Intercellular signaling during terminal differentiation

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DESCRIPTION (provided by applicant): Communication between prespore and prestalk cells coordinates terminal differentiation in Dictyostelium discoideum. The signals include a 34 amino acid peptide, SDF-2, that has a sequence related to that of the neuropeptide DBI (Diazepam Binding Inhibitor). The neurotransmitter GABA is also used as a signal in Dictyostelium by acting through a G-protein coupled receptor similar to the GABAB receptor of mammals. The SDF-2 receptor, DhkA, on the other hand, is a "two-component" histidine kinase completely unrelated to the DBI receptors, GABAA and "peripheral type" BZ receptors. It seems that intercellular signals have been conserved over long periods of evolution while the receptors have changed in some cases. We will further elucidate the detailed mechanisms of intercellular signaling in this genetically tractable system since the results appear to have bearing on signaling in the mammalian central nervous system and may shed light on mechanisms underlying neuropsychiatric disorders. There is a complicated interchange of information between prespore and prestalk cells as they prepare to encapsulate and vacuolize, respectively. SDF-2 is generated by proteolytic processing of a protein, AcbA, released from prespore cells. Processing occurs on the surface of prestalk cells after priming by low levels of SDF-2 or by GABA. Prespore cells respond to priming by rapid release of AcbA from vesicles. AcbA is an acyl-CoA binding protein as is the precursor of DBI. Using site-directed mutagenesis we will determine whether this activity is essential for accumulation of AcbA in endosomes and subsequent release. We will determine whether exocytosis in response to either low levels of SDF-2 or GABA is dependent on signaling through the cAMP dependent protein kinase PKA. Extracellular AcbA is proteolytically cleaved by the membrane embedded prestalk specific protease TagC, which is only exposed following priming by either low levels of SDF-2 or GABA. We will determine whether this process is also dependent on PKA activity. Our results suggest that the precursor of the mammalian neuropeptide DBI, ACBP, may be synthesized in one cell type, stored in vesicles, and processed to the active peptides by another cell type. Such interactions could modulate the levels of this natural ligand that binds at the target of the widely used drug diazepam (Valium). Our observations that GABA appears to induces rapid exocytosis not only accounts for its priming activity in Dictyostelium but provides a test system for better understanding of the basic processes controlling exocytosis. Together these studies will add to the foundation for improvements in the diagnosis and treatment of neurological diseases.