scholarly journals A Putative Ariadne-Like Ubiquitin Ligase Is Required for Dictyostelium discoideum Development

2006 ◽  
Vol 5 (10) ◽  
pp. 1820-1825 ◽  
Author(s):  
Nathaniel Whitney ◽  
Lacey J. Pearson ◽  
Ryan Lunsford ◽  
Lisa McGill ◽  
Richard H. Gomer ◽  
...  
Keyword(s):  
Dictyostelium Discoideum ◽  
Ubiquitin Ligase ◽  
Fruiting Bodies ◽  
Wild Type ◽  
Cell Numbers

ABSTRACT The Dictyostelium rbrA gene encodes a putative Ariadne ubiquitin ligase. rbrA − cells form defective slugs that cannot phototax. Prestalk cell numbers are reduced in rbrA − slugs, and these prestalk cells do not localize to the tip of slugs. Chimeric slugs containing wild-type cells could phototax and form fruiting bodies.

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.

scholarly journals Mitochondrial processing peptidase activity is controlled by the processing of α-MPP during development in Dictyostelium discoideum

Microbiology ◽  
2010 ◽  
Vol 156 (4) ◽  
pp. 978-989 ◽  
Author(s):  
Koki Nagayama ◽  
Tetsuo Ohmachi
Keyword(s):  
Life Cycle ◽  
Dictyostelium Discoideum ◽  
Early Development ◽  
Normal Development ◽  
Peptidase Activity ◽  
Fruiting Bodies ◽  
Wild Type ◽  
Formation Stage ◽  
Mitochondrial Processing Peptidase ◽  
Processing Peptidase

We investigated the expression of the α subunit of the Dictyostelium mitochondrial processing peptidase (Ddα-MPP) during development. Ddα-MPP mRNA is expressed at the highest levels in vegetatively growing cells and during early development, and is markedly downregulated after 10 h of development. The Ddα-MPP protein is expressed as two forms, designated α-MPPH and α-MPPL, throughout the Dictyostelium life cycle. The larger form, α-MPPH, is cleaved to produce the functional α-MPPL form. We were not able to isolate mutants in which the α-mpp gene had been disrupted. Instead, an antisense transformant, αA2, expressing α-MPP at a lower level than the wild-type AX-3 was isolated to examine the function of the α-MPP protein. Development of the αA2 strain was normal until the slug formation stage, but the slug stage was prolonged to ∼24 h. In this prolonged slug stage, only α-MPPH was present, and α-MPPL protein and MPP activity were not detected. After 28 h, α-MPPL and MPP activity reappeared, and normal fruiting bodies were formed after a delay of approximately 8 h compared with normal development. These results indicate that MPP activity is controlled by the processing of α-MPPH to α-MPPL during development in Dictyostelium.

scholarly journals Genetic Interactions of the E3 Ubiquitin Ligase Component FbxA with Cyclic AMP Metabolism and a Histidine Kinase Signaling Pathway during Dictyostelium discoideum Development

2003 ◽  
Vol 2 (3) ◽  
pp. 618-626 ◽  
Author(s):  
Turgay Tekinay ◽  
Herbert L. Ennis ◽  
Mary Y. Wu ◽  
Margaret Nelson ◽  
Richard H. Kessin ◽  
...  
Keyword(s):  
Cyclic Amp ◽  
Signaling Pathway ◽  
Dictyostelium Discoideum ◽  
Ubiquitin Ligase ◽  
Histidine Kinase ◽  
Double Mutant ◽  
E3 Ubiquitin Ligase ◽  
Specific Gene ◽  
Wild Type ◽  
Camp Phosphodiesterase

ABSTRACT Dictyostelium discoideum amoebae with an altered fbxA gene, which is thought to encode a component of an SCF E3 ubiquitin ligase, have defective regulation of cell type proportionality. In chimeras with wild-type cells, the mutant amoebae form mainly spores, leaving the construction of stalks to wild-type cells. To examine the role of fbxA and regulated proteolysis, we have recovered the promoter of fbxA and shown that it is expressed in a pattern resembling that of a prestalk-specific gene until late in development, when it is also expressed in developing spore cells. Because fbxA cells are developmentally deficient in pure culture, we were able to select suppressor mutations that promote sporulation of the original mutant. One suppressor mutation resides within the gene regA, which encodes a cyclic AMP (cAMP) phosphodiesterase linked to an activating response regulator domain. In another suppressor, there has been a disruption of dhkA, a gene encoding a two-component histidine kinase known to influence Dictyostelium development. RegA appears precociously and in greater amounts in the fbxA mutant than in the wild type, but in an fbxA/dhkA double mutant, RegA is restored to wild-type levels. Because the basis of regA suppression might involve alterations in cAMP levels during development, the concentrations of cAMP in all strains were determined. The levels of cAMP are relatively constant during multicellular development in all strains except the dhkA mutant, in which it is reduced at least sixfold. The level of cAMP in the double mutant dhkA/fbxA is relatively normal. The levels of cAMP in the various mutants do not correlate with spore formation, as would be expected on the basis of our present understanding of the signaling pathway leading to the induction of spores. Altered amounts of RegA and cAMP early in the development of the mutants suggest that both fbxA and dhkA genes act earlier than previously thought.

scholarly journals Regulation of Ammonia Homeostasis by the Ammonium Transporter AmtA in Dictyostelium discoideum

2007 ◽  
Vol 6 (12) ◽  
pp. 2419-2428 ◽  
Author(s):  
Ryuji Yoshino ◽  
Takahiro Morio ◽  
Yoko Yamada ◽  
Hidekazu Kuwayama ◽  
Masazumi Sameshima ◽  
...  
Keyword(s):  
Dictyostelium Discoideum ◽  
Growth And Development ◽  
Ammonium Transporter ◽  
Cellular Slime Mold ◽  
Fruiting Bodies ◽  
Wild Type ◽  
Developmental Processes ◽  
Multicellular Aggregates ◽  
Ammonium Transporters ◽  
Cellular Slime

ABSTRACT Ammonia has been shown to function as a morphogen at multiple steps during the development of the cellular slime mold Dictyostelium discoideum; however, it is largely unknown how intracellular ammonia levels are controlled. In the Dictyostelium genome, there are five genes that encode putative ammonium transporters: amtA, amtB, amtC, rhgA, and rhgB. Here, we show that AmtA regulates ammonia homeostasis during growth and development. We found that cells lacking amtA had increased levels of ammonia/ammonium, whereas their extracellular ammonia/ammonium levels were highly decreased. These results suggest that AmtA mediates the excretion of ammonium. In support of a role for AmtA in ammonia homeostasis, AmtA mRNA is expressed throughout the life cycle, and its expression level increases during development. Importantly, AmtA-mediated ammonia homeostasis is critical for many developmental processes. amtA − cells are more sensitive to NH4Cl than wild-type cells in inhibition of chemotaxis toward cyclic AMP and of formation of multicellular aggregates. Furthermore, even in the absence of exogenously added ammonia, we found that amtA − cells produced many small fruiting bodies and that the viability and germination of amtA − spores were dramatically compromised. Taken together, our data clearly demonstrate that AmtA regulates ammonia homeostasis and plays important roles in multiple developmental processes in Dictyostelium.

Investigation of a potential role for aldose reductase AlrA in tetrahydropteridine synthesis in Dictyostelium discoideum Ax2

Pteridines ◽  
2017 ◽  
Vol 28 (2) ◽  
pp. 97-103
Author(s):  
Hye-Lim Kim ◽  
Hyun-Chul Ryu ◽  
Young Shik Park
Keyword(s):  
Dictyostelium Discoideum ◽  
Aldose Reductase ◽  
Potential Role ◽  
Amino Acid Residues ◽  
Wild Type ◽  
High Sequence Identity ◽  
Physiological Conditions ◽  
Antioxidant Function ◽  
Cellular Extract ◽  
Sr Gene

AbstractDictyostelium discoideum Ax2 is well-known for the synthesis of d-threo-tetrahydrobiopterin (DH4) with a smaller amount of l-erythro-tetrahydrobiopterin (BH4). DH4 synthesis from 6-pyruvoyltetrahydropterin (PPH4) is catalyzed by aldose reductase (AR)-like protein and sepiapterin reductase (SR) via an intermediate 1′-oxo-2′-d-hydroxypropyl tetrahydropterin, which is non-enzymatically oxidized to d-sepiapterin in the absence of SR. However, l-sepiapterin was a dominant product in the reaction of a cellular extract of spr− disrupted in the SR gene. In order to investigate its potential role in tetrahydropteridine synthesis, the enzyme catalyzing l-sepiapterin synthesis from PPH4 was purified from spr−. Via mass spectrometry, the protein was identified to be encoded by alrA. AlrA consists of 297 amino acid residues sharing a high sequence identity with human AR. However, in the co-incubation assay, DH4 synthesis was not detected and, furthermore, the recombinant AlrA was observed to suppress BH4 synthesis by SR, which was known to prefer 1′-oxo-2′-d-hydroxypropyl tetrahydropterin to PPH4. Although intracellular DH4 level in alrA− was decreased to 60% of the wild type, it is presumed to result from the antioxidant function of DH4. Therefore, despite the structural and catalytic identities with human AR, AlrA seems to be involved in neither BH4, nor DH4 synthesis under normal physiological conditions.

scholarly journals Functional Promiscuity of Gene Regulation by Serpentine Receptors in Dictyostelium discoideum

1998 ◽  
Vol 18 (10) ◽  
pp. 5744-5749 ◽  
Author(s):  
Irene Verkerke-Van Wijk ◽  
Ji-Yun Kim ◽  
Raymond Brandt ◽  
Peter N. Devreotes ◽  
Pauline Schaap
Keyword(s):  
Gene Expression ◽  
Cell Fate ◽  
Dictyostelium Discoideum ◽  
Developmental Regulation ◽  
Mutant Cell ◽  
Gene Induction ◽  
Specific Gene ◽  
Wild Type ◽  
Animal Development ◽  
Camp Stimulation

ABSTRACT Serpentine receptors such as smoothened and frizzled play important roles in cell fate determination during animal development. InDictyostelium discoideum, four serpentine cyclic AMP (cAMP) receptors (cARs) regulate expression of multiple classes of developmental genes. To understand their function, it is essential to know whether each cAR is coupled to a specific gene regulatory pathway or whether specificity results from the different developmental regulation of individual cARs. To distinguish between these possibilities, we measured gene induction in car1 car3 double mutant cell lines that express equal levels of either cAR1, cAR2, or cAR3 under a constitutive promoter. We found that all cARs efficiently mediate both aggregative gene induction by cAMP pulses and induction of postaggregative and prespore genes by persistent cAMP stimulation. Two exceptions to this functional promiscuity were observed. (i) Only cAR1 can mediate adenosine inhibition of cAMP-induced prespore gene expression, a phenomenon that was found earlier in wild-type cells. cAR1’s mediation of adenosine inhibition suggests that cAR1 normally mediates prespore gene induction. (ii) Only cAR2 allows entry into the prestalk pathway. Prestalk gene expression is induced by differentiation-inducing factor (DIF) but only after cells have been prestimulated with cAMP. We found that DIF-induced prestalk gene expression is 10 times higher in constitutive cAR2 expressors than in constitutive cAR1 or cAR3 expressors (which still have endogenous cAR2), suggesting that cAR2 mediates induction of DIF competence. Since in wild-type slugs cAR2 is expressed only in anterior cells, this could explain the so far puzzling observations that prestalk cells differentiate at the anterior region but that DIF levels are actually higher at the posterior region. After the initial induction of DIF competence, cAMP becomes a repressor of prestalk gene expression. This function can again be mediated by cAR1, cAR2, and cAR3.

Electrical properties of the slime mould grex

Development ◽  
1970 ◽  
Vol 23 (2) ◽  
pp. 311-322
Author(s):  
D. R. Garrod ◽  
J. F. Palmer ◽  
L. Wolpert
Keyword(s):  
Electrical Properties ◽  
Dictyostelium Discoideum ◽  
Fruiting Body ◽  
Fruiting Bodies ◽  
Electrophysiological Investigation ◽  
Slime Mould ◽  
Cellular Pattern ◽  
Electric Potentials

An electrophysiological investigation of the migrating grex of the slime mould, Dictyostelium discoideum, has been carried out with two aims in view. It was hoped to obtain information which would be relevant to, first, the formation and regulation of cellular pattern in the grex, and secondly, the problem of grex movement. During migration the grex develops a simple, linear cellular pattern. The cells at the front become the so-called ‘prestalk’ cells which will form the stalk of the fruiting body while those at the back become ‘prespore’ cells and form spores at culmination (Raper, 1940; Bonner, 1944; Bonner & Slifkin, 1949). Moreover, this cellular pattern is capable of polarized regulation. Raper (1940) has shown that portions isolated from the front or back of the grex are capable of forming normally proportioned fruiting bodies. A number of workers have suggested that bio-electric potentials may be involved in regulation of linear cellular pattern.

The signal from fruiting body and conus tips of Dictyostelium discoideum

Development ◽  
1976 ◽  
Vol 36 (2) ◽  
pp. 261-271
Author(s):  
Jonathan Rubin
Keyword(s):  
Dictyostelium Discoideum ◽  
Fruiting Body ◽  
Beef Heart ◽  
Fruiting Bodies

Tips from fruiting bodies and conuses were transplanted into interphase fields of Dictyostelium discoideum amoebae. Progressively increasing concentrations of beef-heart phosphodiesterase added to the fields significantly decreased the chemotactic range of the responding amoebae. The findings suggest that the tip secretes c-AMP. We also find that the chemotactic range is independent of the size of the tip implying that the tip may produce a regulating gradient.

scholarly journals The ubiquitin-dependent ATPase p97 removes cytotoxic trapped PARP1 from chromatin

2022 ◽  
Author(s):  
Dragomir B. Krastev ◽  
Shudong Li ◽  
Yilun Sun ◽  
Andrew J. Wicks ◽  
Gwendoline Hoslett ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Homologous Recombination ◽  
Ubiquitin Ligase ◽  
Antitumour Activity ◽  
Parp Inhibitor ◽  
Bound State ◽  
Parp Inhibitors ◽  
Tumour Cells ◽  
Wild Type ◽  
Endogenous Proteins

AbstractPoly (ADP-ribose) polymerase (PARP) inhibitors elicit antitumour activity in homologous recombination-defective cancers by trapping PARP1 in a chromatin-bound state. How cells process trapped PARP1 remains unclear. Using wild-type and a trapping-deficient PARP1 mutant combined with rapid immunoprecipitation mass spectrometry of endogenous proteins and Apex2 proximity labelling, we delineated mass spectrometry-based interactomes of trapped and non-trapped PARP1. These analyses identified an interaction between trapped PARP1 and the ubiquitin-regulated p97 ATPase/segregase. We found that following trapping, PARP1 is SUMOylated by PIAS4 and subsequently ubiquitylated by the SUMO-targeted E3 ubiquitin ligase RNF4, events that promote recruitment of p97 and removal of trapped PARP1 from chromatin. Small-molecule p97-complex inhibitors, including a metabolite of the clinically used drug disulfiram (CuET), prolonged PARP1 trapping and enhanced PARP inhibitor-induced cytotoxicity in homologous recombination-defective tumour cells and patient-derived tumour organoids. Together, these results suggest that p97 ATPase plays a key role in the processing of trapped PARP1 and the response of tumour cells to PARP inhibitors.

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