The cyclic nucleotide phosphodiesterase of Dictyostelium discoideum: the structure of the gene and its regulation and role in development

The cyclic nucleotide phosphodiesterase (phosphodiesterase) of Dictyostelium discoideum plays an essential role in development by hydrolyzing the cAMP used as a chemoattractant by aggregating cells. We have studied the biochemistry of the phosphodiesterase and a functionally related protein, the phosphodiesterase inhibitor protein, and have cloned the cognate genes. A 1.8-kb and a 2.2-kb mRNA are transcribed from the single-phosphodiesterase gene. The 2.2-kb mRNA comprises the majority of the phosphodiesterase mRNA found in differentiating cells and is transcribed only during development from a promoter at least 2.5 kb upstream of the translational start site. The 1.8-kb phosphodiesterase mRNA is detected at all stages of growth and development, is present at lower levels than the developmentally induced mRNA, and is transcribed from a site proximal to the protein-coding region. The phosphodiesterase gene contains a minimum of three exons, and a 2.3-kb intron, the longest yet reported for this organism. We have shown that the pdsA gene and four fgd genes affect the accumulation of the phosphodiesterase mRNAs, and we believe that these loci represent a significant portion of the genes regulating expression of the phosphodiesterase. The phosphodiesterase gene was introduced into cells by transformation and used as a tool to explore the effects of cAMP on the terminal stages of development. In cells expressing high levels of phosphodiesterase activity, final morphogenesis cannot be completed, and differentiated spore and stalk cells do not form. We interpret these results to support the hypothesis that cAMP plays an essential role in organizing cell movements in late development as well as in controlling the aggregation of cells in the initial phase of the developmental program.     

A functional splice site in the 5'' untranslated region of a zein gene.

Zein genes, the genes coding for the zein storage proteins of maize, have a unique gene structure where at least two promoters lie upstream of the coding region. Between the P1 promoter (900 base pairs upstream of the coding region) and the translation initiation AUG codon are 18 short reading frames. A discrepancy between the signals obtained by S1-mapping and primer extension and the DNA sequence in the region of one of these signals suggests the presence of a 3' splice site lying 40 nucleotides upstream of the coding region. A splicing event removing all of the short reading frames from the mRNA transcribed from the P1 promoter would bring this mRNA into a translatable form. Further evidence for a functional 3' splice site has been obtained from sequencing of primer extension products and in vitro splicing of a hybrid intron in the HeLa cell in vitro splicing system.