scholarly journals Multiple cysteine proteinase forms during the life cycle of Dictyostelium discoideum revealed by electrophoretic analysis

1988 ◽  
Vol 254 (1) ◽  
pp. 261-268 ◽  
Author(s):  
M J North ◽  
K I Scott ◽  
B C Lockwood
Keyword(s):  
Dictyostelium Discoideum ◽  
Growth And Development ◽  
Developmental Regulation ◽  
Cysteine Proteinase ◽  
Electrophoretic Analysis ◽  
Cysteine Proteinases ◽  
Cellular Slime Mould ◽  
Extracellular Form ◽  
Cellular Slime ◽  
Individual Enzyme

Proteinases of the cellular slime mould Dictyostelium discoideum have been analysed using electrophoresis on polyacrylamide gels containing gelatin (gelatin/PAGE). Multiple proteinase forms were apparent in vegetative myxamoebae, but the presence of individual enzyme forms depended on the manner in which the cells were grown. Axenic cells had a characteristic A-pattern of proteinases consisting of six bands, the most active enzymes having apparent Mr values of 51,000 and 45,000 (these have been named ddCP51 and ddCP45, respectively). Some of the proteinases were also present in the medium, the major extracellular form was ddCP42, a 42,000-Mr enzyme. Cells grown in association with bacteria had a distinct B-pattern with three main enzymes that had apparent Mr values of 48,000, 43,000 and 38,000. All of the A- and B-pattern proteinases were most active at acid pH in the presence of dithiothreitol and were inhibited by various agents such as trans-epoxysuccinyl-L-leucylamido-(4-guanidino)butane (E64), leupeptin and chymostatin, which inactivate cysteine proteinases. One of the enzymes, ddCP30, was identified as cysteine proteinase B which had been purified and characterized previously [North, M.J. & Whyte, A. (1984) J. Gen. Microbiol. 130, 123-134]. During starvation of axenic cells in shaken suspensions some of the vegetative proteinases disappeared, ddCP42 was released from the cells and one new enzyme with an apparent Mr of 48,000 appeared. Addition of cyclic AMP had little effect on these changes. When the axenically grown myxamoebae underwent development on filters, similar changes in band pattern were observed and the aggregation stage was characterized by the presence of three cysteine proteinase bands (apparent Mr values of 48,000, 45,000 and 43,000). Proteinases, especially ddCP42, were released from the cells and could be collected from the buffer-saturated pads which supported the filters. The results demonstrate that cysteine proteinases are present throughout growth and development of D. discoideum and that the forms present are subject to nutritional and developmental regulation.

scholarly journals Properties and developmental regulation of an α-d-galactosidase from Dictyostelium discoideum

1976 ◽  
Vol 158 (2) ◽  
pp. 409-417 ◽  
Author(s):  
D C Kilpatrick ◽  
J L Stirling
Keyword(s):  
Protein Synthesis ◽  
Dictyostelium Discoideum ◽  
Early Development ◽  
Subcellular Distribution ◽  
De Novo ◽  
Specific Activity ◽  
Soluble Fraction ◽  
Developmental Regulation ◽  
Cellular Slime Mould ◽  
Cellular Slime

An alpha-D-galactosidase was detected in cells of the cellular slime mould, Dictyostelium discoideum, at all stages of development. Its specific activity was highest during early development (interphase), and this accumulation of enzyme appears to require protein synthesis de novo. Its subcellular distribution differs from that of other D. discoideum glycosidases, since most activity was recovered in the soluble fraction. No evidence was obtained for more than one isoenzymic form after subjection of extracts to electrophoresis and various chromatographic procedures. It is excreted from the cell during development, but no evidence was found for an extracellular function for the enzyme.

scholarly journals A bacterial factor induces changes in cysteine proteinase forms in the cellular slime mould Dictyostelium discoideum

1988 ◽  
Vol 254 (1) ◽  
pp. 269-275 ◽  
Author(s):  
M J North
Keyword(s):  
Dictyostelium Discoideum ◽  
Cysteine Proteinase ◽  
Gel Filtration ◽  
Electrophoretic Pattern ◽  
Cysteine Proteinases ◽  
Prolonged Incubation ◽  
Cell Extracts ◽  
Latex Beads ◽  
Cellular Slime ◽  
Simultaneous Loss

The electrophoretic pattern of cysteine proteinases in axenically grown myxamoebae of Dictyostelium discoideum can be altered by the addition of either Gram-negative (Klebsiella aerogenes, Escherichia coli) or Gram-positive (Micrococcus lysodeikticus, Bacillus subtilis) bacteria to the culture. No changes occurred, however, if either yeast or latex beads were used in place of bacteria. The changes involved the simultaneous loss of proteinases characteristic of the axenic cells (the A-forms) and the acquisition of those found in cells which have been grown on bacteria (the B-forms). Using K. aerogenes the conversion was complete within 4 h. Extracellular proteinase activity was unaffected during this period. After the D. discoideum cells had been lysed, no equivalent change in proteinase band pattern could be produced either by prolonged incubation of cell extracts or by treatment with proteinases. An identical conversion could be induced in cultures of myxamoebae by a factor, cysteine proteinase converting factor (CPCF), present in the 15,000 g supernatant of a sonicated suspension of K. aerogenes. CPCF was macromolecular, as demonstrated by both ultrafiltration and gel filtration, acid-precipitable, but was soluble in ethanol or alkali. Its activity was unaffected by treatment with trypsin. The results suggested that CPCF might be a component of the bacterial cell wall, and since its activity was affected by lysozyme treatment, peptidoglycan is implicated. The results can be interpreted in terms of a novel nutrient-dependent post-translational change which affected most of the cysteine proteinases present in D. discoideum myxamoebae.

scholarly journals Growth of myxamoebae of the cellular slime mould Dictyostelium discoideum in axenic culture

1970 ◽  
Vol 119 (2) ◽  
pp. 171-174 ◽  
Author(s):  
D. J. Watts ◽  
J. M. Ashworth
Keyword(s):  
Cell Differentiation ◽  
Dictyostelium Discoideum ◽  
Axenic Culture ◽  
Axenic Medium ◽  
Cellular Slime Mould ◽  
Slime Mould ◽  
Solid Media ◽  
Cellular Slime

1. A simple axenic medium suitable for the growth of the myxamoebae of a strain of the cellular slime mould Dictyostelium discoideum is described. 2. Procedures suitable for the growth of this strain in liquid and on solid media are described. 3. Conditions suitable for initiating the cell differentiation of myxamoebae grown axenically are described.

scholarly journals Glycoproteins that exhibit extensive size polymorphisms in Dictyostelium discoideum.

Genetics ◽  
1989 ◽  
Vol 122 (1) ◽  
pp. 59-64 ◽  
Author(s):  
E Smith ◽  
A A Gooley ◽  
G C Hudson ◽  
K L Williams
Keyword(s):  
Linkage Group ◽  
Dictyostelium Discoideum ◽  
Allelic Variation ◽  
Surface Glycoprotein ◽  
Developmentally Regulated ◽  
Electrophoretic Variants ◽  
Cellular Slime Mould ◽  
Repeat Units ◽  
Group I ◽  
Cellular Slime

Abstract Electrophoretic variants which arise from amino acid substitutions, leading to charge differences between proteins are ubiquitous and have been used extensively for genetic analysis. Less well documented are polymorphisms in the size of proteins. Here we report that a group of glycoproteins, which share a common carbohydrate epitope, vary in size in different isolates of the cellular slime mould, Dictyostelium discoideum. One of these proteins, PsA, a developmentally regulated prespore-specific surface glycoprotein, has previously been shown to exist in three size forms due to allelic variation at the pspA locus on linkage group I. In this report, a second glycoprotein, PsB, which is also prespore specific but found inside prespore cells, is studied. PsB maps to linkage group II and exhibits at least four different sizes in the isolates examined. We propose that the size polymorphisms are the product of allelic variation at the pspB locus, due to differences in the number of repeat units.

The role of phosphodiesterase in aggregation of Dictyostelium discoideum

1978 ◽  
Vol 31 (1) ◽  
pp. 233-243
Author(s):  
M. Darmon ◽  
J. Barra ◽  
P. Brachet
Keyword(s):  
Dictyostelium Discoideum ◽  
Direct Contact ◽  
Signal To Noise Ratio ◽  
Signal To Noise ◽  
Extracellular Medium ◽  
Camp Phosphodiesterase ◽  
Cellular Slime Mould ◽  
Cellular Slime ◽  
Mutant Cells

The role of cAMP phosphodiesterase in the cAMP-mediated aggregation of the cellular slime mould Dictyostelium discoideum was investigated with a morphogenetic mutant defective in phosphodiesterase production. Mutant cells become capable of aggregating normally when incubated in the presence of exogenous phosphodiesterase isolated from Idictyostelium or rat brain. Direct contact between enzyme and the cell membrane is not required for this phenotypic suppression. The aggregateless character of this strain presumably results from an over-accumulation of cAMP in the extracellular medium since aggregation can be induced in the absence of added phosphodiesterase under conditions facilitating diffusion of the nucleotide. This suggests that phosphodiesterase is not involved in the generation or recognition of cAMP signals, but that the enzyme is essential in the control of the cAMP signal-to-noise ratio.

Control of cellular differentiation by temperature in the cellular slime mould Dictyostelium discoideum

1984 ◽  
Vol 69 (1) ◽  
pp. 159-165
Author(s):  
M. Maeda
Keyword(s):  
Low Temperature ◽  
Dictyostelium Discoideum ◽  
Cellular Differentiation ◽  
Cellular Slime Mould ◽  
Slime Mould ◽  
Cellular Slime

The effects of low temperature on morphogenesis and cellular differentiation of Dictyostelium discoideum were examined. During incubation at 5 degrees C, the vegetative and preaggregation cells never developed, but cell masses at the aggregation or slug stage developed to form hemispherical, or dumbbell-shaped multicellular structures. By staining with FITC-antispore IgG, the structures formed after 10 days of incubation of tipped aggregates at 5 degrees C were found to be composed of 90% spores, 5% prespore cells and 5% non-stained cells. Since only 20% of the total cells constituting the tipped aggregate had been prespore cells at the beginning of incubation, this showed that spore differentiation proceeded even at low temperature, while stalk differentiation was completely inhibited. Similar results were obtained when the cells were incubated at 3 degrees C. However, at 0 degree C, morphogenesis and cellular differentiation did not occur, although most of the prespore cells at the late culmination stage differentiated incompletely into spores. Possible reasons for the high proportion of spores being induced by low temperature are discussed.

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