dictyNews Electronic Edition Volume 42, number 30 December 16, 2016 Please submit abstracts of your papers as soon as they have been accepted for publication by sending them to dicty@northwestern.edu or by using the form at http://dictybase.org/db/cgi-bin/dictyBase/abstract_submit. Back issues of dictyNews, the Dicty Reference database and other useful information is available at dictyBase - http://dictybase.org. Follow dictyBase on twitter: http://twitter.com/dictybase HAPPY HOLIDAYS! ========= Abstracts ========= Mycobacterium marinum degrades both triacylglycerols and phospholipids from its Dictyostelium host to synthesise its own triacylglycerols and generate lipid inclusions Caroline Barisch and Thierry Soldati PLoS Pathogens, in press During a tuberculosis infection and inside lipid-laden foamy macrophages, fatty acids (FAs) and sterols are the major energy and carbon source for Mycobacterium tuberculosis. Mycobacteria can be found both inside a vacuole and the cytosol, but how this impacts their access to lipids is not well appreciated. Lipid droplets (LDs) store FAs in form of triacylglycerols (TAGs) and are energy reservoirs of prokaryotes and eukaryotes. Using the Dictyostelium discoideum/Mycobacterium marinum infection model we showed that M. marinum accesses host LDs to build up its own intracytosolic lipid inclusions (ILIs). Here, we show that host LDs aggregate at regions of the bacteria that become exposed to the cytosol, and appear to coalesce on their hydrophobic surface leading to a transfer of diacylglycerol O-acyltransferase 2 (Dgat2)-GFP onto the bacteria. Dictyostelium knockout mutants for both Dgat enzymes are unable to generate LDs. Instead, the excess of exogenous FAs is esterified predominantly into phospholipids, inducing uncontrolled proliferation of the endoplasmic reticulum (ER). Strikingly, in absence of host LDs, M. marinum alternatively exploits these phospholipids, resulting in rapid reversal of ER-proliferation. In addition, the bacteria are unable to restrict their acquisition of lipids from the dgat1 and 2 double knockout leading to vast accumulation of ILIs. Recent data indicate that the presence of ILIs is one of the characteristics of dormant mycobacteria. During Dictyostelium infection, ILI formation in M. marinum is not accompanied by a significant change in intracellular growth and a reduction in metabolic activity, thus providing evidence that storage of neutral lipids does not necessarily induce dormancy. submitted by: Thierry Soldati [thierry.soldati@unige.ch] ——————————————————————————————————————— Adenylate cyclase A acting on PKA mediates induction of stalk formation by c-di-GMP at the Dictyostelium organizer Zhi-hui Chen a, Reema Singh a,b, Christian Cole b, Hajara Lawal a, Christina Schilde a, Melanie Febrer c, Geoffrey J. Barton b and Pauline Schaap a* Divisions of Cell and Developmental Biology a, Computational Biology b, Molecular Medicine c, School of Life Sciences, University of Dundee, Dundee, UK. Proc. Natl. Acad. Sci. USA, in press Coordination of cell movement with cell differentiation is a major feat of embryonic development. The Dictyostelium stalk always forms at the organizing tip by a mechanism that is not understood. We previously reported that c-di-GMP, synthesized by diguanylate cyclase A (DgcA), induces stalk formation. We here used transcriptional profiling of dgca- structures to identify target genes for c-di-GMP, and used these genes to investigate the c-di-GMP signal transduction pathway. We found that knock-down of PKA activity in prestalk cells reduced stalk gene induction by c-di-GMP, while PKA activation bypassed the c-di-GMP requirement for stalk gene expression. c-di-GMP caused a persistent increase in cAMP, which still occurred in mutants lacking the adenylate cyclases ACG or ACR, or the cAMP phosphodiesterase RegA. However, both inhibition of adenylate cyclase A (ACA) with SQ22536, and incubation of a temperature- sensitive ACA mutant at the restrictive temperature prevented c-di-GMP induced cAMP synthesis as well as c-di-GMP induced stalk gene transcription. ACA produces the cAMP signals that coordinate Dictyostelium morphogenetic cell movement and is highly expressed at the organizing tip. The stalk-less dgca- mutant regained its stalk by expression of a light- activated adenylate cyclase from the ACA promoter and exposure to light, indicating that cAMP is also the intermediate for c-di-GMP in vivo. Our data show that the more widely expressed DgcA activates tip-expressed ACA, which then acts on PKA to induce stalk genes. These results explain why stalk formation in Dictyostelia always initiates at the site of the morphogenetic organizer. submitted by: Pauline Schaap [p.schaap@dundee.ac.uk] ——————————————————————————————————————— A polycycstin-type transient receptor potential (Trp) channel that is activated by ATP David Traynor & Robert R. Kay Open Biology, in press ATP and ADP are ancient extra-cellular signalling molecules that in Dictyostelium amoebae cause rapid, transient increases in cytosolic calcium due to an influx through the plasma membrane. This response is independent of hetero-trimeric G-proteins, the putative IP3 receptor IplA and all P2X channels. We show, unexpectedly, that it is abolished in mutants of the polycystin-type transient receptor potential channel, TrpP. Responses to the chemoattractants cyclic-AMP and folic acid are unaffected in TrpP mutants. We report that the DIF morphogens, cyclic-di-GMP, GABA, glutamate and adenosine all induce strong cytoplasmic calcium responses, likewise independently of TrpP. Thus TrpP is dedicated to purinergic signalling. ATP treatment causes cell blebbing within seconds but this does not require TrpP, implicating a separate purinergic receptor. We could detect no effect of ATP on chemotaxis and TrpP mutants grow, chemotax and develop almost normally in standard conditions. No gating ligand is known for the human homologue of TrpP, polycystin-2, which causes polycystic kidney disease. Our results now show that TrpP mediates purinergic signalling in Dictyostelium and is directly or indirectly gated by ATP. submitted by: Rob Kay [rrk@mrc-lmb.cam.ac.uk] ============================================================== [End dictyNews, volume 42, number 30]