scholarly journals Microtubule sliding in mutant Chlamydomonas axonemes devoid of outer or inner dynein arms.

1986 ◽  
Vol 103 (5) ◽  
pp. 1895-1902 ◽  
Author(s):  
T Okagaki ◽  
R Kamiya
Keyword(s):  
Functional Differentiation ◽  
The Other ◽  
Sliding Velocity ◽  
Wild Type ◽  
Michaelis Constant ◽  
The Poor ◽  
Special Procedure ◽  
Bacterial Protease ◽  
Mutant Strains ◽  
Dynein Arms

To clarify the functional differentiation between the outer and inner dynein arms in eukaryotic flagella, their mechanochemical properties were assessed by measuring the sliding velocities of outer-doublet microtubules in disintegrating axonemes of Chlamydomonas, using wild-type and mutant strains that lack either of the arms. A special procedure was developed to induce sliding disintegration in Chlamydomonas axonemes which is difficult to achieve by ordinary methods. The flagella were first fragmented by sonication, demembranated by Nonidet P-40, and then perfused under a microscope with Mg-ATP and nagarse, a bacterial protease with broad substrate specificity. The sliding velocity varied with the Mg-ATP concentration in a Michaelis-Menten manner in the axonemes from the wild type and a motile mutant lacking the outer dynein arm (oda38). The maximal sliding velocity and apparent Michaelis constant for Mg-ATP were measured to be 13.2 +/- 1.0 micron/s and 158 +/- 36 microM for the wild type and 2.0 +/- 0.1 micron/s and 64 +/- 18 microM for oda38. These maximal sliding velocities were significantly smaller than those estimated in beating axonemes; the reason is not clear. The velocities in the presence or absence of 10(-5) M Ca2+ did not differ noticeably. The axonemes of nonmotile mutants lacking either outer arms (pf13A, pf22) or inner arms (pf23) were examined for their ability to undergo sliding disintegration in the presence of 0.1 mM Mg-ATP. Whereas pf13A axonemes underwent normal sliding disintegration, the other two species displayed it only very poorly. The poor ability of pf23 axonemes to undergo sliding disintegration raises the possibility that the outer dynein arm cannot function well in the absence of the inner arm.

scholarly journals act Operon Control of Developmental Gene Expression in Myxococcus xanthus

2002 ◽  
Vol 184 (4) ◽  
pp. 1172-1179 ◽  
Author(s):  
Thomas M. A. Gronewold ◽  
Dale Kaiser
Keyword(s):  
Fruiting Body ◽  
Myxococcus Xanthus ◽  
The Other ◽  
Loss Of Function ◽  
Wild Type ◽  
Developmental Gene ◽  
Developmental Gene Expression ◽  
Mutant Strains ◽  
Genes Encoding ◽  
Signal Production

ABSTRACT Cell-bound C-signal guides the building of a fruiting body and triggers the differentiation of myxospores. Earlier work has shown that transcription of the csgA gene, which encodes the C-signal, is directed by four genes of the act operon. To see how expression of the genes encoding components of the aggregation and sporulation processes depends on C-signaling, mutants with loss-of-function mutations in each of the act genes were investigated. These mutations were found to have no effect on genes that are normally expressed up to 3 h into development and are C-signal independent. Neither the time of first expression nor the rate of expression increase was changed in actA, actB, actC, or actD mutant strains. Also, there was no effect on A-signal production, which normally starts before 3 h. By contrast, the null act mutants have striking defects in C-signal production. These mutations changed the expression of four gene reporters that are related to aggregation and sporulation and are expressed at 6 h or later in development. The actA and actB null mutations substantially decreased the expression of all these reporters. The other act null mutations caused either premature expression to wild-type levels (actC) or delayed expression (actD), which ultimately rose to wild-type levels. The pattern of effects on these reporters shows how the C-signal differentially regulates the steps that together build a fruiting body and differentiate spores within it.

scholarly journals IC138 Defines a Subdomain at the Base of the I1 Dynein That Regulates Microtubule Sliding and Flagellar Motility

2009 ◽  
Vol 20 (13) ◽  
pp. 3055-3063 ◽  
Author(s):  
Raqual Bower ◽  
Kristyn VanderWaal ◽  
Eileen O'Toole ◽  
Laura Fox ◽  
Catherine Perrone ◽  
...  
Keyword(s):  
Double Mutant ◽  
Essential Role ◽  
Null Allele ◽  
Flagellar Motility ◽  
Wild Type ◽  
Gene Encoding ◽  
Radial Spoke ◽  
Mutant Strains ◽  
Dynein Arms ◽  
Image Averaging

To understand the mechanisms that regulate the assembly and activity of flagellar dyneins, we focused on the I1 inner arm dynein (dynein f) and a null allele, bop5-2, defective in the gene encoding the IC138 phosphoprotein subunit. I1 dynein assembles in bop5-2 axonemes but lacks at least four subunits: IC138, IC97, LC7b, and flagellar-associated protein (FAP) 120—defining a new I1 subcomplex. Electron microscopy and image averaging revealed a defect at the base of the I1 dynein, in between radial spoke 1 and the outer dynein arms. Microtubule sliding velocities also are reduced. Transformation with wild-type IC138 restores assembly of the IC138 subcomplex and rescues microtubule sliding. These observations suggest that the IC138 subcomplex is required to coordinate I1 motor activity. To further test this hypothesis, we analyzed microtubule sliding in radial spoke and double mutant strains. The results reveal an essential role for the IC138 subcomplex in the regulation of I1 activity by the radial spoke/phosphorylation pathway.

scholarly journals Analysis of the movement of Chlamydomonas flagella:" the function of the radial-spoke system is revealed by comparison of wild-type and mutant flagella.

1982 ◽  
Vol 92 (3) ◽  
pp. 722-732 ◽  
Author(s):  
C J Brokaw ◽  
D J Luck ◽  
B Huang
Keyword(s):  
Recovery Phase ◽  
Wild Type ◽  
Radial Spoke ◽  
Suppressor Mutations ◽  
Mutant Strains ◽  
Type Pattern ◽  
Dynein Arms ◽  
Beta Frequency ◽  
Amplitude Pattern ◽  
Asymmetric Bending

The mutation uni-1 gives rise to uniflagellate Chlamydomonas cells which rotate around a fixed point in the microscope field, so that the flagellar bending pattern can be photographed easily. This has allowed us to make a detailed analysis of the wild-type flagellar bending pattern and the bending patterns of flagella on several mutant strains. Cells containing uni-1, and recombinants of uni-1 with the suppressor mutations, suppf-1 and suppf-3, show the typical asymmetric bending pattern associated with forward swimming in Chlamydomonas, although suppf-1 flagella have about one-half the normal beta frequency, apparently as the result of defective function of the outer dynein arms. The pf-17 mutation has been shown to produce nonmotile flagella in which radial spoke heads and five characteristic axonemal polypeptides are missing. Recombinants containing pf-17 and either suppf-2 or suppf-3 have motile flagella, but still lack radial-spoke heads and the associated polypeptides. The flagellar bending pattern of these recombinants lacking radial-spoke heads is a nearly symmetric, large amplitude pattern which is quite unlike the wild-type pattern. However, the presence of an intact radial-spoke system is not required to convert active sliding into bending and is not required for bend initiation and bend propagation, since all of these processes are active in suppfpf-17 recombinants. The function of the radial-spoke system appears to be to convert the symmetric bending pattern displayed by these recombinants into the asymmetric bending pattern required for efficient swimming, by inhibiting the development of reverse bends during the recovery phase of the bending cycle.

scholarly journals Functional reconstitution of Chlamydomonas outer dynein arms from alpha-beta and gamma subunits: requirement of a third factor.

1994 ◽  
Vol 126 (3) ◽  
pp. 737-745 ◽  
Author(s):  
S Takada ◽  
R Kamiya
Keyword(s):  
Cross Section ◽  
Cross Sections ◽  
Attachment Site ◽  
The Other ◽  
Wild Type ◽  
Heavy Chains ◽  
Sds Page ◽  
Gamma Subunits ◽  
Dynein Arms ◽  
Alpha Beta

The outer dynein arm of Chlamydomonas flagella, when isolated under Mg(2+)-free conditions, tends to dissociate into an 11 to 12S particle (12S dynein) containing the gamma heavy chain and a 21S particle (called 18S dynein) containing the alpha and beta heavy chains. We show here that functional outer arms can be reconstituted by the addition of 12S and 18S dyneins to the axonemes of the outer armless mutants oda1-oda6. A third factor that sediments at integral 7S is required for efficient reconstitution of the outer arms on the axonemes of oda1 and oda3. However, this factor is not necessary for reconstitution on the axonemes of oda2, oda4, oda5, and oda6. SDS-PAGE analysis indicates that the axonemes of the former two mutants lack a integral of 70-kD polypeptide that is present in those of the other mutants as well as in the 7S fraction from the wild-type extract. Furthermore, electron micrographs of axonemal cross sections revealed that the latter four mutants, but not oda1 or oda3, have small pointed structures on the outer doublets, at a position in cross section where outer arms normally occur. We suggest that the 7S factor constitutes the pointed structure on the outer doublets and facilitates attachment of the outer arm. The discovery of this structure raises a new question as to how the attachment site for the outer arm dynein is determined within the axoneme.

Developmentally related changes in the production and expression of endo-β-1,4-glucanases in Aspergillus nidulans

Genome ◽  
1989 ◽  
Vol 32 (2) ◽  
pp. 288-292 ◽  
Author(s):  
P. S. Bagga ◽  
S. Sharma ◽  
D. K. Sandhu
Keyword(s):  
Aspergillus Nidulans ◽  
The Other ◽  
Wild Type ◽  
Stages Of Development ◽  
Experimental Conditions ◽  
The Third ◽  
Developmental Mutants ◽  
Mutant Strains ◽  
In The Wild ◽  
Low Levels

The production and electrophoretic expression of endoglucanase(s) were compared in the wild-type and three developmental mutants of Aspergillus nidulans. In the wild type, the production of endoglucanase and its distribution in extracellular and intracellular fractions varied with the age of the culture and the yield was better in stable cultures (production of conidia and cleistothecia) as compared with shake cultures (vegetative hyphae only). Two developmental mutants, aco-T69 and aco-40, which lack the development of conidia and cleistothecia, produced low levels of endoglucanase enzymes as compared with the wild type grown under similar conditions. On the other hand, in aco-90, a mutant capable of producing cleistothecia but no conidia, endoglucanase production was better. The results indicate a correlation between cleistothecial development and endoglucanase level. The electrophoretic studies revealed the presence of three forms of endoglucanase, i.e., EGI, EGII, and EGIII. The first two were detectable in the wild type as well as in mutant strains when grown under various experimental conditions and at all the stages of development. However, the third form could be observed only during cleistothecial development, indicating that this isozyme is developmentally regulated.Key words: endoglucanases, development, Aspergillus nidulans.

Genes other than TLR4 are involved in the response to inhaled LPS

2001 ◽  
Vol 281 (5) ◽  
pp. L1106-L1114 ◽  
Author(s):  
Eva Lorenz ◽  
Michael Jones ◽  
Christine Wohlford-Lenane ◽  
Nicole Meyer ◽  
Kathy L. Frees ◽  
...  
Keyword(s):  
Airway Hyperreactivity ◽  
The Other ◽  
Mouse Strains ◽  
Wild Type ◽  
Conserved Residues ◽  
Biological Responses ◽  
Mutant Strains ◽  
Tlr4 Gene ◽  
Response Phenotype ◽  
The Relationship

For several decades, the mouse strains C3H/HeJ and C57BL/10ScNCr have been known to be hyporesponsive to endotoxin or lipopolysaccharide (LPS). Recently, mutations in Toll-like receptor (TLR) 4 have been shown to underlie this aberrant response to LPS. To further determine the relationship between TLR4 and responsiveness to LPS, we genotyped 18 strains of mice for TLR4 and evaluated the physiological and biological responses of these strains to inhaled LPS. Of the 18 strains tested, 6 were wild type for TLR4 and 12 had mutations in TLR4. Of those strains with TLR4 mutations, nine had mutations in highly conserved residues. Among the strains wild type for TLR4, the inflammatory response in the airway induced by inhalation of LPS showed a phenotype ranging from very sensitive (DBA/2) to hyporesponsive (C57BL/6). A broad spectrum of airway hyperreactivity after inhalation of LPS was also observed among strains wild type for TLR4. Although the TLR4 mutant strains C3H/HeJ and C57BL/10ScNCr were phenotypically distinct from the other strains with mutations in the TLR4 gene, the other strains with mutations for TLR4 demonstrated a broad distribution in their physiological and biological responses to inhaled LPS. The results of our study indicate that although certain TLR4 mutations can be linked to a change in the LPS response phenotype, additional genes are clearly involved in determining the physiological and biological responses to inhaled LPS in mammals.

scholarly journals Two types of Chlamydomonas flagellar mutants missing different components of inner-arm dynein.

1991 ◽  
Vol 112 (3) ◽  
pp. 441-447 ◽  
Author(s):  
R Kamiya ◽  
E Kurimoto ◽  
E Muto
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Cross Section ◽  
Double Mutant ◽  
The Other ◽  
Flagellar Motility ◽  
Wild Type ◽  
Single Group ◽  
Heavy Chains ◽  
Section Electron ◽  
Dynein Arms

Two types of Chlamydomonas reinhardtii flagellar mutants (idaA and idaB) lacking partial components of the inner-arm dynein were isolated by screening mutations that produce paralyzed phenotypes when present in a mutant missing outer-arm dynein. Of the currently identified three inner-arm subspecies I1, I2, and I3, each containing two heterologous heavy chains (Piperno, G., Z. Ramanis, E. F. Smith, and W. S. Sale. 1990. J. Cell Biol. 110:379-389), idaA and idaB lacked I1 and I2, respectively. The 13 idA isolates comprised three genetically different groups (ida1, ida2, ida3) and the two idaB isolates comprised a single group (ida4). In averaged cross-section electron micrographs, inner dynein arms in wild-type axonemes appeared to have two projections pointing to discrete directions. In ida1-3 and ida4 axonemes, on the other hand, either one of them was missing or greatly diminished. Both projections were weak in the double mutant ida1-3 x ida4. These observations suggest that the inner dynein arms in Chlamydomonas axonemes are aligned not in a single straight row, but in a staggered row or two discrete rows. Both ida1-3 and ida4 swam at reduced speed. Thus, the inner-arm subspecies missing in these mutants are not necessary for flagellar motility. However, the double mutants ida1-3 x ida4 were nonmotile, suggesting that axonemes with significant defects in inner arms cannot function. The inner-arm dynein should be important for the generation of axonemal beating.

scholarly journals GENETICS OF YEAST HEXOKINASE

Genetics ◽  
1977 ◽  
Vol 86 (4) ◽  
pp. 727-744
Author(s):  
Zita Lobo ◽  
P K Maitra
Keyword(s):  
Saccharomyces Cerevisiae ◽  
The Other ◽  
Missense Mutations ◽  
Structural Genes ◽  
Wild Type ◽  
Hexokinase Activity ◽  
Mutant Strains ◽  
Mutant Genes ◽  
Yeast Hexokinase ◽  
Respective Wild Type

ABSTRACT Two independent isolates of Saccharomyces cerevisiae lacking hexokinase activity (EC 2.7.1.1) are described. Both mutant strains grow on glucose but are unable to grow on fructose, and contain two mutant genes h×k1 and h×k2 each. The mutations are recessive and noncomplementing. Genetic analysis suggests that these two unlinked genes h×k1 and h×k2 determine, independently of each other, the synthesis of hexokinase isozymes P1 and P2, respectively. h×k1 is located on chromosome VIR distal to met10, and h×k2 is on chromosome IIIR distal to MAL2. Of four hexokinase-positive spontaneous reversions, one is very tightly linked to h×k1 and the other three to the h×k2 locus. The reverted enzymes are considerably more thermolabile than the respective wild-type enzymes, and in one case show altered immunological properties. Data are presented which suggest that the h×k1 and h×k2 mutations are missense mutations in the structural genes of hexokinase P1 and hexokinase P2, respectively. These are presumably the only enzymes that allow S. cerevisiae to grow on fructose.

scholarly journals Hypoxanthine Incorporation Is Nonmutagenic in Escherichia coli

2006 ◽  
Vol 188 (18) ◽  
pp. 6553-6560 ◽  
Author(s):  
Brian Budke ◽  
Andrei Kuzminov
Keyword(s):  
Escherichia Coli ◽  
Nitrous Acid ◽  
Double Mutant ◽  
The Other ◽  
Wild Type ◽  
Purine Base ◽  
Spontaneous Mutagenesis ◽  
Endonuclease V ◽  
Mutant Strains ◽  
Induced Mutagenesis

ABSTRACT Endonuclease V, encoded by the nfi gene, initiates removal of the base analogs hypoxanthine and xanthine from DNA, acting to prevent mutagenesis from purine base deamination within the DNA. On the other hand, the RdgB nucleotide hydrolase in Escherichia coli is proposed to prevent hypoxanthine and xanthine incorporation into DNA by intercepting the noncanonical DNA precursors dITP and dXTP. Because many base analogs are mutagenic when incorporated into DNA, it is intuitive to think of RdgB as acting to prevent similar mutagenesis from deaminated purines in the DNA precursor pools. To test this idea, we used a set of Claire Cupples' strains to detect changes in spontaneous mutagenesis spectra, as well as in nitrous acid-induced mutagenesis spectra, in wild-type cells and in rdgB single, nfi single, and rdgB nfi double mutants. We found neither a significant increase in spontaneous mutagenesis in rdgB and nfi single mutants or the double mutant nor any changes in nitrous acid-induced mutagenesis for rdgB mutant strains. We conclude that incorporation of deaminated purines into DNA is nonmutagenic.

scholarly journals A STUDY OF THE TRANSMISSION AND STRUCTURE OF DOUBLE STRANDED RNAs ASSOCIATED WITH THE KILLER PHENOMENON IN SACCHAROMYCES CEREVISIAE

Genetics ◽  
1976 ◽  
Vol 84 (1) ◽  
pp. 27-42
Author(s):  
T Kevin Sweeney ◽  
Ann Tate ◽  
Gerald R Fink
Keyword(s):  
Electron Microscopy ◽  
Saccharomyces Cerevisiae ◽  
The Other ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Mutant Strains ◽  
Different Types ◽  
Minus Strain

ABSTRACT Killer strains contain two double stranded RNAs, L and M. The M dsRNA appears to be necessary for production of a toxin and for resistance to that toxin. Mutant strains have been found that are defective in their ability to kill and in their resistance to toxin. These sensitive, non-killer strains have altered dsRNA composition. One class has no M dsRNA. Another class of sensitive, non-killers called suppressives has no M dsRNA but instead has smaller dsRNAs called S. In diploids resulting from a cross of a wild-type killer by a suppressive the transmission of the M dsRNA is suppressed by the S dsRNA. When a suppressive is crossed by a strain with no M dsRNA, the diploids and all four meiotic spores have the S dsRNA characteristic of the parental suppressive strain. Suppressive strains do not suppress each other. Intercrosses between two different suppressives yields diploids with both parental S dsRNAs. These two S dsRNAs are transmitted to all 4 meiotic progeny. Another class of mutants has been found which is defective for one of the traits but retains the other. One type, temperature-sensitive killers, has a normal dsRNA composition but is unable to kill at 30°. The other type, immunity-minus, has a complex dsRNA pattern. The immunity-minus strain is extremely unstable during mitotic growth and segregates several different types of non-killers. Analysis of the dsRNAs from wild type and the mutants by electron microscopy shows that the L, M, and S dsRNAs are linear. All strains regardless of killer phenotype appear to have the same size L dsRNA.

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