scholarly journals The metabolic profile of lag and exponential phases of Saccharomyces cerevisiae growth changes continuously

2016 ◽  
Author(s):  
Rafael N. Bento ◽  
Miguel A. Aradhya ◽  
Valdir A. R. Semedo ◽  
Carlos E. S. Bernardes ◽  
Manuel E. M. Piedade ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Metabolic Profile ◽  
Environmental Changes ◽  
Exponential Phase ◽  
Cellular Growth ◽  
Lag Phase ◽  
Continuous Increase ◽  
Flow Microcalorimetry ◽  
Growth Changes ◽  
Two Phases

AbstractCellular growth is usually separated in well-defined phases. For microorganism like Saccharomyces cerevisiae, two phases usually defined are (1) a lag phase, in which no growth is observed and cells adapt to a new environment, followed by (2) an exponential phase, in which rapid proliferation occurs. Here we investigate whether these well-defined phases are uniform. By using flow-microcalorimetry, we found that the metabolic profile of the culture is continuously changing, both in the lag and exponential phases of growth. Along the lag phase there is a continuous increase in the energy that is dissipated irreversibly as heat, while in the exponential phase the opposite occurs. We also confirm recent observations that the oxidative component of metabolism decreases along the exponential phase. Interestingly, nutrient limitation further decreases the amount of energy that is dissipated irreversibly. Altogether, this points to a picture in which cells respond rapidly to minute environmental changes by adjusting their metabolic profile.

Conditions Affecting the Responses of Human Platelets to Epinephrine

1988 ◽  
Vol 60 (02) ◽  
pp. 209-216 ◽  
Author(s):  
Chantal Lalau Keraly ◽  
Raelene L Kinlough-Rathbone ◽  
Marian A Packham ◽  
Hidenori Suzuki ◽  
J Fraser Mustard
Keyword(s):  
Platelet Rich Plasma ◽  
Shape Change ◽  
Human Platelets ◽  
Dense Granule ◽  
Primary Phase ◽  
Lag Phase ◽  
Acid Citrate Dextrose ◽  
Two Phases ◽  
Citrate Dextrose ◽  
Thromboxane Formation

SummaryConditions affecting the responses of human platelets to epinephrine were examined. In platelet-rich plasma prepared from blood anticoagulated with hirudin or PPACK (D-pheny- lalanyl-L-prolyl-L-arginine chloromethyl ketone), epinephrine did not cause shape change or aggregation. In a Tyrode-albumin- apyrase solution containing a concentration of Ca2+ in the physiological range, and fibrinogen, epinephrine in concentrations as high as 40 μM did not induce platelet shape change, caused either no primary aggregation or very slight primary aggregation, and did not induce thromboxane formation, release of dense granule contents, or secondary aggregation. In contrast, in citrated platelet-rich plasma, epinephrine induced two phases of aggregation. This is not attributable to the generation of traces of thrombin since the same effects were evident when blood was taken into a combined citrate-hirudin anticoagulant or a combined citrate-PPACK anticoagulant. In a modified Tyrode-albu- min-apyrase solution containing approximately 20 μM Ca2+, 1 mM Mg2+, and fibrinogen, epinephrine induced extensive aggregation after a lag phase, but no primary phase was evident; thromboxane formation and release of dense granule contents accompanied the aggregation response. These responses were also observed when PPACK was included with the acid-citrate- dextrose anticoagulant, and in the washing and resuspending fluids. In the presence of aspirin or the thromboxane receptor blocker BM 13.177 a few small aggregates were detected by particle counting and by scanning electron microscopy; with the latter inhibitor, the platelets in the aggregates retained their disc shape; secondary aggregation and the responses associated with it did not occur. Thus thromboxane A2 formation is not necessary for the formation of these small aggregates, but is required for extensive aggregation and release. As with other weak agonists, the close platelet-to-platelet contact in the low Ca2+ medium appears to be necessary for full secondary aggregation. Omission of fibrinogen from the low Ca2+ medium prevented both primary and secondary aggregation in response to epinephrine. An antibody (10E5) to the glycoprotein Ilb/IIIa complex was completely inhibitory in the presence of fibrinogen. Thus the response of human platelets to epinephrine is influenced by the concentration of Ca2+ and the presence of fibrinogen in the medium in which they are suspended.

scholarly journals Expansion of EasyClone-MarkerFree toolkit for Saccharomyces cerevisiae genome with new integration sites

2021 ◽  
Author(s):  
Mahsa Babaei ◽  
Luisa Sartori ◽  
Alexey Karpukhin ◽  
Dmitrii Abashkin ◽  
Elena Matrosova ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Recombinant Dna ◽  
Green Fluorescence Protein ◽  
Cellular Growth ◽  
Green Fluorescence ◽  
Biotechnological Production ◽  
Yeast Saccharomyces Cerevisiae ◽  
Integration Sites ◽  
Fluorescence Protein ◽  
Integration Efficiency

Abstract Biotechnological production requires genetically stable recombinant strains. To ensure genomic stability, recombinant DNA is commonly integrated into the genome of the host strain. Multiple genetic tools have been developed for genomic integration into baker's yeast Saccharomyces cerevisiae. Previously, we had developed a vector toolkit EasyClone-MarkerFree for stable integration into eleven sites on chromosomes X, XI, and XII of S. cerevisiae. The markerless integration was enabled by CRISPR-Cas9 system. In this study, we have expanded the kit with eight additional intergenic integration sites located on different chromosomes. The integration efficiency into the new sites was above 80%. The expression level of green fluorescence protein (gfp) for all eight sites was similar or above XI-2 site from the original EasyClone-MarkerFree toolkit. The cellular growth was not affected by the integration into any of the new eight locations. The eight-vector expansion kit is available from AddGene.

scholarly journals Reciprocal hemizygosity analysis reveals that the Saccharomyces cerevisiae CGI121 gene affects lag time duration in synthetic grape must

2021 ◽  
Author(s):  
Runze Li ◽  
Rebecca C Deed
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Low Temperatures ◽  
Lag Time ◽  
Lag Phase ◽  
Phenotypic Traits ◽  
Time Duration ◽  
Phase Duration ◽  
Grape Must ◽  
The Impact ◽  
Reciprocal Hemizygosity Analysis

Abstract It is standard practice to ferment white wines at low temperatures (10-18 °C). However, low temperatures increase fermentation duration and risk of problem ferments, leading to significant costs. The lag duration at fermentation initiation is heavily impacted by temperature; therefore, identification of Saccharomyces cerevisiae genes influencing fermentation kinetics is of interest for winemaking. We selected 28 S. cerevisiae BY4743 single deletants, from a prior list of open reading frames (ORFs) mapped to quantitative trait loci (QTLs) on chromosomes VII and XIII, influencing the duration of fermentative lag time. Five BY4743 deletants, Δapt1, Δcgi121, Δclb6, Δrps17a, and Δvma21, differed significantly in their fermentative lag duration compared to BY4743 in synthetic grape must (SGM) at 15 °C, over 72 h. Fermentation at 12.5 °C for 528 h confirmed the longer lag times of BY4743 Δcgi121, Δrps17a, and Δvma21. These three candidate ORFs were deleted in S. cerevisiae RM11-1a and S288C to perform single reciprocal hemizygosity analysis (RHA). RHA hybrids and single deletants of RM11-1a and S288C were fermented at 12.5 °C in SGM and lag time measurements confirmed that the S288C allele of CGI121 on chromosome XIII, encoding a component of the EKC/KEOPS complex, increased fermentative lag phase duration. Nucleotide sequences of RM11-1a and S288C CGI121 alleles differed by only one synonymous nucleotide, suggesting that intron splicing, codon bias, or positional effects might be responsible for the impact on lag phase duration. This research demonstrates a new role of CGI121 and highlights the applicability of QTL analysis for investigating complex phenotypic traits in yeast.

Growth, development and allometry of Philophthalmus nocturnus in the eyes of domestic chicks

1992 ◽  
Vol 66 (2) ◽  
pp. 100-107 ◽  
Author(s):  
V. G. M. Swarnakumari ◽  
R. Madhavi
Keyword(s):  
Growth Rate ◽  
Developmental Stages ◽  
Adult Stage ◽  
The Body ◽  
Lag Phase ◽  
Allometric Growth ◽  
Body Width ◽  
Rapid Phase ◽  
Growth Development ◽  
Two Phases

ABSTRACTFifty day-old chicks were each infected with 10 excysted metaccreariae of Philophthalimus nocturnus Looss. 1907 around each orbit and growth, development and allometry were studied. The growth rate showed two phases over a period of 35 days, a limited lag phase lasting two days post-infection in which flukes did not exceed 440 μm in length, and a rapid phase during which growth was rapid and flukes reached a size of 3·008–3·504 mm on day 35. Five developmental stages were noticed during the course of development of the metacercaria to the egg-producing adult stage. Eggs appeared in the uterus on day 14 and oculate miracidia on day 25. The hindhody, testes and ovary showed positive allometric growth, the pharnyx less so, whereas negative allometric growth was shown by the forebody. Body width, oral sucker and ventral sucker were close to isometry, growing at the same rate as the body length.

scholarly journals Ser/Thr phospho-regulation by PknB and Stp mediates bacterial quiescence and antibiotic persistence in Staphylococcus aureus

2021 ◽  
Author(s):  
Markus Huemer ◽  
Srikanth Mairpady Shambat ◽  
Sandro Pereira ◽  
Lies Van Gestel ◽  
Judith Bergada-Pijuan ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Ribosomal Proteins ◽  
Environmental Changes ◽  
Acid Stress ◽  
Elongation Factor ◽  
Growth Delay ◽  
Lag Phase ◽  
Cellular Processes ◽  
Translational Activity

Staphylococcus aureus colonizes 30 to 50% of healthy adults and can cause a variety of diseases, ranging from superficial to life-threatening invasive infections such as bacteraemia and endocarditis. Often, these infections are chronic and difficult-to-treat despite adequate antibiotic therapy. Most antibiotics act on metabolically active bacteria in order to eradicate them. Thus, bacteria with minimized energy consumption resulting in metabolic quiescence, have increased tolerance to antibiotics. The most energy intensive process in cells - protein synthesis - is attenuated in bacteria entering into quiescence. Eukaryote-like serine/threonine kinases (STKs) and phosphatases (STPs) can fine-tune essential cellular processes, thereby enabling bacteria to quickly respond to environmental changes and to modulate quiescence. Here, we show that deletion of the only annotated functional STP, named Stp, in S. aureus leads to increased bacterial lag-phase and phenotypic heterogeneity under different stress challenges, including acidic pH, intracellular milieu and in vivo abscess environment. This growth delay was associated with reduced intracellular ATP levels and increased antibiotic persistence. Using phosphopeptide enrichment and mass spectrometry-based proteomics, we identified possible targets of Ser/Thr phosphorylation that regulate cellular processes and bacterial growth, such as ribosomal proteins including the essential translation elongation factor EF-G. Finally, we show that acid stress leads to a reduced translational activity in the stp deletion mutant indicating metabolic quiescence correlating with increased antibiotic persistence.

scholarly journals Electromagnetic Fields (0.04 to 0.39) mT effect on cellular growth cycles of Saccharomyces cerevisiae wine strains

2019 ◽  
Vol 17 (2) ◽  
pp. 196
Author(s):  
Eliseo Amado-González ◽  
Alveiro Álvarez Ovallos ◽  
Alfonso Quijano Parra
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Electromagnetic Fields ◽  
Biomass Production ◽  
Exposure Time ◽  
Culture Media ◽  
Dry Weight ◽  
Cellular Growth ◽  
Biomass Growth ◽  
Yeast Saccharomyces Cerevisiae ◽  
Growth Cycles

Low frecuency electromagnetic fields effect (EMF) on growth cycles of yeast Saccharomyces cerevisiae wine strains Rv1 and Rhône were studied.  A cylindrical coil induced magnetic fields with inductions up to 0,39 mT. Exposure time to EMF varied between (1 – 10) min at 30 °C.  The biomass growth were monitored in the reactor culture media (yeast extract + by measurement optical density from (0 to 32) h. The biomass was found by dry weight. After yeast expose to the different EMF, the number of growth cycles decreased from 4 cycles to 2 or 1. However, the biomass production increased almost 50 %.  The best biomass production was found at 0.39 mT and 10 min exposure time.  Keywords: Electromagnetic fields, Saccharomyces cerevisiae, biomass production, RV1

scholarly journals Different resource allocation in a Bacillus subtilis population displaying bimodal motility

2021 ◽  
Author(s):  
Simon Syvertsson ◽  
Biwen Wang ◽  
Jojet Staal ◽  
Yongqiang Gao ◽  
Remco Kort ◽  
...  
Keyword(s):  
Gene Expression ◽  
Bacillus Subtilis ◽  
Exponential Growth ◽  
Environmental Changes ◽  
Feedback Regulation ◽  
Bet Hedging ◽  
Hedging Strategy ◽  
Negative Feedback Regulation ◽  
Continuous Increase ◽  
Motile Cells

To cope with sudden changes in their environment, bacteria can use a bet-hedging strategy by dividing the population into cells with different properties. This so-called bimodal or bistable cellular differentiation is generally controlled by positive feedback regulation of transcriptional activators. Due to the continuous increase in cell volume, it is difficult for these activators to reach an activation threshold concentration when cells are growing exponentially. This is one reason why bimodal differentiation is primarily observed from the onset of the stationary phase when exponential growth ceases. An exception is the bimodal induction of motility in Bacillus subtilis, which occurs early during exponential growth. Several mechanisms have been put forward to explain this, including double negative-feedback regulation and the stability of the mRNA molecules involved. In this study, we used fluorescence-assisted cell sorting to compare the transcriptome of motile and non-motile cells and noted that expression of ribosomal genes is lower in motile cells. This was confirmed using an unstable GFP reporter fused to the strong ribosomal rpsD promoter. We propose that the reduction in ribosomal gene expression in motile cells is the result of a diversion of cellular resources to the synthesis of the chemotaxis and motility systems. In agreement, single-cell microscopic analysis showed that motile cells are slightly shorter than non-motile cells, an indication of slower growth. We speculate that this growth rate reduction can contribute to the bimodal induction of motility during exponential growth. IMPORTANCE To cope with sudden environmental changes, bacteria can use a bet-hedging strategy and generate different types of cells within a population, so called bimodal differentiation. For example, a Bacillus subtilis culture can contain both motile and non-motile cells. In this study we compared the gene expression between motile and non-motile cells. It appeared that motile cells express less ribosomes. To confirm this, we constructed a ribosomal promoter fusion that enabled us to measure expression of this promoter in individual cells. This reporter fusion confirmed our initial finding. The re-allocation of cellular resources from ribosome synthesis towards synthesis of the motility apparatus results in a reduction in growth. Interestingly, this growth reduction has been shown to stimulate bimodal differentiation.

scholarly journals Isolation and characterization of PEP3, a gene required for vacuolar biogenesis in Saccharomyces cerevisiae

1991 ◽  
Vol 11 (12) ◽  
pp. 5801-5812
Author(s):  
R A Preston ◽  
M F Manolson ◽  
K Becherer ◽  
E Weidenhammer ◽  
D Kirkpatrick ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Product ◽  
Zinc Finger ◽  
Genomic Library ◽  
Sequence Similarity ◽  
Carboxypeptidase Y ◽  
Lag Phase ◽  
Significant Similarity ◽  
Reading Frame ◽  
Isolation And Characterization

The Saccharomyces cerevisiae PEP3 gene was cloned from a wild-type genomic library by complementation of the carboxypeptidase Y deficiency in a pep3-12 strain. Subclone complementation results localized the PEP3 gene to a 3.8-kb DNA fragment. The DNA sequence of the fragment was determined; a 2,754-bp open reading frame predicts that the PEP3 gene product is a hydrophilic, 107-kDa protein that has no significant similarity to any known protein. The PEP3 predicted protein has a zinc finger (CX2CX13CX2C) near its C terminus that has spacing and slight sequence similarity to the adenovirus E1a zinc finger. A radiolabeled PEP3 DNA probe hybridized to an RNA transcript of 3.1 kb in extracts of log-phase and diauxic lag-phase cells. Cells bearing pep3 deletion/disruption alleles were viable, had decreased levels of protease A, protease B, and carboxypeptidase Y antigens, had decreased repressible alkaline phosphatase activity, and contained very few normal vacuolelike organelles by fluorescence microscopy and electron microscopy but had an abundance of extremely small vesicles that stained with carboxyfluorescein diacetate, were severely inhibited for growth at 37 degrees C, and were incapable of sporulating (as homozygotes). Fractionation of cells expressing a bifunctional PEP3::SUC2 fusion protein indicated that the PEP3 gene product is present at low abundance in both log-phase and stationary cells and is a vacuolar peripheral membrane protein. Sequence identity established that PEP3 and VPS18 (J. S. Robinson, T. R. Graham, and S. D. Emr, Mol. Cell. Biol. 11:5813-5824, 1991) are the same gene.

The induction of cytoplasmic petite mutants of Saccharomyces cerevisiae by hydrostatic pressure

1977 ◽  
Vol 26 (1) ◽  
pp. 373-385
Author(s):  
M.P. Rosin ◽  
A.M. Zimmerman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Hydrostatic Pressure ◽  
Cell Survival ◽  
Growth Stage ◽  
Lag Phase ◽  
Pressure Treatment ◽  
Tetrad Analysis ◽  
Petite Mutants ◽  
Wide Range ◽  
Inductive Agent

This study demonstrates that hydrostatic pressure is a potent inductive agent of the petite mutation in cultures of Saccharomyces cerevisiae. The inductive capacity of this mutagen is dependent on the magnitude and the duration of the pressure treatment. Furthermore, the extent of petite induction varies with the growth stage of the culture. Induction occurs in pressure-treated (1-4 X 1-(4) lbf in.-2 or 9–66 X 10(4) kN m-2 for 4 h) log growth cultures but not in stationary or lag phase cultures. Petite induction and cell survival are also dependent on the particular strain of yeast which is pressure-treated. Tetrad analysis and complementation assays demonstrate that pressure-induced petite cells are cytoplasmic in nature. Moreover, induced petite cells show a wide range of suppressivity (2–99%) with a large proportion of the petite cells being highly suppressive.

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