scholarly journals Developmental Commitment in Dictyostelium discoideum

2007 ◽  
Vol 6 (11) ◽  
pp. 2038-2045 ◽  
Author(s):  
Mariko Katoh ◽  
Guokai Chen ◽  
Emily Roberge ◽  
Gad Shaulsky ◽  
Adam Kuspa
Keyword(s):  
Amino Acids ◽  
Cell Proliferation ◽  
Cyclic Amp ◽  
Dictyostelium Discoideum ◽  
Growth Phase ◽  
Camp Signaling ◽  
Phagocytic Function ◽  
Fruiting Bodies ◽  
Mutant Strains ◽  
Multicellular Organisms

ABSTRACT Upon starvation, Dictyostelium discoideum cells halt cell proliferation, aggregate into multicellular organisms, form migrating slugs, and undergo morphogenesis into fruiting bodies while differentiating into dormant spores and dead stalk cells. At almost any developmental stage cells can be forced to dedifferentiate when they are dispersed and diluted into nutrient broth. However, migrating slugs can traverse lawns of bacteria for days without dedifferentiating, ignoring abundant nutrients and continuing development. We now show that developing Dictyostelium cells revert to the growth phase only when bacteria are supplied during the first 4 to 6 h of development but that after this time, cells continue to develop regardless of the presence of food. We postulate that the cells’ inability to revert to the growth phase after 6 h represents a commitment to development. We show that the onset of commitment correlates with the cells’ loss of phagocytic function. By examining mutant strains, we also show that commitment requires extracellular cyclic AMP (cAMP) signaling. Moreover, cAMP pulses are sufficient to induce both commitment and the loss of phagocytosis in starving cells, whereas starvation alone is insufficient. Finally, we show that the inhibition of development by food prior to commitment is independent of contact between the cells and the bacteria and that small soluble molecules, probably amino acids, inhibit development during the first few hours and subsequently the cells become unable to react to the molecules and commit to development. We propose that commitment serves as a checkpoint that ensures the completion of cooperative aggregation of developing Dictyostelium cells once it has begun, dampening the response to nutritional cues that might inappropriately block development.

Action of a slowly hydrolysable cyclic AMP analogue on developing cells of Dictyostelium discoideum

1979 ◽  
Vol 35 (1) ◽  
pp. 321-338
Author(s):  
C. Rossier ◽  
G. Gerisch ◽  
D. Malchow
Keyword(s):  
Cyclic Amp ◽  
Dictyostelium Discoideum ◽  
Type Strain ◽  
Wild Type Strain ◽  
Agar Plate ◽  
Cell Aggregation ◽  
Fruiting Bodies ◽  
Wild Type ◽  
Camp Analogue ◽  
Order Of Magnitude

Adenosine 3′,5′-cyclic phosphorothioate (cAMP-S) is a cyclic AMP (cAMP) analogue which is only slowly hydrolysed by phosphodiesterases of Dictyostelium discoideum. The affinity of cAMP-S to cAMP receptors at the cell surface is only one order of magnitude lower than that of cAMP. cAMP-S can replace cAMP as a stimulant with respect to all receptor-mediated responses tested, including chemotaxis and the induction of cAMP pulses. cAMP-S does not affect growth of D. discoideum but it blocks cell aggregation at a uniform concentration of 5 × 10(−7) M in agar plate cultures of strain NC-4 as well as its axenically growing derivative, Ax-2. Another wild-type strain of D. discoideum, v-12, is able to aggregate on agar plates supplemented with 1 mM cAMP-S. The development of Polysphondylium pallidum and P. violaceum is also highly cAMP-S resistant. In Ax-2 both differentiation from the growth phase to the aggregation-competent stage and chemotaxis are cAMP-S sensitive, whereas in v-12 only chemotaxis is inhibited. v-12 can still form streams of cohering cells and fruiting bodies when chemotaxis is inhibited by cAMP-S. Whereas cAMP induces differentiation into stalk cells at concentrations of 10(−3) or 10(−4) M, cAMP-S has the same effect in strain v-12 at the much lower concentration of 10(−6) M.

Identification of the sequences controlling cyclic AMP regulation and cell-type-specific expression of a prestalk-specific gene in Dictyostelium discoideum

1987 ◽  
Vol 7 (1) ◽  
pp. 149-159
Author(s):  
S Datta ◽  
R A Firtel
Keyword(s):  
Amino Acids ◽  
Cyclic Amp ◽  
Dictyostelium Discoideum ◽  
Base Pair ◽  
Regulatory Region ◽  
Specific Gene ◽  
Cell Type ◽  
Specific Expression ◽  
Endogenous Gene ◽  
Cell Type Specific Expression

We have cloned and analyzed a developmentally and spatially regulated prestalk cell-specific gene from Dictyostelium discoideum. The gene encodes a protein highly homologous to the lysosomal cysteine proteinases cathepsin H and cathepsin B. Amino acid comparisons between these enzymes showed that the active-site amino acids were conserved, as were amino acids known to be important for catalysis and residues which form the intramolecular cysteine bridges. We have constructed a series of internal deletions, duplications, and linker scanner mutations within the region 300 base pairs 5' to the cap site. Analysis of expression of the mutations in transformants identified a approximately 35-base pair GC-rich region containing a dAdC/dGdT palindromic repeat and a G-rich box which is homologous to the 3' GT half of the palindromic repeat. Deletion or disruption of the G box resulted in a approximately 50-fold drop in the level of expression of the gene fusion in transformants in response to cyclic AMP in single-cell culture but did not affect the temporal pattern of regulation or control by cyclic AMP. The expression of such constructs during normal multicellular differentiation paralleled that of the endogenous gene; however, the level of RNA from the constructs was only approximately 10-fold lower than that of constructs containing the G box. Deletion of the 3' half of the palindromic sequence and the G box region resulted in a dramatic decrease in the level of transcription, although the constructs still showed proper temporal expression. These results suggest that this 35-base-pair region acts as an important part of the regulatory region for cell type and cyclic AMP regulation.

Dictyostelium discoideum lipids modulate cell-cell cohesion and cyclic AMP signaling

1991 ◽  
Vol 11 (1) ◽  
pp. 468-475
Author(s):  
D R Fontana ◽  
C S Luo ◽  
J C Phillips
Keyword(s):  
Cyclic Amp ◽  
Adenylyl Cyclase ◽  
Dictyostelium Discoideum ◽  
Membrane Lipids ◽  
Camp Signaling ◽  
Cell Contact ◽  
Complete Elimination ◽  
Cyclase Activation ◽  
Independent Events ◽  
Cell Cell

During Dictyostelium discoideum development, cell-cell communication is mediated through cyclic AMP (cAMP)-induced cAMP synthesis and secretion (cAMP signaling) and cell-cell contact. Cell-cell contact elicits cAMP secretion and modulates the magnitude of a subsequent cAMP signaling response (D. R. Fontana and P. L. Price, Differentiation 41:184-192, 1989), demonstrating that cell-cell contact and cAMP signaling are not independent events. To identify components involved in the contact-mediated modulation of cAMP signaling, amoebal membranes were added to aggregation-competent amoebae in suspension. The membranes from aggregation-competent amoebae inhibited cAMP signaling at all concentrations tested, while the membranes from vegetative amoebae exhibited a concentration-dependent enhancement or inhibition of cAMP signaling. Membrane lipids inhibited cAMP signaling at all concentrations tested. The lipids abolished cAMP signaling by blocking cAMP-induced adenylyl cyclase activation. The membrane lipids also inhibited amoeba-amoeba cohesion at concentrations comparable to those which inhibited cAMP signaling. The phospholipids and neutral lipids decreased cohesion and inhibited the cAMP signaling response. The glycolipid/sulfolipid fraction enhanced cohesion and cAMP signaling. Caffeine, a known inhibitor of cAMP-induced adenylyl cyclase activation, inhibited amoeba-amoeba cohesion. These studies demonstrate that endogenous lipids are capable of modulating amoeba-amoeba cohesion and cAMP-induced activation of the adenylyl cyclase. These results suggest that cohesion may modulate cAMP-induced adenylyl cyclase activation. Because the complete elimination of cohesion is accompanied by the complete elimination of cAMP signaling, these results further suggest that cohesion may be necessary for cAMP-induced adenylyl cyclase activation in D. discoideum.

scholarly journals Rates of degradation and synthesis of glycosidases de novo during growth and differentiation of Dictyostelium discoideum

1975 ◽  
Vol 148 (2) ◽  
pp. 169-177 ◽  
Author(s):  
D Every ◽  
J M Ashworth
Keyword(s):  
Amino Acids ◽  
Enzyme Activity ◽  
Life Cycle ◽  
Dictyostelium Discoideum ◽  
Growth Medium ◽  
Growth Phase ◽  
Enzyme Activities ◽  
De Novo ◽  
Antibody Preparation ◽  
Differentiated Cells

1. Injection of a purified preparation of β-N-acetylglucosaminidase from the spent growth medium of myxamoebae of Dictyostelium discoideum into rabbits gave rise to an antibody preparation containing both anti-α-glucosidase and anti-β-acetylglucosaminidase activities. 2. These two activities were shown to reside in different immunoglobulin molecules and it was concluded that the β-N-acetylglucosaminidase preparation contained trace amounts of highly antigenic α-glucosidase. 3. A single precipitin band having β-N-acetylglucosaminidase activity was formed in Ouchterlony plates when this antibody preparation was tested against extracts obtained from differentiated cells or from myxamoebae grown either axenically or on bacteria. 4. The antibody preparation was used to show that both β-N-acetylglucosaminidase and α-glucosidase molecules are synthesized de novo from isotopically labelled amino acids during both the growth and differentiation phases of the life cycle and to show that neither of these proteins is significantly degraded during the growth phase or during the first 9h of differentiation. 5. The rates of accumulation of these assayable enzyme activities are thus equal to their rates of synthesis during growth and early differentiation. 6. The factors regulating cellular enzyme activity during the life cycle of D. discoideum are discussed.

scholarly journals Identification of the sequences controlling cyclic AMP regulation and cell-type-specific expression of a prestalk-specific gene in Dictyostelium discoideum.

1987 ◽  
Vol 7 (1) ◽  
pp. 149-159 ◽  
Author(s):  
S Datta ◽  
R A Firtel
Keyword(s):  
Amino Acids ◽  
Cyclic Amp ◽  
Dictyostelium Discoideum ◽  
Base Pair ◽  
Regulatory Region ◽  
Specific Gene ◽  
Cell Type ◽  
Specific Expression ◽  
Endogenous Gene ◽  
Cell Type Specific Expression

We have cloned and analyzed a developmentally and spatially regulated prestalk cell-specific gene from Dictyostelium discoideum. The gene encodes a protein highly homologous to the lysosomal cysteine proteinases cathepsin H and cathepsin B. Amino acid comparisons between these enzymes showed that the active-site amino acids were conserved, as were amino acids known to be important for catalysis and residues which form the intramolecular cysteine bridges. We have constructed a series of internal deletions, duplications, and linker scanner mutations within the region 300 base pairs 5' to the cap site. Analysis of expression of the mutations in transformants identified a approximately 35-base pair GC-rich region containing a dAdC/dGdT palindromic repeat and a G-rich box which is homologous to the 3' GT half of the palindromic repeat. Deletion or disruption of the G box resulted in a approximately 50-fold drop in the level of expression of the gene fusion in transformants in response to cyclic AMP in single-cell culture but did not affect the temporal pattern of regulation or control by cyclic AMP. The expression of such constructs during normal multicellular differentiation paralleled that of the endogenous gene; however, the level of RNA from the constructs was only approximately 10-fold lower than that of constructs containing the G box. Deletion of the 3' half of the palindromic sequence and the G box region resulted in a dramatic decrease in the level of transcription, although the constructs still showed proper temporal expression. These results suggest that this 35-base-pair region acts as an important part of the regulatory region for cell type and cyclic AMP regulation.

scholarly journals Dictyostelium discoideum lipids modulate cell-cell cohesion and cyclic AMP signaling.

1991 ◽  
Vol 11 (1) ◽  
pp. 468-475 ◽  
Author(s):  
D R Fontana ◽  
C S Luo ◽  
J C Phillips
Keyword(s):  
Cyclic Amp ◽  
Adenylyl Cyclase ◽  
Dictyostelium Discoideum ◽  
Membrane Lipids ◽  
Camp Signaling ◽  
Cell Contact ◽  
Complete Elimination ◽  
Cyclase Activation ◽  
Independent Events ◽  
Cell Cell

During Dictyostelium discoideum development, cell-cell communication is mediated through cyclic AMP (cAMP)-induced cAMP synthesis and secretion (cAMP signaling) and cell-cell contact. Cell-cell contact elicits cAMP secretion and modulates the magnitude of a subsequent cAMP signaling response (D. R. Fontana and P. L. Price, Differentiation 41:184-192, 1989), demonstrating that cell-cell contact and cAMP signaling are not independent events. To identify components involved in the contact-mediated modulation of cAMP signaling, amoebal membranes were added to aggregation-competent amoebae in suspension. The membranes from aggregation-competent amoebae inhibited cAMP signaling at all concentrations tested, while the membranes from vegetative amoebae exhibited a concentration-dependent enhancement or inhibition of cAMP signaling. Membrane lipids inhibited cAMP signaling at all concentrations tested. The lipids abolished cAMP signaling by blocking cAMP-induced adenylyl cyclase activation. The membrane lipids also inhibited amoeba-amoeba cohesion at concentrations comparable to those which inhibited cAMP signaling. The phospholipids and neutral lipids decreased cohesion and inhibited the cAMP signaling response. The glycolipid/sulfolipid fraction enhanced cohesion and cAMP signaling. Caffeine, a known inhibitor of cAMP-induced adenylyl cyclase activation, inhibited amoeba-amoeba cohesion. These studies demonstrate that endogenous lipids are capable of modulating amoeba-amoeba cohesion and cAMP-induced activation of the adenylyl cyclase. These results suggest that cohesion may modulate cAMP-induced adenylyl cyclase activation. Because the complete elimination of cohesion is accompanied by the complete elimination of cAMP signaling, these results further suggest that cohesion may be necessary for cAMP-induced adenylyl cyclase activation in D. discoideum.

scholarly journals Characterization of glutathione S-transferase enzymes in Dictyostelium discoideum suggests a functional role for the GSTA2 isozyme in cell proliferation and development

PLoS ONE ◽  
2021 ◽  
Vol 16 (4) ◽  
pp. e0250704
Author(s):  
Mamatha Garige ◽  
Eric Walters
Keyword(s):  
Cell Proliferation ◽  
Dictyostelium Discoideum ◽  
Functional Role ◽  
Model Organism ◽  
Fruiting Bodies ◽  
Percent Reduction ◽  
Glutathione S Transferase ◽  
Gst Activity ◽  
Starvation Conditions

In this report, we extend our previous characterization of Dictyostelium discoideum glutathione S-transferase (DdGST) enzymes that are expressed in the eukaryotic model organism. Transcript profiling of gstA1-gstA5 (alpha class) genes in vegetative, log phase cells identified gstA2 and gstA3 with highest expression (6–7.5-fold, respectively) when compared to other gstA transcripts. Marked reductions in all gstA transcripts occurred under starvation conditions, with gstA2 and gstA3 exhibiting the largest decreases (-96% and -86.6%, respectively). When compared to their pre-starvation levels, there was also a 60 percent reduction in total GST activity. Glutathione (GSH) pull-down assay and mass spectroscopy detected three isozymes (DdGSTA1, DdGSTA2 and DdGSTA3) that were predominantly expressed in vegetative cells. Biochemical and kinetic comparisons between rDdGSTA2 and rDdGSTA3 shows higher activity of rDdGSTA2 to the CDNB (1-chloro-2,4-dinitrobenzene) substrate. RNAi-mediated knockdown of endogenous DdGSTA2 caused a 60 percent reduction in proliferation, delayed development, and altered morphogenesis of fruiting bodies, whereas overexpression of rDdGSTA2 enzyme had no effect. These findings corroborate previous studies that implicate a role for phase II GST enzymes in cell proliferation, homeostasis, and development in eukaryotic cells.

scholarly journals The Cyclic AMP-Dependent Protein Kinase A Network Regulates Development and Virulence in Aspergillus fumigatus

2004 ◽  
Vol 72 (9) ◽  
pp. 5193-5203 ◽  
Author(s):  
Burghard Liebmann ◽  
Meike Müller ◽  
Armin Braun ◽  
Axel A. Brakhage
Keyword(s):  
Signal Transduction ◽  
Protein Kinase ◽  
Cyclic Amp ◽  
Aspergillus Fumigatus ◽  
Protein Kinase A ◽  
Gene Fusion ◽  
Camp Signaling ◽  
Crude Extracts ◽  
Mutant Strains ◽  
Kinase A

ABSTRACT Aspergillus fumigatus is an important pathogen of immunocompromised hosts, causing pneumonia and invasive disseminated disease with high mortality. To determine the importance of the cyclic AMP (cAMP) signaling pathway for virulence, the pkaC1 gene encoding a protein kinase A (PKA) catalytic subunit was cloned and characterized. Deletion of pkaC1 led to reduced conidiation and growth. PKA activity was not detectable in ΔpkaC1, ΔgpaB, and ΔacyA mutant strains. gpaB and acyA encode a G protein α subunit involved in cAMP signal transduction and adenylate cyclase, respectively. Addition of cAMP led to PKA activity in crude extracts of both the ΔgpaB and ΔacyA strains but not in crude extracts of the ΔpkaC1 strain. These findings provide evidence that PKAC1 represents the predominant form of PKA under the conditions tested, and GPAB and ACYA are members of the cAMP signaling cascade. Analysis of a pksPp-lacZ gene fusion indicated that the expression of the pathogenicity determinant-encoding pksP gene was reduced in ΔpkaC1 mutant strains compared with the expression of the gene fusion in the parental strain. In a low-dose murine inhalation model, conidia of both the ΔpkaC1 and ΔgpaB mutant strains were almost avirulent. Taken together, these findings indicate that the cAMP-PKA signal transduction pathway is required for A. fumigatus pathogenicity.

PROTECTIVE EFFECT OF POLYPEPTIDE AND AMINO ACIDS ON THE DEVELOPMENT OF THE NERVOUS TISSUE CULTURE IN THE PRESENCE OF CYCLOPHOSPHAMIDE

2017 ◽  
pp. 22-26
Author(s):  
V. B. Dolgo-Saburov ◽  
N. I. Chalisova ◽  
L. V. Lyanginen ◽  
E. S. Zalomaeva
Keyword(s):  
Amino Acids ◽  
Cell Proliferation ◽  
Tissue Culture ◽  
Protective Effect ◽  
Organotypic Culture ◽  
Inhibitory Effect ◽  
Mustard Gas ◽  
Synthesis Inhibitor ◽  
Combined Administration ◽  
The Central Nervous System

In an organotypic culture, an investigation was conducted into combined effects of cyclophosphamide DNA as synthesis inhibitor used to model a resorptive action of mustard gas, and cortexin polypeptide or each of 20 encoded amino acids on the development of cell proliferation in cerebral cortex explants of the rat. The combined administration of cyclophosphamide together with cortexin or with each of the 20 encoded amino acids, except glycine, showed suppression of the cytostatic agent inhibitory effect. Thus, cortexin and amino acids have a protective effect on cell proliferation in the tissue culture of the central nervous system under the action of mustardlike substances.

Characterization of YY1 OPB Peptide for its Anticancer Activity

2019 ◽  
Vol 19 (6) ◽  
pp. 504-511 ◽  
Author(s):  
Yige Qi ◽  
Ting Yan ◽  
Lu Chen ◽  
Qiang Zhang ◽  
Weishu Wang ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Amino Acids ◽  
Cell Proliferation ◽  
Cancer Cell ◽  
Breast Cancer Cell ◽  
Expression Vectors ◽  
Cancer Cell Proliferation ◽  
Breast Cancer Cell Proliferation ◽  
Oncogene Products ◽  
And Migration

Background:The oncoprotein binding (OPB) domain of Yin Yang 1 (YY1) consists of 26 amino acids between G201 and S226, and is involved in YY1 interaction with multiple oncogene products, including MDM2, AKT, EZH2 and E1A. Through the OPB domain, YY1 promotes the oncogenic or proliferative regulation of these oncoproteins in cancer cells. We previously demonstrated that a peptide with the OPB sequence blocked YY1-AKT interaction and inhibited breast cancer cell proliferation.Objective:In the current study, we characterized the OPB domain and determined a minimal region for peptide design to suppress cancer cellMethods:Using alanine-scan method, we identified that the amino acids at OPB C-terminal are essential to YY1 binding to AKT. Further studies suggested that serine and threonine residues, but not lysines, in OPB play a key role in YY1-AKT interaction. We generated GFP fusion expression vectors to express OPB peptides with serially deleted N-terminal and found that OPB1 (i.e. G201-S226) is cytoplasmic, but OPB2 (i.e. E206-S226), OPB3 (i.e. E206-S226) and control peptide were both nuclear and cytoplasmic.Results:Both OPB1 and 2 inhibited breast cancer cell proliferation and migration, but OPB3 exhibited similar effects to control. OPB1 and 2 caused cell cycle arrest at G1 phase, increased p53 and p21 expression, and reduced AKT(S473) phosphorylation in MCF-7 cells, but not in MDA-MB-231 cells.Conclusion:: Overall, the serines and threonines of OPB are essential to YY1 binding to oncoproteins, and OPB peptide can be minimized to E206-S226 that maintain inhibitory activity to YY1- promoted cell proliferation.

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