for extracellular cAMP-mediated expression. This region was used as a probe in gel shift assays to detect sequence-specific DNA/protein interactions. This 106-bp sequence is bound by nuclear proteins that are synthesized de novo during development. The level of binding activity is enhanced significantly by cAMP treatment. Treatment of nuclear extracts with phosphatase indicated that protein phosphorylation is essential for DNA binding activity. An element with the sequence of 5'-TGTGTGTGT-3' within the 106-bp cAMP responsive region of the pde promoter is homologous to the identified CRE of many Dictyostelium genes with a rough consensus (T/G)G(G/T)G(T/G)G(T/G).

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