The GTS1 gene product facilitates the self-organization of the energy metabolism oscillation in the continuous culture of the yeast Saccharomyces cerevisiae

To study the role of the GTS1 gene in the energy metabolism oscillation in continuous cultures of yeast from the physical aspect, time-series data of dissolved oxygen oscillations were analyzed by transforming them into power spectra and by creating two-dimensional trajectories using time delay embedding technique. We found that the wild-type cells organized themselves into a stable limit cycle oscillation and that the GTS1-deleted mutant, gts1Delta, usually showed transient oscillations whose power spectra resembled those of 1/f noise. Thus, we suggested that GTS1 plays an important role in the self-organization of the energy metabolism oscillation.     

Heterogeneous conditions in dissolved oxygen affect N-glycosylation but not productivity of a monoclonal antibody in hybridoma cultures

It is known that heterogeneous conditions exist in large-scale animal cell cultures. However, little is known about how heterogeneities affect cells, productivities, and product quality. To study the effect of non-constant dissolved oxygen tension (DOT), hybridomas were subjected to sinusoidal DOT oscillations in a one-compartment scale-down simulator. Oscillations were forced by manipulating the inlet oxygen partial pressure through a feedback control algorithm in a 220-mL bioreactor maintained at a constant agitation. Such temporal DOT oscillations simulate spatial DOT gradients that can occur in large scales. Different oscillation periods, in the range of 800 to 12,800 s (axis of 7% (air saturation) and amplitude of 7%), were tested and compared to constant DOT (10%) control cultures. Oscillating DOT decreased maximum cell concentrations, cell growth rates, and viability indexes. Cultures at oscillating DOT had an increased glycolytic metabolism that was evidenced by a decrease in yield of cells on glucose and an increase in lactate yield. DOT gradients, even several orders of magnitude higher than those expected under practical large-scale conditions, did not significantly affect the maximum concentration of an IgG(1) monoclonal antibody (MAb). The glycosylation profile of the MAb produced at a constant DOT of 10% was similar to that reported in the literature. However, MAb produced under oscillating culture conditions had a higher amount of triantennary and sialylated glycans, which can interfere with effector functions of the antibody. It was shown that transient excursions of hybridomas to limiting DOT, as occurs in deficiently mixed large-scale bioreactors, is important to culture performance as the oscillation period, and thus the time cells spent at low DOT, affected cell growth, metabolism, and the glycosylation pattern of MAb. Such results underline the importance of monitoring protein characteristics for the development of large-scale processes.     

Action Spectra for the Transient and the Normal Photosynthetic Oxygen Evolution in Wheat Leaves

An action spectrum within the wavelength region 430–700 nm has been determined for an oxygen Iransient isolated from the main photosynthetic oxygen evolution in wheat leaves. the spectrum follows the absorption spectrum of chlorophyll b. In order to compare this oxygen evolving process with the normal photosynthetic oxygen evolution, an action spectrum for this later process was also determined but only in the wavelength region 600–700 nm. This action spectrum closely corresponds to other action spectra for the photosynthetic oxygen evolution found in the literature. The behaviour of the oxygen transient after various treatments was also studied. It was found that the transient oxygen evolution was influenced by the dark period between irradiations, and by a previous irradiation with a wavelength longer than 695 nm, both conditions having a promoting effect on the oxygen evolution. The action spectra and the other findings are discussed on the basis of the two-pigment hypothesis.     

The influence of organic substrates and inhibitors on the induction of a respiratory oscillation in the cyanobacterium Anacystis nidulans

The rate of respiratory oxygen uptake of the cyanobacterium (blue-green alga) Anacystis nidulans oscillated under certain physiological conditions after light pulses or after addition of sodium acetate. The oscillation started either by the photosynthetic inhibition of respiration or by the stimulation of oxygen uptake caused by sodium acetate. The photosynthetic inhibition of respiration decreased the rate of oxygen uptake to about 20% of the rate in the dark.Starved cells (48 h dark) had lost the inducibility of the oscillation. In starved as well as in non-starved cells oscillations were inducible in the presence of fructose or glucose. Well developed oscillations were not promoted further. All other substances tested as substrates did not restore the inducibility of the oscillation in starved cells. The induction of the oscillation was inhibited by iodoacetamide (0.1 mM), p-hydroxymercuribenzoate (0.1 mM) and sodium fluoride (100 mM). It is suggested that a flow from glyceraldehyde-3-phosphate to the incomplete tricarboxylic acid cycle is a prerequisite for the oscillation.     

Pergressive Fourier analysis of encephalograms and renal potassium excretion data

Abstract

Through pergressive Fourier analysis of samples typical properties of this method are demonstrated. The most important difference from other mostly applied methods is that not only frequences (or period lengths, respectively) and amplitude values are used, but also phase relations of hidden oscillations, analogously to optical imaging when, e.g., a laser is added to lenses so that also phase relations of the wave trains in the optical field are used. In this way, from a three dimensional object not a plane picture is produced, but a picture equivalent to the original.

In the pergressive Fourier analysis initial phase and period length of hidden oscillations are accurately obtained through linear regression in a phase diagram, regardless whether the at random selected analysing interval is an integer multiple of the hidden period length, or not. As compared to spectrum and correlation methods, pergressive Fourier analysis produces a consideraly higher accuracy in the calculation of oscillation parameters.     

Clock Control of Ultradian Respiratory Oscillation Found during Yeast Continuous Culture

A short-period autonomous respiratory ultradian oscillation (period approximately 40 min) occurs during aerobic Saccharomyces cerevisiae continuous culture and is most conveniently studied by monitoring dissolved O(2) concentrations. The resulting data are high quality and reveal fundamental information regarding cellular dynamics. The phase diagram and discrete fast Fourier transformation of the dissolved O(2) values revealed a square waveform with at least eight harmonic peaks. Stepwise changes in temperature revealed that the oscillation was temperature compensated at temperatures ranging from 27 to 34 degrees C when either glucose (temperature quotient [Q(10)] = 1.02) or ethanol (Q(10) = 0.82) was used as a carbon source. After alteration of the temperature beyond the temperature compensation region, phase coherence events for individual cells were quickly lost. As the cell doubling rate decreased from 15.5 to 9.2 h (a factor of 1.68), the periodicity decreased by a factor of 1.26. This indicated that there was a degree of nutrient compensation. Outside the range of dilution rates at which stable oscillation occurred, the mode of oscillation changed. The oscillation in respiratory output is therefore under clock control.     

Relationship between outer membrane protein and lipopolysaccharide profiles and serotypes of enterotoxigenic Escherichia coli isolated in Brazil

Abstract When the yeast Saccharomyces cerevisiae was grown under aerobic continuous culture conditions with a medium containing ethanol as carbon source, an autonomous sustained metabolic oscillation appeared. This oscillation was observed in rates and concentrations of various parameters such as, ethanol, oxygen uptake rate, carbon dioxide evolution rate, NaOH addition rate for pH control, acetate, and intracellular pH. No changes were observed in concentrations of stock carbohydrates. Intracellular pH changes were out of phase with oxygen uptake rate, which was reverse of the results with glucose-based oscillation. These results suggested that changes in glycolytic flux and intracellular pH were not regulating the oscillation. Analysis suggested that one of the oscillatory regulation points was located in the ethanol assimilation pathway.     

Asymmetry in the self-sustained oscillation ofPhysarum plasmodial strands

Summary The characteristics of the self-sustained oscillation in the plasmodial strand ofPhysarum polycephalum have been investigated in one steady and two transient conditions. An isolatedPhysarum strand changes its length periodically when it is suspended. In the behaviour of the self-sustained oscillation under the conditions, we provide the first demonstration that the changes in the periods of the oscillation can be ascribed to effects on the shortening phase, while the lengthening periods are almost unaffected. This result means that the asymmetric self-sustained oscillation of thePhysarum strand is composed of an active contracting process, presumably due to actin filaments and myosin-like molecules in the strand, and a passive lengthening process which is merely an extension of the strand under a load.     

An association between vasomotion and oxygen extraction

Vasomotion is defined as a spontaneous local oscillation in vascular tone whose function is unclear but may have a beneficial effect on tissue oxygenation. Optical reflectance spectroscopy and laser Doppler fluximetry provide unique insights into the possible mechanisms of vasomotion in the cutaneous microcirculation through the simultaneous measurement of changes in concentration of oxyhemoglobin ([HbO(2)]), deoxyhemoglobin ([Hb]), and mean blood saturation (S(mb)O(2)) along with blood volume and flux. The effect of vasomotion at frequencies <0.02 Hz attributed to endothelial activity was studied in the dorsal forearm skin of 24 healthy males. Fourier analysis identified periodic fluctuations in S(mb)O(2) in 19 out of 24 subjects, predominantly where skin temperatures were >29.3°C (X(2) = 6.19, P < 0.02). A consistent minimum threshold in S(mb)O(2) (mean: 39.4%, range: 24.0-50.6%) was seen to precede a sudden transient surge in flux, inducing a fast rise in S(mb)O(2). The integral increase in flux correlated with the integral increase in [HbO(2)] (Pearson's correlation r(2) = 0.50, P < 0.001) and with little change in blood volume suggests vasodilation upstream, responding to a low S(mb)O(2) downstream. This transient surge in flux was followed by a sustained period where blood volume and flux remained relatively constant and a steady decrease in [HbO(2)] and equal and opposite increase in [Hb] was considered to provide a measure of oxygen extraction. A measure of this oxygen extraction has been approximated by the mean half-life of the decay in S(mb)O(2) during this period. A comparison of the mean half-life in the 8 normal subjects [body mass index (BMI) <26.0 kg/m(2)] of 12.2 s and the 11 obese subjects (BMI >29.5 kg/m(2)) of 18.8 s was statistically significant (Mann Whitney, P < 0.004). The S(mb)O(2) fluctuated spontaneously in this saw tooth manner by an average of 9.0% (range 4.0-16.2%) from mean S(mb)O(2) values ranging from 30 to 52%. These observations support the hypothesis that red blood cells may act as sensors of local tissue hypoxia, through the oxygenation status of the hemoglobin, and initiate improved local perfusion to the tissue through hypoxic vasodilation.     

Oxygen, pH value, and carbon source induced changes of the mode of oscillation in synchronous continuous culture of Saccharomyces cerevisiae

Oscillations of measured process parameters occur in continuous cultures of Saccharomyces cerevisiae owing to a partial synchronization of budding. Intentional changes of the oxygen concentration, pH value, and carbon source cause effects on the period length similar to those known from variations of the dilution rate. The generation times of parent and daughter cells frequently differ in synchronous culture. To analyze the oscillation the term mode IJ of oscillation is used, which is defined as the ratio IJ of the generation times of parent and daughter cells. When the dissolved oxygen concentration was reduced to zero, the mode of oscillation changed within two periods from mode 12 to mode 11, caused by a decrease of the generation time of daughter cells and an increase of that of the parent cells. When the pH value was slowly reduced from 5.0 to 3.9, a change from mode 112 to mode 13 was observed. Mode 13, representing one parent and three daughter cell populations (the start of budding of each of the three being delayed by one period), denotes an elongated generation time of the daughter cells compared to mode 112, marked by one parent and two different daughter cell classes. When the carbon source galactose was replaced by glucose a mode change from mode 12 to mode 11 was observed. This alteration of the mode was found to be dependent on the status of the cell cycle at the time when the carbon source is changed. The population distribution in batch cultures with glucose or galactose as a substrate was analysed by dyeing the DNA and counting the bud scars. Galactose provoked higher growth rates for the older cells. According to the model for stationary synchronous growth parameters like DO, pH value or the type of carbon source can be varied within a certain range without effecting the period length. If the variation imposes a certain stress, the culture switches to a new mode. These kinds of parameters therefore provide selective measures to influence the period lengths and the modes of oscillation.     

酒精酵母在连续发酵中的振荡行为研究

初步分析酒精酵母在连续发酵中的振荡行为的产生条件及产生机理。通过改变稀释率、pH值、溶氧和进料葡萄糖浓度等条件 ,观察不同操作条件对酒精酵母菌生长和代谢行为的影响。在 10～ 15 g/L的较低葡萄糖浓度 ,0 .10～ 0 .2 0h-1的较低稀释率 ,以及 70 %左右的适度的溶氧浓度等发酵条件下 ,酒精酵母会出现同步的代谢振荡现象。一定条件下 ,菌体浓度处于振荡状态 ,残余葡萄糖浓度不可测或在很低水平振荡 ,这些发现预示着控制机制的新发展。     

A TEMPERATURE-COMPENSATED ULTRADIAN CLOCK OF TETRAHYMENA: OSCILLATIONS IN RESPIRATORY ACTIVITY AND CELL DIVISION

Both a circadian clock and an ultradian clock (period 4—5 h) have previously been described for the ciliated protozoon Tetrahymena. The present communication demonstrates the existence of yet another cellular clock: an ultradian rhythm with a period of about 30 min. The period was found to be well temperature-compensated over the range studied, i.e., between 19°C and 33°C. Ultradian rhythmicity was initiated by dilution of stationary-phase cultures, which were kept previously in a light-dark cycle, into fresh medium. LD treatment during stationary phase was an absolute requirement, since cultures kept in either LL or DD did not produce the ultradian rhythmicity after refeeding. The clock exerts control over respiration; the observed oscillation in oxygen uptake is just a hand of the clock: after a limitation of oxygen supply had ended, the rhythm resumed with the same phase and period as that in control cultures. The clock exerts temporal control also over cell division; in the refed culture cell division resumed with an oscillation in the number of dividing organisms. The period of this oscillation corresponded to that of the rhythm in respiratory activity, indicating that the same ultradian clock may exert control over different cellular functions. Analysis of a second Tetrahymena strain indicates that period length of the ultradian clock is a strain-specific characteristic.     

Autocrine activation of EGF receptor promotes oscillation of glutamate-induced calcium increase in astrocytes cultured in rat cerebral cortex

We previously reported that astrocytes cultured for more than 2 days in a defined medium containing epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) showed calcium oscillation in response to glutamate, whereas the response pattern was transient in the absence of the exogenous growth factors. In the present study, we found that astrocytes showed glutamate-induced calcium oscillation, even in growth factor-free medium, if the cells had been cultured for more than 5 days. The calcium oscillation promoted by the prolonged culture period was suppressed by an inhibitor of EGF receptor tyrosine kinase, but not by a neutralizing antibody to bFGF, indicating that the accumulation of an autocrine factor that activates the EGF receptor leads to calcium oscillation. Astrocytes in our culture system expressed EGF, transforming growth factor alpha (TGFalpha), bFGF and acidic fibroblast growth factor (aFGF). Exogenous aFGF, which induced astrocyte immediate early gene expression to the same extent as EGF or bFGF, did not affect calcium oscillation. Exogenous EGF and bFGF promoted astrocyte hypertrophic morphology and proliferation, as well as calcium oscillation. In contrast, these properties did not accompany calcium oscillation induced by the prolonged culture period. These results suggest that astrocytes possess the ability to promote their own calcium oscillation, which is independent of hypertrophic changes to reactive astrocytes.     

Myocardial sarcomeres spontaneously oscillate with the period of heartbeat under physiological conditions

During heartbeat, the repeated contractions of myocardium are induced by the oscillation of intracellular Ca(2+) concentration. On the other hand, when intermediately activated at a certain Ca(2+) concentration, cardiac myofibrils exhibit the spontaneous sarcomeric oscillation (Ca-SPOC) under steady ionic conditions. In the present study, we found that Ca-SPOC occurred over a wide range of Ca(2+) concentrations, including physiological contractile conditions, in skinned myocardium prepared from various animal species (rat, rabbit, pig, and cow). The period of sarcomeric oscillation fell within the same range as the period of heartbeat of each animal species. On the basis of these results we propose that the intrinsic auto-oscillatory property of sarcomeres (myofibrils) significantly contributes to myocardial beating in vivo.     

Interaction of 1,4-benzoquinones with Photosystem II in thylakoids and Photosystem II membrane fragments from spinach

The interaction of exogenous quinones with the Photosystem II (PS II) acceptor side has been analyzed by measurements of flash-induced 320 nm absorption changes, transient flash-induced variable fluorescence changes, thermoluminescence emission and oxygen yield in dark-adapted thylakoids and PS II membrane fragments. Two classes of 1,4-benzoquinones were shown to give rise to remarkably different reaction patterns. (A) Phenyl-p-benzoquinone (Ph-p-BQ) -type compounds give rise to a marked binary oscillation of the initial amplitudes of 320 nm absorption changes induced by a flash train in dark-adapted PS II membrane fragments and a retardation of the decay kinetics of the flash-induced variable fluorescence. The electron transfer reactions to these type of quinones are severely inhibited by 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU). (B) In the presence of tribromotoluquinone (TBTQ) a different oscillation pattern of the 320 nm absorption changes is observed characterized by a marked relaxation after the first flash in the 5 ms domain. This relaxation is insensitive to 10 μM DCMU. Likewise the decay of the flash-induced variable fluorescence in TBTQ-treated samples is much less sensitive to DCMU than in control. The thermoluminescence emission exhibits an oscillation in samples incubated for 5 min with TBTQ before addition of 30 μM DCMU. Under the same conditions a significant flash-induced oxygen evolution is observed only after the third and fourth flash, respectively, whereas in the presence of TBTQ alone a normal oscillation pattern is observed. The different functional patterns of PS II caused by the two types of classes of exogenous quinones are interpreted by their binding properties: a noncovalent association with the Q_B-site of Ph-p-BQ-type quinones versus a tight (covalent?) binding in the vicinity of Q_A (possibly also at the Q_B-site) in the case of halogenated 1,4-benzoquinones. The mechanistic implications of these findings are discussed.     

Oscillatory response of photosynthesis in leaves to environmental perturbations: a mathematical model

Oscillations in the yield of chlorophyll fluorescence, in oxygen evolution, and in CO2 uptake observed with leaves upon perturbation of steady-state conditions are suggested to be due to the interdependence of turnover of adenylates and Calvin cycle intermediates. This suggestion is quantified in a mathematical model; the behavior of the model system in the neighborhood of the singular point of the system is analyzed. The linearized system is solved analytically, a condition for the occurrence of oscillations is given, and explicit expressions for the oscillation period and the damping constant are derived. The model is shown to be capable of exhibiting oscillations with the period observed with algae or leaves, whereas calculated values of the damping constant are higher than those measured for leaves or algae.     

High-frequency oscillations and circadian rhythm of the membrane potential in Spinach leaves

The microelectrode technique was used to follow oscillations in membrane potential in mesophyll cells of spinach (Spinacia oleracea L.) during exposure do different photoperiodic conditions. Both high-frequency oscillations and circadian variations were observed. The circadian rhythm was imposed on the period of high-frequency oscillation during short days as well as in continuous light: The free-running period was 25.2 h. The average period of high-frequency oscillation increased from 7.64 min in the dark to 19.95 min in the light within several minutes after dark to light transition. This period length coincides with the established period length for oscillations in the redox potential in the chloroplast suspensions of spinach.Abbreviations CL continuous light - SD short day - MP membrane potential     

Rhythmic gene expression in a purple photosynthetic bacterium, Rhodobacter sphaeroides

Circadian rhythms are known to exist in all groups of eukaryotic organisms as well as oxygenic photosynthetic bacteria, cyanobacteria. However, little information is available regarding the existence of rhythmic behaviors in prokaryotes other than cyanobacteria. Here we report biological rhythms of gene expression in a purple bacterium Rhodobacter sphaeroides by using a luciferase reporter gene system. Self-bioluminescent strains of Rb. sphaeroides were constructed, which produced a bacterial luciferase and its substrate, a long chain fatty aldehyde, to sustain the luminescence reaction. After being subjected to a temperature or light entrainment regime, the reporter strains with the luciferase genes driven by an upstream endogenous promoter expressed self-sustained rhythmicity in the constant free-running period. The rhythms were controlled by oxygen and exhibited a circadian period of 20.5 h under aerobic conditions and an ultradian period of 10.6-12.7 h under anaerobic conditions. The data suggest a novel endogenous oscillation mechanism in purple photosynthetic bacteria. Elucidation of the clock-like behavior in purple bacteria has implications in understanding the origin and evolution of circadian rhythms.