scholarly journals An Adenylyl Cyclase, CyaB, Acts as an Osmosensor in Myxococcus xanthus

2005 ◽  
Vol 187 (10) ◽  
pp. 3593-3598 ◽  
Author(s):  
Yoshio Kimura ◽  
Mika Ohtani ◽  
Kaoru Takegawa
Keyword(s):  
Cyclic Amp ◽  
Adenylyl Cyclase ◽  
Growth Retardation ◽  
Spore Germination ◽  
Germination Rate ◽  
Myxococcus Xanthus ◽  
Receptor Type ◽  
Wild Type ◽  
Vegetative Cells ◽  
Mutant Cells

ABSTRACT We have previously reported that a receptor-type adenylyl cyclase (CyaA) of Myxococcus xanthus undergoes an osmosensor mainly during spore germination (Y. Kimura et al., J. Bacteriol. 184:3578-3585, 2002). In the present study, we cloned another receptor-type adenylyl cyclase gene (cyaB) and characterized the function of the cyaB-encoded protein. Disruption of cyaB generates a mutant that showed growth retardation at high ionic (NaCl) or high nonionic (sucrose) osmolarity. When vegetative cells were stimulated with 0.15 M NaCl, the increases in intracellular cyclic AMP levels of cyaB mutant cells were lower than those of wild-type cells. Under nonionic osmostress, the cyaB mutant exhibited reduced spore germination; however, the germination rate of the cyaB mutant was significantly higher than that of the cyaA mutant.

scholarly journals An Adenylyl Cyclase, CyaA, of Myxococcus xanthus Functions in Signal Transduction during Osmotic Stress

2002 ◽  
Vol 184 (13) ◽  
pp. 3578-3585 ◽  
Author(s):  
Yoshio Kimura ◽  
Yukako Mishima ◽  
Hiromi Nakano ◽  
Kaoru Takegawa
Keyword(s):  
Osmotic Stress ◽  
Adenylyl Cyclase ◽  
Spore Germination ◽  
Developmental Stages ◽  
Catalytic Domain ◽  
Myxococcus Xanthus ◽  
Normal Growth ◽  
Wild Type ◽  
Amino Terminal ◽  
Mutant Cells

ABSTRACT An adenylyl cyclase gene (cyaA) present upstream of an osmosensor protein gene (mokA) was isolated from Myxococcus xanthus. cyaA encoded a polypeptide of 843 amino acids with a predicted molecular mass of 91,187 Da. The predicted cyaA gene product had structural similarity to the receptor-type adenylyl cyclases that are composed of an amino-terminal sensor domain and a carboxy-terminal catalytic domain of adenylyl cyclase. In reverse transcriptase PCR experiments, the transcript of the cyaA gene was detected mainly during development and spore germination. A cyaA mutant, generated by gene disruption, showed normal growth, development, and germination. However, a cyaA mutant placed under conditions of ionic (NaCl) or nonionic (sucrose) osmostress exhibited a marked reduction in spore formation and spore germination. When wild-type and cyaA mutant cells at developmental stages were stimulated with 0.2 M NaCl or sucrose, the mutant cells increased cyclic AMP accumulation at levels similar to those of the wild-type cells. In contrast, the mutant cells during spore germination had mainly lost the ability to respond to high-ionic osmolarity. In vegetative cells, the cyaA mutant responded normally to osmotic stress. These results suggested that M. xanthus CyaA functions mainly as an ionic osmosensor during spore germination and that CyaA is also required for osmotic tolerance in fruiting formation and sporulation.

An adenylyl cyclase that functions during late development of Dictyostelium

Development ◽  
1999 ◽  
Vol 126 (23) ◽  
pp. 5463-5471 ◽  
Author(s):  
F. Soderbom ◽  
C. Anjard ◽  
N. Iranfar ◽  
D. Fuller ◽  
W.F. Loomis
Keyword(s):  
Adenylyl Cyclase ◽  
Critical Role ◽  
Fruiting Bodies ◽  
Wild Type ◽  
Adenylyl Cyclase Activity ◽  
Extracellular Signals ◽  
Mutant Strains ◽  
Morphological Mutants ◽  
Low Levels ◽  
Mutant Cells

A variety of extracellular signals lead to the accumulation of cAMP which can act as a second message within cells by activating protein kinase A (PKA). Expression of many of the essential developmental genes in Dictyostelium discoideum are known to depend on PKA activity. Cells in which the receptor-coupled adenylyl cyclase gene, acaA, is genetically inactivated grow well but are unable to develop. Surprisingly, acaA(−) mutant cells can be rescued by developing them in mixtures with wild-type cells, suggesting that another adenylyl cyclase is present in developing cells that can provide the internal cAMP necessary to activate PKA. However, the only other known adenylyl cyclase gene in Dictyostelium, acgA, is only expressed during germination of spores and plays no role in the formation of fruiting bodies. By screening morphological mutants generated by Restriction Enzyme Mediated Integration (REMI) we discovered a novel adenylyl cyclase gene, acrA, that is expressed at low levels in growing cells and at more than 25-fold higher levels during development. Growth and development up to the slug stage are unaffected in acrA(−) mutant strains but the cells make almost no viable spores and produce unnaturally long stalks. Adenylyl cyclase activity increases during aggregation, plateaus during the slug stage and then increases considerably during terminal differentiation. The increase in activity following aggregation fails to occur in acrA(−) cells. As long as ACA is fully active, ACR is not required until culmination but then plays a critical role in sporulation and construction of the stalk.

scholarly journals Ras Proteins Have Multiple Functions in Vegetative Cells of Dictyostelium

2010 ◽  
Vol 9 (11) ◽  
pp. 1728-1733 ◽  
Author(s):  
Parvin Bolourani ◽  
George Spiegelman ◽  
Gerald Weeks
Keyword(s):  
Cyclic Amp ◽  
Vegetative Cell ◽  
Ras Proteins ◽  
Random Motility ◽  
Vegetative Cells ◽  
Content Type ◽  
Multiple Functions ◽  
Mutant Cells

ABSTRACT During the aggregation of Dictyostelium cells, signaling through RasG is more important in regulating cyclic AMP (cAMP) chemotaxis, whereas signaling through RasC is more important in regulating the cAMP relay. However, RasC is capable of substituting for RasG for chemotaxis, since rasG − cells are only partially deficient in chemotaxis, whereas rasC −/rasG − cells are totally incapable of chemotaxis. In this study we have examined the possible functional overlap between RasG and RasC in vegetative cells by comparing the vegetative cell properties of rasG −, rasC −, and rasC −/rasG − cells. In addition, since RasD, a protein not normally found in vegetative cells, is expressed in vegetative rasG − and rasC −/rasG − cells and appears to partially compensate for the absence of RasG, we have also examined the possible functional overlap between RasG and RasD by comparing the properties of rasG − and rasC −/rasG − cells with those of the mutant cells expressing higher levels of RasD. The results of these two lines of investigation show that RasD is capable of totally substituting for RasG for cytokinesis and growth in suspension, whereas RasC is without effect. In contrast, for chemotaxis to folate, RasC is capable of partially substituting for RasG, but RasD is totally without effect. Finally, neither RasC nor RasD is able to substitute for the role that RasG plays in regulating actin distribution and random motility. These specificity studies therefore delineate three distinct and none-overlapping functions for RasG in vegetative cells.

scholarly journals Genetic and Molecular Analysis of cglB, a Gene Essential for Single-Cell Gliding in Myxococcus xanthus

1999 ◽  
Vol 181 (14) ◽  
pp. 4381-4390 ◽  
Author(s):  
Ana M. Rodriguez ◽  
Alfred M. Spormann
Keyword(s):  
Single Cell ◽  
Myxococcus Xanthus ◽  
Gliding Motility ◽  
Open Reading Frames ◽  
Wild Type Allele ◽  
Wild Type ◽  
Isolated Cells ◽  
Type Allele ◽  
C Terminus ◽  
Mutant Cells

ABSTRACT Gliding movements of individual isolated Myxococcus xanthus cells depend on the genes of the A-motility system (agl and cgl genes). Mutants carrying defects in those genes are unable to translocate as isolated cells on solid surfaces. The motility defect of cgl mutants can be transiently restored to wild type by extracellular complementation upon mixing mutant cells with wild-type or other motility mutant cells. To develop a molecular understanding of the function of a Cgl protein in gliding motility, we cloned the cglB wild-type allele by genetic complementation of the mutant phenotype. The nucleotide sequence of a 2.85-kb fragment was determined and shown to encode two complete open reading frames. The CglB protein was determined to be a 416-amino-acid putative lipoprotein with an unusually high cysteine content. The CglB antigen localized to the membrane fraction. The swarming and gliding defects of a constructed ΔcglBmutant were fully restored upon complementation with thecglB wild-type allele. Experiments with a cglBallele encoding a CglB protein with a polyhistidine tag at the C terminus showed that this allele also promoted wild-type levels of swarming and single-cell gliding, but was unable to stimulate ΔcglB cells to move. Possible functions of CglB as a mechanical component or as a signal protein in single cell gliding are discussed.

scholarly journals Phototaxis and Impaired Motility in Adenylyl Cyclase and Cyclase Receptor Protein Mutants of Synechocystis sp. Strain PCC 6803

2006 ◽  
Vol 188 (20) ◽  
pp. 7306-7310 ◽  
Author(s):  
Devaki Bhaya ◽  
Kenlee Nakasugi ◽  
Fariba Fazeli ◽  
Matthew S. Burriesci
Keyword(s):  
Cyclic Amp ◽  
Adenylyl Cyclase ◽  
Receptor Protein ◽  
Wild Type ◽  
Catabolite Activator Protein ◽  
Activator Protein ◽  
Protein Mutants ◽  
Synechocystis Sp ◽  
Pcc 6803

ABSTRACT We have carefully characterized and reexamined the motility and phototactic responses of Synechocystis sp. adenylyl cyclase (Cya1) and catabolite activator protein (SYCRP1) mutants to different light regimens, glucose, 3-(3,4-dichlorophenyl)-1,1-dimethylurea, and cyclic AMP. We find that contrary to earlier reports, cya1 and sycrp1 mutants are motile and phototactic but are impaired in one particular phase of phototaxis in comparison with wild-type Synechocystis sp.

scholarly journals Characterization of bcsA Mutations That Bypass Two Distinct Signaling Requirements for Myxococcus xanthus Development

2002 ◽  
Vol 184 (18) ◽  
pp. 5141-5150 ◽  
Author(s):  
John K. Cusick ◽  
Elizabeth Hager ◽  
Ronald E. Gill
Keyword(s):  
Cell Density ◽  
Myxococcus Xanthus ◽  
Wild Type ◽  
Amino Acid Similarity ◽  
A Cell ◽  
Protease Deficient ◽  
Mutant Cells ◽  
Low Cell Density ◽  
Significant Amino Acid

ABSTRACT The BsgA protease is required for starvation-induced development in Myxococcus xanthus. Bypass suppressors of a bsgA mutant were isolated to identify genes that may encode additional components of BsgA protease-dependent regulation of development. Strain M951 was isolated following Tn5 mutagenesis of a bsgA mutant and was capable of forming fruiting bodies and viable spores in the absence of the BsgA protease. The Tn5Ω951 insertion was localized to a gene, bcsA, that encodes a protein that has significant amino acid similarity to a group of recently described flavin-containing monooxygenases involved in styrene catabolism. Mutations in bcsA bypassed the developmental requirements for both extracellular B and C signaling but did not bypass the requirement for A signaling. Bypass of the B-signaling requirement by the bcsA mutation was accompanied by restored expression of a subset of developmentally induced lacZ fusions to the BsgA protease-deficient strain. bcsA mutant cells developed considerably faster than wild-type cells at low cell density and altered transcriptional levels of a developmentally induced, cell-density-regulated gene (Ω4427), suggesting that the bcsA gene product may normally act to inhibit development in a cell-density-regulated fashion. Bypass of the requirements for both B and C signaling by bcsA mutations suggests a possible link between these two genetically, biochemically, and temporally distinct signaling requirements.

Cytoskeletons from a mutant of Dictyostelium discoideum with flattened cells

1986 ◽  
Vol 86 (1) ◽  
pp. 69-82
Author(s):  
M. Claviez ◽  
M. Brink ◽  
G. Gerisch
Keyword(s):  
Cyclic Amp ◽  
Dictyostelium Discoideum ◽  
Cell Shape ◽  
Wild Type ◽  
Aggregation Pattern ◽  
Mutant Cells ◽  
Moving Front ◽  
Microtubular System

Development of a mutant of Dictyostelium discoideum, HG403, is described whose cells spread strongly on a substratum. Although the mutant cells were less clearly polarized into the front and rear ends, and usually less extensively elongated than wild-type cells, their aggregation pattern was only slightly less regular. Cells of the mutant responded well to cyclic AMP by chemotaxis, although their capability of stabilizing cell shape and maintaining dominance of a single moving front appeared to be reduced. Mutant HG403 proved to be ideal for the preparation of cytoskeletons in which the organization of the microtubular system, the network of filaments between them, the dense texture of the microfilament network at the periphery of the cells, as well as the bundling of microfilaments in spike-like extensions, could be observed.

Ultrastructure and Morphology of the Neurospora Crassa Cell Wall Mutants, Slime And Slime X, Using Tem and Sem

1976 ◽  
Vol 34 ◽  
pp. 54-55
Author(s):  
Karen S. Howard ◽  
H. D. Braymer ◽  
M. D. Socolofsky ◽  
S. A. Milligan
Keyword(s):  
Cell Wall ◽  
Neurospora Crassa ◽  
Wild Type ◽  
Morphological Comparison ◽  
Small Areas ◽  
Solid Media ◽  
Thin Sectioning ◽  
X Cells ◽  
Mutant Cells ◽  
Isolated Cell

The recently isolated cell wall mutant slime X of Neurospora crassa was prepared for ultrastructural and morphological comparison with the cell wall mutant slime. The purpose of this article is to discuss the methods of preparation for TEM and SEM observations, as well as to make a preliminary comparison of the two mutants.TEM: Cells of the slime mutant were prepared for thin sectioning by the method of Bigger, et al. Slime X cells were prepared in the same manner with the following two exceptions: the cells were embedded in 3% agar prior to fixation and the buffered solutions contained 5% sucrose throughout the procedure.SEM: Two methods were used to prepare mutant and wild type Neurospora for the SEM. First, single colonies of mutant cells and small areas of wild type hyphae were cut from solid media and fixed with OSO4 vapors similar to the procedure used by Harris, et al. with one alteration. The cell-containing agar blocks were dehydrated by immersion in 2,2-dimethoxypropane (DMP).

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