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Published on: June 18, 2016
Regulation of Dictyostelium early development genes in signal transduction mutants
1Department of Anatomy and Cell Biology, College of Physicians and Surgeons, Columbia University, New York, New York 10032, USA.
Abstract:
The secreted cyclic nucleotide phosphodiesterase (PDE) and its glycoprotein inhibitor (PDI) are among the first genes expressed when Dictyostelium amoebae begin their development. We used a series of mutants with defects in signal transduction to ask whether cAMP receptors 1 and 3, G alpha2, G beta, adenylyl cyclase (ACA), or the protein kinase A catalytic subunit (PKAcat) are required for the initial appearance or later regulation of the PDE and the PDI transcripts. The PDE gene produces a 1.9-kb transcript during vegetative growth and then a 2.4-kb transcript during the early hours of development. Regulation of the 2.4-kb transcript in CAR1, G alpha2, G beta, and ACA mutants is similar to that of isogenic parental strains, although its level is reduced in strains that carry the CAR1 mutation. CAR1/CAR3 double mutants also produce less PDE transcript, but the PDE gene remains inducible by cAMP. The PKAcat mutant produces the 2.4-kb PDE transcript, but in this mutant the vegetative transcript is retained in development. CAR1 and CAR3 are not required for transcription of the PDI gene, but deleting CAR1 leads to overproduction of the PDI transcript. Induction or repression of the PDI gene does not require G alpha2, G beta, or ACA. PKAcat is required for synthesis of the PDI transcript. The results suggest a two-stage regulation of these early genes through a G alpha2/G beta-independent mechanism and an absolute dependence of PDI on the PKAcat.
Insights
Dictyostelium development involves early gene expression of cyclic nucleotide phosphodiesterase (PDE) and its inhibitor (PDI). While PDE regulation is complex, PDI synthesis absolutely requires protein kinase A catalytic subunit (PKAcat).
Area of Science:
- Cellular and Molecular Biology
- Developmental Biology
- Biochemistry
Background:
- Early Dictyostelium development involves the coordinated expression of secreted cyclic nucleotide phosphodiesterase (PDE) and its glycoprotein inhibitor (PDI).
- Understanding the regulatory pathways governing these early developmental genes is crucial for deciphering the signaling cascades initiating multicellularity.
Purpose of the Study:
- To investigate the roles of specific signal transduction components, including cAMP receptors (CAR1, CAR3), G proteins (G alpha2, G beta), adenylyl cyclase (ACA), and protein kinase A catalytic subunit (PKAcat), in regulating PDE and PDI gene expression during Dictyostelium development.
Main Methods:
- Utilized a series of Dictyostelium mutants with defects in key signal transduction pathways.
- Analyzed transcript levels of PDE (1.9-kb and 2.4-kb) and PDI genes under various mutant conditions using Northern blotting or similar transcript analysis techniques.
- Assessed the inducibility and repression of PDE and PDI genes by cAMP in different genetic backgrounds.
Main Results:
- PDE gene regulation involves a G alpha2/G beta-independent mechanism, with CAR1 and CAR3 influencing transcript levels but not inducibility.
- PKAcat is essential for PDI transcript synthesis, while CAR1 and CAR3 are not required for PDI transcription, though CAR1 deletion leads to PDI overproduction.
- The vegetative PDE transcript is retained in PKAcat mutants, indicating a role for PKAcat in transcript processing or turnover.
Conclusions:
- Dictyostelium PDE and PDI gene expression during early development are differentially regulated by distinct signal transduction pathways.
- A two-stage regulatory mechanism, independent of G alpha2/G beta, controls early gene expression.
- Protein kinase A catalytic subunit (PKAcat) plays an indispensable role in the synthesis of the PDI transcript, highlighting its critical function in developmental signaling.
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