dictyNews
Electronic Edition
Volume 39, number 35
December 13, 2013

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=========
Abstracts
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The cyclic AMP phosphodiesterase RegA critically regulates encystation in social 
and pathogenic amoebas.

Du Q, Schilde C, Birgersson E, Chen ZH, McElroy S, Schaap P.

College of Life Sciences, University of Dundee, Dundee DD15EH, UK.


Cell Signal. 2013 Oct 31. pii: S0898-6568(13)00319-7. 
doi: 10.1016/j.cellsig.2013.10.008. [Epub ahead of print]

Amoebas survive environmental stress by differentiating into encapsulated cysts. 
As cysts, pathogenic amoebas resist antibiotics, which particularly counteracts 
treatment of vision-destroying Acanthamoeba keratitis. Limited genetic tractability 
of amoeba pathogens has left their encystation mechanisms unexplored. The social 
amoeba Dictyostelium discoideum forms spores in multicellular fruiting bodies to 
survive starvation, while other dictyostelids, such as Polysphondylium pallidum can 
additionally encyst as single cells. Sporulation is induced by cAMP acting on PKA, 
with the cAMP phosphodiesterase RegA critically regulating cAMP levels. We show here
that RegA is deeply conserved in social and pathogenic amoebas and that deletion of 
the RegA gene in P. pallidum causes precocious encystation and prevents cyst 
germination. We heterologously expressed and characterized Acanthamoeba RegA and 
performed a compound screen to identify RegA inhibitors. Two effective inhibitors 
increased cAMP levels and triggered Acanthamoeba encystation. Our results show that 
RegA critically regulates Amoebozoan encystation and that components of the cAMP 
signalling pathway could be effective targets for therapeutic intervention with 
encystation.


Submitted by Qingyou Du [q.du@dundee.ac.uk]
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A RabGAP Regulates Life-Cycle Duration via Trimeric G-protein Cascades in 
Dictyostelium discoideum

Hidekazu Kuwayama, Yukihiro Miyanaga, Hideko Urushihara and Masahiro Ueda.


PLOS ONE  in press

Background: The life-cycle of cellular slime molds comprises chronobiologically 
regulated processes. During the growth phase, the amoeboid cells proliferate at 
a definite rate. Upon starvation, they synthesize cAMP as both first and second 
messengers in signalling pathways and form aggregates, migrating slugs, and 
fruiting bodies, consisting of spores and stalk cells, within 24 h. In Dictyostelium 
discoideum, because most growth-specific events cease during development, 
proliferative and heterochronic mutations are not considered to be interrelated 
and no genetic factor governing the entire life-cycle duration has ever been 
identified. 
Methodology/Principal Findings: Using yeast 2-hybrid library screening, we isolated
a Dictyostelium discoideum RabGAP, Dd Rbg-3, as a candidate molecule by which the
Dictyostelium Galpha2 subunit directs its effects. Rab GTPase-activating protein, 
RabGAP, acts as a negative regulator of Rab small GTPases, which orchestrate the 
intracellular membrane trafficking involved in cell proliferation. Deletion mutants 
of Dd rbg-3 exhibited an increased growth rate and a shortened developmental period, 
while an overexpression mutant demonstrated the opposite effects. We also show that 
Dd Rbg-3 interacts with 2 Galpha2 subunits in an activity-dependent manner in vitro.
Furthermore, both human and Caenorhabditis elegans rbg-3 homologs complemented the 
Ddrbg-3Ðdeletion phenotype in D. discoideum, indicating that similar pathways may 
be generally conserved in multicellular organisms. 

Conclusions/Significance: Our findings suggest that Dd Rbg-3 acts as a key element 
regulating the duration of D. discoideum life-span potentially via trimeric 
G-protein cascades. 


Submitted by Hidekazu Kuwayama [hidekuwayama@biol.tsukuba.ac.jp]
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[End dictyNews, volume 39, number 35]