Zygote giant cell differentiation in Dictyostelium discoideum: biochemical markers of specific stages of sexual development

1992 ◽  
Vol 70 (10-11) ◽  
pp. 1200-1208 ◽  
Author(s):  
Darren D. Browning ◽  
Keith E. Lewis ◽  
Danton H. O'Day
Keyword(s):  
Alkaline Phosphatase ◽  
Cell Differentiation ◽  
Giant Cell ◽  
Dictyostelium Discoideum ◽  
Cell Fusion ◽  
Sexual Development ◽  
Giant Cells ◽  
Second Phase ◽  
Developmental Phase ◽  
Calcium Dependent

Sexual development in Dictyostelium discoideum has many unique features making it an attractive eukaryotic model system for the study of biomembrane fusion and intercellular communication. The work presented here provides primary biochemical evidence for two distinct phases during early sexual development that appear to be defined by calcium-dependent gamete cell fusion. In addition, we introduce a novel procedure for the enrichment of zygote giant cells and use this method to define certain wheat-germ agglutinin binding glycoproteins which are specifically located in zygote giant cells and others which are markers for surrounding amoebae in the second phase of development. In addition, a G protein which is present in high amounts early in development is unique to giant cells in the second phase, suggesting a role in phagocytosis. Finally, alkaline phosphatase activity was found to mark the first phase of sexual development, suggesting a role in cell fusion. This contrasts with the patterns of α-mannosidase and β-glucosidase activity that increase late in the second developmental phase, where they likely function in endocyte digestion during the cytophagic period. The developmental significance of these findings is discussed.Key words: zygote giant cell differentiation, Ca2+, glycoproteins, GTP-binding proteins, alkaline phosphatase, glycosidase, cell fusion.

The regulation of sexual development in Dictyostelium discoideum: cannibalistic behaviour of the giant cell

1985 ◽  
Vol 31 (5) ◽  
pp. 423-428 ◽  
Author(s):  
Keith E. Lewis ◽  
Danton H. O'Day
Keyword(s):  
Giant Cell ◽  
Dictyostelium Discoideum ◽  
Sexual Development ◽  
Giant Cells ◽  
Neutral Red ◽  
Cannibalistic Behaviour ◽  
Sexual Cultures ◽  
Drug Studies ◽  
Temperature Optima ◽  
Aerobic And Anaerobic

Sexual development leading to macrocyst formation in Dictyostelium discoideum involves the development of zygote giant cells which subsequently chemoattract and cannibalize local unfused amoebae of the same genotypes from which they formed. A system for assessing this process was developed wherein purified giant cells were fed amoebae under various conditions with the production of endocytes being assessed after neutral red staining of living cells. The phagocytic rate was linear during the 3-h assay period, was directly related to the concentration of amoebae, and demonstrated limited pH and temperature optima. Light which inhibits macrocyst development had no effect on sexual endocytosis. Cell-free conditioned medium from sexual cultures actively involved in phagocytosis enhanced giant cell endocytosis in the assay system. Drug studies revealed that microfilaments and microtubules are involved in the endocytic process and energy for it is supplied by both aerobic and anaerobic metabolic pathways. The data are discussed in terms of phagocytosis in D. discoideum and other organisms.

Phagocytic specificity during sexual development in Dictyostelium discoideum

1986 ◽  
Vol 32 (2) ◽  
pp. 79-82 ◽  
Author(s):  
Keith E. Lewis ◽  
Danton H. O'Day
Keyword(s):  
Dictyostelium Discoideum ◽  
Concanavalin A ◽  
Sexual Development ◽  
Wheat Germ ◽  
Giant Cells ◽  
Inhibitory Effect ◽  
Sexual Cycle ◽  
Type I ◽  
Further Development ◽  
Type Receptor

During the sexual cycle of Dictyostelium discoideum, zygote giant cells develop and serve as foci for further development by chemoattracting and cannibalizing hundreds of local amoebae. Previous work has shown that the phagocytic process bears similarities to and differences from asexual endocytosis. In the present study, sexual phagocytosis in D. discoideum was found to be species and developmental stage specific. It was inhibited selectively by glucose and concanavalin A. Although a partial, inhibitory effect of mannose on phagocytosis was not statistically significant, alpha-methylmannosamine, like alpha-methyl-glucose, significantly restored the phagocytic competence of giant cells treated with concanavalin A. Other sugars (N-acetyl glucosamine, N-acetylgalactosamine, and galactose) and lectins (wheat germ agglutinin, Ulex europus type I, and Ricinis communis agglutinin type I) had no significant effect on sexual phagocytosis. Together these data indicate that a glucose-type receptor is involved in selective uptake of D. discoideum amoebae by giant cells.

The immediate induction of extensive cell fusion by Ca2+ addition in Dictyostelium discoideum

1986 ◽  
Vol 64 (12) ◽  
pp. 1281-1287 ◽  
Author(s):  
David R. McConachie ◽  
Danton H. O'Day
Keyword(s):  
Dictyostelium Discoideum ◽  
Cell Fusion ◽  
Cell Population ◽  
Giant Cells ◽  
Binucleate Cells ◽  
Large Numbers ◽  
Synchronous Cell ◽  
Extensive Cell ◽  
Chloride Salts

In mated cultures (NC4 × V12) of Dictyostelium discoideum containing 1.0 mM CaCl2, cell fusion generates large numbers of binucleate cells which develop into zygote giant cells. In the absence of Ca2+, binucleate formation does not occur. When 1.0 mM CaCl2 is added to Ca2+-deficient cultures at 18 h, 50% of the cells fuse within 45 min producing large multinucleate syncytia. Small, presumptive gametes appear in Ca2+-deficient cultures and reach a peak of about 20% of the cell population by 10 h, but they maintain this plateau and do not fuse. Upon the addition of CaCl2, the presumptive gametes immediately fuse, producing binucleate cells which develop rapidly into morphologically distinct giant cells. Cell fusion continues, resulting in the formation of extremely large (40–80 μm diameter) multinucleate syncytia by 45 min. The induction of this extensive, synchronous cell fusion does not occur in the presence of other chloride salts and EGTA inhibits it, revealing that Ca+ is the regulatory ion.

scholarly journals Enzyme synthesis in the cellular slime mould Dictyostelium discoideum during the differentiation of myxamoebae grown axenically

1972 ◽  
Vol 126 (3) ◽  
pp. 609-615 ◽  
Author(s):  
J. Quance ◽  
J. M. Ashworth
Keyword(s):  
Alkaline Phosphatase ◽  
Acid Phosphatase ◽  
Cell Differentiation ◽  
Dictyostelium Discoideum ◽  
Specific Activity ◽  
Enzyme Synthesis ◽  
Cellular Slime Mould ◽  
Slime Mould ◽  
Cellular Slime ◽  
Developmental Programme

1. Myxamoebae of the cellular slime mould Dictyostelium discoideum Ax-2 were grown on different media, and were harvested either in the stationary or exponential phases of the growth cycle to yield samples of myxamoebae differing in enzymic composition. 2. Morphogenesis and cell differentiation phenomena in D. discoideum appear to be similar in myxamoebae grown and harvested under different conditions. 3. The specific activity of the enzymes β-N-acetylglucosaminidase, acid phosphatase, α-mannosidase, β-glucosidase and alkaline phosphatase have been determined during cell differentiation of myxamoebae grown and harvested under different conditions. 4. The pattern of synthesis of these enzymes, all of which have been claimed to be part of the ‘developmental programme’, either remains unaffected despite the origin of the myxamoebae (alkaline phosphatase) or is qualitatively similar but quantitatively affected (acid phosphatase, β-glucosidase) or is both qualitatively and quantitatively affected by changes in the myxamoebae (α-mannosidase, β-N-acetylglucosaminidase). 5. The implications of these results for the concept of a ‘developmental programme‘ are discussed.

scholarly journals Differentiation of Trophoblast Giant Cells and Their Metabolic Functions Are Dependent on Peroxisome Proliferator-Activated Receptor β/δ

2006 ◽  
Vol 26 (8) ◽  
pp. 3266-3281 ◽  
Author(s):  
Karim Nadra ◽  
Silvia I. Anghel ◽  
Elisabeth Joye ◽  
Nguan Soon Tan ◽  
Sharmila Basu-Modak ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
Giant Cell ◽  
Giant Cells ◽  
Peroxisome Proliferator ◽  
Phosphatidylinositol 3 Kinase ◽  
Peroxisome Proliferator Activated Receptor ◽  
Metabolic Functions ◽  
Integrin Linked Kinase ◽  
Trophoblast Giant Cells

ABSTRACT Mutation of the nuclear receptor peroxisome proliferator-activated receptor β/δ (PPARβ/δ) severely affects placenta development, leading to embryonic death at embryonic day 9.5 (E9.5) to E10.5 of most, but not all, PPARβ/δ-null mutant embryos. While very little is known at present about the pathway governed by PPARβ/δ in the developing placenta, this paper demonstrates that the main alteration of the placenta of PPARβ/δ-null embryos is found in the giant cell layer. PPARβ/δ activity is in fact essential for the differentiation of the Rcho-1 cells in giant cells, as shown by the severe inhibition of differentiation once PPARβ/δ is silenced. Conversely, exposure of Rcho-1 cells to a PPARβ/δ agonist triggers a massive differentiation via increased expression of 3-phosphoinositide-dependent kinase 1 and integrin-linked kinase and subsequent phosphorylation of Akt. The links between PPARβ/δ activity in giant cells and its role on Akt activity are further strengthened by the remarkable pattern of phospho-Akt expression in vivo at E9.5, specifically in the nucleus of the giant cells. In addition to this phosphatidylinositol 3-kinase/Akt main pathway, PPARβ/δ also induced giant cell differentiation via increased expression of I-mfa, an inhibitor of Mash-2 activity. Finally, giant cell differentiation at E9.5 is accompanied by a PPARβ/δ-dependent accumulation of lipid droplets and an increased expression of the adipose differentiation-related protein (also called adipophilin), which may participate to lipid metabolism and/or steroidogenesis. Altogether, this important role of PPARβ/δ in placenta development and giant cell differentiation should be considered when contemplating the potency of PPARβ/δ agonist as therapeutic agents of broad application.

scholarly journals The HAND1 Basic Helix-Loop-Helix Transcription Factor Regulates Trophoblast Differentiation via Multiple Mechanisms

2000 ◽  
Vol 20 (2) ◽  
pp. 530-541 ◽  
Author(s):  
Ian C. Scott ◽  
Lynn Anson-Cartwright ◽  
Paul Riley ◽  
Danny Reda ◽  
James C. Cross
Keyword(s):  
Transcription Factor ◽  
Cell Differentiation ◽  
Giant Cell ◽  
Giant Cells ◽  
Mutant Phenotype ◽  
Bhlh Transcription Factor ◽  
Placental Development ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
Ectoplacental Cone

ABSTRACT The basic helix-loop-helix (bHLH) transcription factor genesHand1 and Mash2 are essential for placental development in mice. Hand1 promotes differentiation of trophoblast giant cells, whereas Mash2 is required for the maintenance of giant cell precursors, and its overexpression prevents giant cell differentiation. We found that Hand1 expression and Mash2 expression overlap in the ectoplacental cone and spongiotrophoblast, layers of the placenta that contain the giant cell precursors, indicating that the antagonistic activities ofHand1 and Mash2 must be coordinated. MASH2 and HAND1 both heterodimerize with E factors, bHLH proteins that are the DNA-binding partners for most class B bHLH factors and which are also expressed in the ectoplacental cone and spongiotrophoblast. In vitro, HAND1 could antagonize MASH2 function by competing for E-factor binding. However, the Hand1 mutant phenotype cannot be solely explained by ectopic activity of MASH2, as the Hand1mutant phenotype was not altered by further mutation ofMash2. Interestingly, expression of E-factor genes (ITF2 and ALF1) was down-regulated in the trophoblast lineage prior to giant cell differentiation. Therefore, suppression of MASH2 function, required to allow giant cell differentiation, may occur in vivo by loss of its E-factor partner due to loss of its expression and/or competition from HAND1. In giant cells, where E-factor expression was not detected, HAND1 presumably associates with a different bHLH partner. This may account for the distinct functions of HAND1 in giant cells and their precursors. We conclude that development of the trophoblast lineage is regulated by the interacting functions of HAND1, MASH2, and their cofactors.

The low molecular weight autoinhibitor of sexual development in Dictyostelium discoideum inhibits cell fusion and zygote differentiation

1984 ◽  
Vol 62 (8) ◽  
pp. 722-731 ◽  
Author(s):  
Stephen P. Szabo ◽  
Danton H. O'Day
Keyword(s):  
Molecular Weight ◽  
Giant Cell ◽  
Dictyostelium Discoideum ◽  
Cell Fusion ◽  
Cell Formation ◽  
Sexual Cycle ◽  
Low Molecular Weight ◽  
Heat Stable ◽  
Giant Cell Formation ◽  
Zygote Development

A previous study has shown that, during the sexual cycle of Dictyostelium discoideum, zygote giant cell formation is regulated by an autoinhibitor. Experiments reported here show that the autoinhibitor inhibits two events of zygote development: cell fusion and subsequent giant cell differentiation. The autoinhibitor is heat stable and has a molecular weight around 500. Medium containing the autoinhibitor can be diluted 500-fold without loss of activity. Preliminary experiments show that, although levels of ammonia double during the 8-h period of autoinhibitor production, added ammonia does not mimic the inhibiting effect. cAMP at 1 mM inhibits both binucleate formation and differentiation, but the concentration of cyclic AMP in 28-h cultures is only 13.4 pmol, a level which does not affect zygote development. Thus, it is established that neither of these critical regulators of other developmental processes in D. discoideum is the autoinhibitor.

Macrocyst development in Dictyostelium discoideum. III. Cell-fusion inducing factor secreted by giant cells

1983 ◽  
Vol 62 (1) ◽  
pp. 237-248
Author(s):  
Y. Saga ◽  
K. Yanagisawa
Keyword(s):  
Dictyostelium Discoideum ◽  
Cell Fusion ◽  
Actinomycin D ◽  
External Medium ◽  
Giant Cells ◽  
Culture Conditions ◽  
Sexual Cycle ◽  
Opposite Mating Type ◽  
Fusion Products ◽  
Type Strains

A factor was discovered that markedly enhances the degree of fusion activity between cells of the opposite mating-type strains, MI1 and NC4, in the sexual cycle of Dictyostelium discoideum. This factor, designated cell-fusion inducing factor (CFIF), was detected initially in supernatants taken from 24 h dark-grown mixed cultures of HM1 and NC4 cells, and subsequently shown to be secreted by giant cells - the fusion products of HM1 and NC4 cells. HM1 cells, cultured in the dark at appropriate temperature, normally acquire fusion-competence specific to NC4 cells. The addition of CFIF to such dark-grown HM1 cultures results in a marked increase in their fusion-competence. In addition, when CFIF is added to light-grown HM1 cultures, in which cells normally do not acquire the ability to fuse with NC4 cells, fusion-competence is induced. The fusion-competence of NC4 cells is unaffected by CFIF, being quite high under the culture conditions used here. Experiments, using actinomycin D, daunomycin and cycloheximide showed that the secretion of CFIF from giant cells requires synthesis of RNA and protein. Possibly, cell fusion triggers production of CFIF, which is rapidly released into the external medium.

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