Periodic biological variations reflect interactions among molecules and cells, or even organisms. The Dictyostelium cAMP oscillatory circuit is a highly robust example. cAMP oscillations in Dictyostelium arise intracellularly by a complex interplay of activating and inhibiting pathways, are transmitted extracellularly, and synchronize an entire local population. Once established, cAMP signal-relay persists stably for hours. On a two-dimensional surface, >100,000 cells may form a single coordinated territory. In suspension culture, >10 10 cells can oscillate in harmony. This review focuses on molecular mechanisms that cyclically activate and attenuate signal propagation and on chemotactic responses to oscillatory wave progression.

SUMMARY Presenilin (PS) is the catalytic moiety of the γ-secretase complex. PS and other γ-secretase components are well conserved among metazoa, but their presence and function in more-distant species are not resolved. Because inappropriate γ-secretase processing of amyloid precursor protein (APP) in humans is associated with familial Alzheimer’s disease, understanding essential elements within each γ-secretase component is crucial to functional studies. Diverged proteins have been identified in primitive plants but experiments have failed to demonstrate γ-secretase activity. We have identified highly diverged orthologs for each γ-secretase component in the ancient eukaryote Dictyostelium, which lacks equivalents of APP, Notch and other characterized PS/γ-secretase substrates. We show that wild-type (WT) Dictyostelium is capable of amyloidogenic processing of ectopically expressed human APP to generate amyloid-β peptides Aβ40 and Aβ42; strains deficient in γ-secretase cannot produce Aβ peptides but accumulate processed intermediates of APP that co-migrate with the C-terminal fragments α- and β-CTF of APP that are found in mammalian cells. We further demonstrate that Dictyostelium requires PS for phagocytosis and cell-fate specification in a cell-autonomous manner, and show that regulation of phagocytosis requires an active γ-secretase, a pathway suggested, but not proven, to occur in mammalian and Drosophila cells. Our results indicate that PS signaling is an ancient process that arose prior to metazoan radiation, perhaps independently of Notch. Dictyostelium might serve to identify novel PS/γ-secretase signaling targets and provide a unique system for high-throughput screening of small-molecule libraries to select new therapeutic targets for diseases associated with this pathway.

Cryphonectria parasitica, the causative agent of chestnut blight, has proven to be a tractable experimental system for studying fungal pathogenesis. Moreover, the development of infectious cDNA clones of C. parasitica hypoviruses, capable of attenuating fungal virulence, has provided the opportunity to examine molecular aspects of fungal plant pathogenesis in the context of biological control. In order to establish a genomic base for future studies of C. parasitica, the authors have analysed a collection of expressed sequences. A mixed cDNA library was prepared from RNA isolated from wild-type (virus-free) and hypovirus-infected C. parasitica strains. Plasmid DNA was recovered from individual transformants and sequenced from the 59 end of the insert. Contig analysis of the collected sequences revealed that they represented approximately 2200 individual ORFs. An assessment of functional diversity present in this collection was achieved by using the BLAST software utilities and the NCBI protein database. Candidate genes were identified with significant potential relevance to C. parasitica growth, development, pathogenesis and vegetative incompatibility. Additional investigations of a 12?9 kbp genomic region revealed microsynteny between C. parasitica and both Neurospora crassa and Magnaporthe grisea, two closely related fungi. These data represent the largest collection of sequence information currently available for C. parasitica and are now forming the basis of further studies using microarray analyses to determine global changes in transcription that occur in response to hypovirus infection.

Using an established spotted cDNA microarray platform, the nature of changes in the transcriptional profiles of 2200 unique genes from the chestnut blight fungus Cryphonectria parasitica in response to the absence of either the Ga subunit CPG-1 or the Gb subunit CPGB-1 has been explored. It is reported that 216 transcripts were altered in accumulation in the Dcpg-1 strain and 163 in the Dcpgb-1 strain, with a considerable overlap (100 genes) that were changed in both cases. Of note, these commonly altered transcripts were changed in the same direction in every instance, thus suggesting a considerable redundancy in pathway control or extensive crosstalk. To further knowledge of the potential impact on G-protein-signalling of infection by hypovirus CHV1-EP713, the accumulation of CPG-1 and CPGB-1 was also investigated by Western analysis. It was demonstrated that both signalling components were reduced in abundance to approximately 25 % of wild-type levels, while their transcripts were slightly elevated. Comparison of a list of genes with altered expression in the presence of CHV1-EP713 to the data obtained in the absence of either G-protein subunit showed that more than one-half of all the transcripts changed by hypovirus infection were also changed in at least one G-protein mutant strain, with one-third being changed in both. Significantly, 95 % of the co-changed genes were altered in the same direction. These data provide the first evidence for modulation of Gb protein levels as well as the Gbc-signalling pathways by hypovirus infection, and support the hypothesis that modification of G-protein-signalling via both Ga and Gbc provides for a significant contribution to hypovirus-mediated phenotype.

SUMMARYIn Dictyostelium, the interaction of secreted cAMP with specific cell surface receptors regulates the activation/de-activation of GSK3, which mediates developmental cell patterning. In addition, Dictyostelium cells polarize in response to extracellular cAMP, although a potential role for GSK3 in this pathway has not been investigated. Previously, we had shown that ZAK1 was an activating tyrosine kinase for GSK3 function in Dictyostelium and we now identify ZAK2 as the other tyrosine kinase in the cAMPactivation pathway for GSK3; no additional family members exist. We also now show that tyrosine phosphorylation/activation of GSK3 by ZAK2 and ZAK1 separately regulate GSK3 in distinct differentiated cell populations, and that ZAK2 acts in both autonomous and non-autonomous pathways to regulate these cell-type differentiations. Finally, we demonstrate that efficient polarization of Dictyostelium towards cAMP depends on ZAK1-mediated tyrosine phosphorylation of GSK3. Combinatorial regulation of GSK3 by ZAK kinases in Dictyostelium guides cell polarity, directional cell migration and cell differentiation, pathways that extend the complexity of GSK3 signaling throughout the development of Dictyostelium.