Relationship between endoplasmic reticulum- and Golgi-associated calcium homeostasis and 4-NQO-induced DNA repair in Saccharomyces cerevisiae

Calcium (Ca²⁺) is an important ion that is necessary for the activation of different DNA repair mechanisms. However, the mechanism by which DNA repair and Ca²⁺ homeostasis cooperate remains unclear. We undertook a systems biology approach to verify the relationship between proteins associated with Ca²⁺ homeostasis and DNA repair for Saccharomyces cerevisiae. Our data indicate that Pmr1p, a Ca²⁺ transporter of Golgi complex, interacts with Cod1p, which regulates Ca²⁺ levels in the endoplasmic reticulum (ER), and with Rad4p, which is a nucleotide excision repair (NER) protein. This information was used to construct single and double mutants defective for Pmr1p, Cod1p, and Rad4p followed by cytotoxic, cytostatic, and cell cycle arrest analyses after cell exposure to different concentrations of 4-nitroquinoline 1-oxide (4-NQO). The results indicated that cod1Δ, cod1Δrad4Δ, and cod1Δpmr1Δ strains have an elevated sensitivity to 4-NQO when compared to its wild-type (WT) strain. Moreover, both cod1Δpmr1Δ and cod1Δrad4Δ strains have a strong arrest at G₂/M phases of cell cycle after 4-NQO treatment, while pmr1Δrad4Δ have a similar sensitivity and cell cycle arrest profile when compared to rad4Δ after 4-NQO exposure. Taken together, our results indicate that deletion in Golgi- and ER-associated Ca²⁺ transporters affect the repair of 4-NQO-induced DNA damage.     

Glutathione peroxidase3 of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> protects phospholipids during cadmium-induced oxidative stress

The present study was undertaken to determine the role of glutathione peroxidase3 (gpx3) in phospholipid protection in cells. Wild-type (WT) cells showed an overall increase in phospholipids upon 50 μM cadmium (Cd)-treatment, whereas an untreated gpx3Δ strain showed a drastic reduction in overall phospholipids which was further reduced with 50 μM Cd. In WT cells, Cd-exposure increased the short chain fatty acids and decreased the unsaturated fatty acids and the magnitude was high in Cd-treated gpx3Δ cells. Purified recombinant gpx3p showed higher activity with phospholipid hydroperoxides than shorter hydroperoxides. An increase in gpx activity was observed in Cd-treated WT cells and no such alteration was observed in gpx3Δ. WT cells treated with Cd showed an increase in MDA over untreated, while untreated gpx3Δ cells themselves showed a higher level of MDA which was further enhanced with Cd-treatment. Iron, zinc and calcium levels were significantly altered in WT and gpx3Δ cells during Cd-treatment.     

Tyroserleutide tripeptide affects calcium homeostasis of human hepatocarcinoma BEL-7402 cells

This study aimed to observe the effects of tyroserleutide (tyrosyl-seryl-leucine, YSL) on the growth of human hepatocarcinoma BEL-7402 that was transplanted into nude mice, and explore its anti-tumor mechanism preliminarily. YSL, at doses of 80 μg-kg^-1 · d^-1, 160 μg·kg^-1 ·d^-1 and 320 μg · kg^-1 · d^-1 significantly inhibited the growth of the human hepatocarcinoma BEL-7402 tumor in nude mice, producing inhibition of 21.66%, 41.34%, and 34.78%, respectively. Ultra structure of BEL-7402 tumor in nude mice showed that YSL could induce tumor cells apoptosis and necrosis, cell organelle mitochondria and endoplasmic reticulum damage, and calcium overload. By confocal laser scanning microscopy and flow cytometry, we found that 10 μg/mL YSL rapidly induced an increase of the concentration of cytoplasmic free calcium in BEL-7402 cells in vitro, and maintained high concentrations of cytoplasmic free calcium for 1 h. Then the calcium concentration began to decrease after 2 h, and was lower than that of the control group at 4 h and 24 h (P< 0.05). YSL also decreased the mitochondrial transmembrane potential of BEL-7402 cells in vitro, but had no effect on the calcium homeostasis or mitochondrial transmembrane potential of Chang liver hepatocytes. So affecting calcium homeostasis, then inducing apoptosis and necrosis may be a mechanism by which YSL inhibits the tumor growth in animal model.     

Effect of aPMR1 disruption on the processing of heterologous glycoproteins secreted in the yeastSaccharomyces cerevisiae

TheSaccharomyces cerevisiae PMR1 gene encodes a Ca²⁺-ATPase localized in the Golgi. We have investigated the effects ofPMR1 disruption inS. cerevisiae on the glycosylation and secretion of three heterologous glycoproteins, human α₁-antitrypsin (α₁-AT), human antithrombin III (ATHIII), andAspergillus niger glucose oxidase (GOD). Thepmr1 null mutant strain secreted larger amounts of ATHIII and GOD proteins per a unit cell mass than the wild type strain. Despite a lower growth rate of thepmr1 mutant, two-fold higher level of human ATHIII was detected in the culture supernatant from thepmr1 mutant compared to that of the wild-type strain. Thepmr1 mutant strain secreted α₁-AT and the GOD proteins mostly as core-glycosylated forms, in contrast to the hyperglycosylated proteins secreted in the wild-type strain. Furthermore, the core-glycosylated forms secreted in thepmr1 mutant migrated slightly faster on SDS-PAGE than those secreted in themnn9 deletion mutant and the wild type strains. Analysis of the recombinant GOD with anti-α1,3-mannose antibody revealed that GOD secreted in thepmr1 mutant did not have terminal α1,3-linked mannoses unlike those secreted in themnn9 mutant and the wild type strains. The present results indicate that thepmr1 mutant, with the super-secretion phenotype, is useful as a host system to produce recombinant glycoproteins lacking high-mannose outer chains.     

Mitochondrial sequestration of BCECF after ester loading in the giant alga Chara australis

Summary. Ratiometric fluorescent dyes are often used to monitor free ion concentrations in vivo, especially in cells that are recalcitrant to transformation with genetically encoded fluorescent markers. Although intracellular dye distributions are often found to be cytosolic, dye localisation has often not been examined in detail. We began exploring the use of BCECF (2′,7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein) to monitor pH in the giant alga Chara australis and discovered that younger leaf cells could be loaded using the acetoxymethyl ester of BCECF. However, we were puzzled to find in microphotometric measurements that the fluorescence ratio appeared insensitive to manipulations affecting cytosolic pH. Confocal imaging of C. australis cells loaded with BCECF showed an accumulation of the dye in two locations: (1) on the outside of the chloroplasts in irregularly shaped stationary bodies; (2) within 1–1.5 μm structures that moved rapidly with the pericellular cytoplasmic streaming. Together with the streaming cytoplasm, these organelles were rendered stationary with 50 μM cytochalasin D. Rhodamine 123, a mitochondrionspecific dye, highlighted organelles outside of the chloroplasts, similar to those shown by BCECF in location 1. We conclude that in the cytoplasmic compartment, BCECF was sequestered within cytoplasmic mitochondria in immature and fast-growing cells and within the cortical mitochondrial system in older and slowly growing cells. Thus, BCECF-AM is unsuitable for reporting changes in cytosolic pH in C. australis but might be employed in future to study pH changes in the mitochondria. Correspondence: M. J. Beilby, Biophysics, School of Physics, University of New South Wales, Sydney 2052, Australia.     

Overexpression of the <Emphasis Type="Italic">PHO84</Emphasis> gene causes heavy metal accumulation and induces Ire1p-dependent unfolded protein response in <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> cells

Pho84p, the protein responsible for the high-affinity uptake and transport of inorganic phosphate across the plasma membrane, is also involved in the low-affinity uptake of heavy metals in the Saccharomyces cerevisiae cells. In the present study, the effect of PHO84 overexpression upon the heavy metal accumulation by yeast cells was investigated. As PHO84 overexpression triggered the Ire1p-dependent unfolded protein response, abundant plasma membrane Pho84p could be achieved only in ire1Δ cells. Under environmental surplus, PHO84 overexpression augmented the metal accumulation by the wild type, accumulation that was exacerbated by the IRE1 deletion. The pmr1Δ cells, lacking the gene that encodes the P-type ATPase ion pump that transports Ca²⁺ and Mn²⁺ into the Golgi, hyperaccumulated Mn²⁺ even from normal medium when overexpressing PHO84, a phenotype which is rather restricted to metal-hyperaccumulating plants.     

Photoreceptor ultrastructure in the Antarctic mussel shrimp Acetabulastoma (Crustacea; Ostracoda), a parasite of Glyptonotus antarcticus (Crustacea; Isopoda)

The ultrastructure of the nauplius eye of the tiny Antarctic ostracode Acetabulastoma sp. is described and conclusions about its possible function are drawn. Each of the three eye-cups measures approximately 20 μm in diameter and is optically isolated from its neighbour by screening pigments, which are contained in pigment cells behind a tapetum of concentrically arranged, ca. 1-μm-long and 0.1-μm-thick, crystals. Three and sometimes four separate rhabdoms with microvilli measuring 50–60 nm in diameter project from the concave side of the tapetum up to 5 μm deep into the eye-cup interior, which is filled by the retinula cell bodies with their spherical nuclei and various organelles. Desmosomes and microtubules are seen and light-induced cell or membrane damage was minimal. The observations suggest that the Acetabulastoma eye has photoreceptors that can tolerate an exposure to bright light and it may be used to inform its owner of the approach of danger, the depth of water, and/or the season. Accepted: 27 August 1998     

Evaluation of the maximum specific growth rate of a yeast indicating non-linear growth trends in batch culture

The maximum specific growth rate (μ_max) of an ethanolic D-xylose-fermenting yeast, Pichia stipitis, showing non-linear growth trends in batch culture, was calculated using the rate equation μ₂ = (1/Δt) ln(x ₂/x ₁). The absolute error Δμ, affecting μ₂, was derived using an equation given by Borzani (1994). Based on the assumption of linearity of growth curves between two closest time points, the relation between the two rate formulae, μ₁ = (1/xˉ)dx _t /dt and μ₂ = (1/Δt) ln(x ₂/x ₁) was established. In a particular condition, when μ₁ = μ₂, an equation has been developed, the roots of which are the specific growth rates at different time points. This revised version was published online in July 2006 with corrections to the Cover Date.     

Biodegradation of dioxins by recombinant Escherichia coli expressing rat CYP1A1 or its mutant

Among polychlorinated dibenzo-p-dioxins (PCDDs), 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TetraCDD) is the most toxic one. Recently, we reported that rat CYP1A1 mutant, F240A, expressed in yeast showed metabolic activity toward 2,3,7,8-TetraCDD. In this study, we successfully expressed N-terminal truncated P450s (Δ1A1 and ΔF240A) in Escherichia coli cells. Kinetic analysis using membrane fractions prepared from the recombinant E. coli cells revealed that ΔF240A has enzymatic properties similar to F240A expressed in yeast. The metabolism of PCDDs by recombinant E. coli cells expressing both ΔF240A and human NADPH-P450 reductase was also examined. When 2,3,7-TriCDD was added to the E. coli cell culture at a final concentration of 10 μM, approximately 90% of the 2,3,7-TriCDD was converted into multiple metabolites within 8 h. These results indicate the possible application of prokaryotic cells expressing ΔF240A to the bioremediation of PCDD-contaminated soil.     

Mitochondrial dysfunction in platelets and hippocampi of senescence-accelerated mice

Senescence-accelerated mice (SAM) strains are useful models to understand the mechanisms of age-dependent degeneration. In this study, measurements of the mitochondrial membrane potential (Δψ_m) of platelets and the Adenosine 5^′-triphosphate (ATP) content of hippocampi and platelets were made, and platelet mitochondria were observed in SAMP8 (faster aging mice) and SAMR1 (aging resistant control mice) at 2, 6 and 9 months of age. In addition, an Aβ-induced (Amyloid beta-protein) damage model of platelets was established. After the addition of Aβ, the Δψ_m of platelets of SAMP8 at 1and 6 months of age were measured. We found that platelet Δψ_m, and hippocampal and platelet ATP content of SAMP8 mice decreased at a relatively early age compared with SAMR1. The platelets of 6 month-old SAMP8 showed a tolerance to Aβ-induced damages. These results suggest that mitochondrial dysfunction might be one of the mechanisms leading to age-associated degeneration in SAMP mice at an early age and the platelets could serve as a biomarker for detection of mitochondrial function and age related disease.     

Citrate Uptake into Tonoplast Vesicles from Acid Lime (Citrus aurantifolia) Juice Cells

Citrate transport into the vacuoles of acid lime juice cells was investigated using isolated tonoplast vesicles. ATP stimulated citrate uptake in the presence or in the absence of a ΔμH⁺. Energization of the vesicles only by an artificial K⁺ gradient (establishing an inside-positive Δψ) also resulted in citrate uptake as was the case of a ΔpH dominated ΔμH⁺. Addition of inhibitors to endomembrane ATPases showed no direct correlation between the inhibition to the tonoplast bound H⁺/ATPase and citrate uptake. The data indicated that, although some citrate uptake can be accounted for by Δψ and by a direct primary active transport mechanism involving ATP, under in vivo conditions of vacuolar pH of 2.0, citrate uptake is driven by ΔpH. Received: 27 April 1998/Revised: 8 September 1998     

Identification of protein kinase disruptions as suppressors of the calcium sensitivity of S. cerevisiae Δptp2 Δmsg5 protein phosphatase double disruptant

The double disruptant of the S. cerevisiae protein phosphatase (PPase) genes, PTP2 (phosphotyrosine-specific PPase) and MSG5 (phosphotyrosine and phosphothreonine/serine-PPase) causes calcium-sensitive growth (Ca^s). Previous study using Fluorescent-activated cell sorting (FACS) analysis showed that this growth defect with calcium occurs at G1–S transition in the cell cycle. We discovered that six non-essential protein kinase (PKase) disruptions (Δbck1, Δmkk1, Δslt2/Δmpk1, Δmck1, Δssk2 and Δyak1) suppressed the Ca^s-phenotype of the Δptp2 Δmsg5 double disruptant. Bck1p, Mkk1p and Slt2p are components of the mitogen-activated protein kinase (MAPK) cascade of cell wall integrity pathway (Slt2 pathway), and Mck1p is its down regulator. Ssk2p is the MAPK kinase kinase of the high-osmolarity glycerol (HOG) pathway, while Yak1p is a negative regulator for the cAMP-dependent PKA pathway. FACS analysis revealed that only the disruption of Δssk2 and Δyak1 but not Δbck1, Δmkk1, Δslt2 and Δmck1 was able to suppress the delayed G1–S transition, suggesting that suppression of the growth defect is not always accompanied by suppression of the G1–S transition delay. The discovery of these PKases as suppressors revealed that in addition to the previously anticipated Slt2 pathway, HOG, Yak1p and Mck1p regulatory pathways may also be involved in the calcium sensitivity of the Δptp2 Δmsg5 double disruptant.     

Energization of <Emphasis Type="Italic">Bacillus subtilis</Emphasis> membrane vesicles increases catalytic activity of succinate: Menaquinone oxidoreductase

In this work, high ΔμH⁺-dependent succinate oxidase activity has been demonstrated for the first time with membrane vesicles isolated from Bacillus subtilis. The maximal specific rate of succinate oxidation by coupled inside-out membrane vesicles isolated from a B. subtilis strain overproducing succinate:menaquinone oxidoreductase approaches the specific rate observed with the intact cells. Deenergization of the membrane vesicles with ionophores or alamethicin brings about an almost complete inhibition of succinate oxidation. An apparent K _m for succinate during the energy-dependent succinate oxidase activity of the vesicles (2.2 mM) is higher by an order of magnitude than the K _m value measured for the energy-independent reduction of 2,6-dichlorophenol indophenol. The data reveal critical importance of ΔμH⁺ for maintaining active electron transfer by succinate:menaquinone oxidoreductase. The role of ΔμH⁺ might consist in providing energy for thermodynamically unfavorable menaquinone reduction by succinate by virtue of transmembrane electron transport within the enzyme down the electric field; alternatively, ΔμH⁺ could play a regulatory role by maintaining the electroneutrally operating enzyme in a catalytically active conformation.     

Probing binding site of bacteriochlorophyll a and carotenoid in the reconstituted LH1 complex from Rhodospirillum rubrum S1 by Stark spectroscopy

Stark spectroscopy is a powerful technique to investigate the electrostatic interactions between pigments as well as between the pigments and the proteins in photosynthetic pigment–protein complexes. In this study, Stark spectroscopy has been used to determine two nonlinear optical parameters (polarizability change Tr(Δα) and static dipole-moment change |Δμ| upon photoexcitation) of isolated and of reconstituted LH1 complexes from the purple photosynthetic bacterium, Rhodospirillum (Rs.) rubrum. The integral LH1 complex was prepared from Rs. rubrum S1, while the reconstituted complex was assembled by addition of purified carotenoid (all-trans-spirilloxanthin) to the monomeric subunit of LH1 from Rs. rubrum S1. The reconstituted LH1 complex has its Q_y absorption maximum at 878 nm. This is shifted to the blue by 3 nm in comparison to the isolated LH1 complex. The energy transfer efficiency from carotenoid to bacteriochlorophyll a (BChl a), which was determined by fluorescence excitation spectroscopy of the reconstituted LH1 complex, is increased to 40%, while the efficiency in the isolated LH1 complex is only 28%. Based on the differences in the values of Tr(Δα) and |Δμ|, between these two preparations, we can calculate the change in the electric field around the BChl a molecules in the two situations to be E _Δ ≈ 3.4 × 10⁵ [V/cm]. This change can explain the 3 nm wavelength shift of the Q_y absorption band in the reconstituted LH1 complex.     

Genetic evaluation of peroxisomal and cytosolic acetoacetyl-CoA thiolase isozymes in n-alkane-assimilating diploid yeast, Candida tropicalis

The n-alkane-assimilating diploid yeast, Candida tropicalis, possesses two acetoacetyl-CoA thiolase (Thiolase I) isozymes encoded by one allele: peroxisomal and cytosolic Thiolase Is encoded by both CT-T1A and CT-T1B. To clarify the function of peroxisomal and cytosolic. Thiolase Is, the site-directed mutation leading Thiolase I ΔC6 without a putative C-terminal peroxisomal targeting signal was introduced on CT-T1A locus in the ct-t1bΔ-null mutant. The C-terminus-truncated Thiolase I was active and solely present in the cytosol. Although the ct-t1aΔ/t1bΔ-null mutants showed mevalonate auxotrophy, the mutants having the C-terminus-truncated Thiolase I did not require mevalonate for growth, as did the strains having cytosolic Thiolase I. These results demonstrated that the presence of Thiolase I in the cytoplasm is indispensable for the sterol synthesis in this yeast. It is of greater interest that peroxisomal and cytosolic Thiolase I isozymes, products of the same genes, play different roles in the respective compartments, although further investigations will be necessary to analyze how to be sorted into peroxisomes and the cytosol.     

Effects of medium composition and nutrient limitation on loss of the recombinant plasmid pLG669-z and β -galactosidase expression by Saccharomyces cerevisiae

The effects of medium composition, nutrient limitation and dilution rate on the loss of the recombinant plasmid pLG669-z and plasmid-borne β -galactosidase expression were studied in batch and chemostat cultures of Saccharomyces cerevisiae strain CGpLG. The difference in growth rates between plasmid-free and plasmid-containing cells (Δμ) and the rate of segregation (R) were determined and some common factors resulting from the effect of medium composition on plasmid loss were identified. Glucose-limited chemostat cultures of CGpLG grown on defined medium were more stable at higher dilution rates and exhibited Δμ -dominated plasmid loss kinetics. Similar cultures grown on complex medium were more stable at lower dilution rates and exhibited R-dominated plasmid loss kinetics. Overall plasmid stability was greatest in phosphate-limited chemostat cultures grown on defined medium and was least stable in magnesium-limited cultures grown on defined medium. Δμ decreased and R increased with increased dilution rate, irrespective of medium composition. Increased plasmid loss rates at high or low dilution rates would appear to be characteristic of loss kinetics dominated by R or Δμ, respectively. Growth of glucose-limited chemostat cultures on complex medium decreased Δμ values but increased R values, in comparison to those cultures grown on defined medium. Any increased stability that a complex medium-induced reduction of Δμ may have conferred was counteracted by an increased R value. Increased β-galactosidase productivity was correlated with increased plasmid stability only in glucose-limited chemostat cultures grown on defined medium and not in those grown on complex medium. Previous studies have yielded contrasting responses with regard to the effect of dilution rate on recombinant plasmid loss from S. cerevisiae. Our findings can account for these differences and may be generally valid for the stability of similar yeast plasmid constructs. This information would facilitate the design of bioprocesses, where recombinant plasmid instability results in reduced culture productivity. Received 08 July 1996/ Accepted in revised form 14 January 1997     

Rescue of Dysfunctional ΔF508-CFTR Chloride Channel Activity by IBMX

Nucleotide-dependent gating of ΔF508-CFTR was evaluated in membrane patches excised from HEK 293 and mouse L-cells and compared to observations on wt-CFTR channels recorded in the same expression systems. ΔF508-CFTR exhibited PKA activated, ATP-dependent channel gating. When compared to wt-CFTR, the K _m for ATP was increased by ninefold (260 μm vs. 28 μm) and maximal open probability (P _o ) was reduced by 49% (0.21 ± 0.06 vs. 0.41 ± 0.02). Additionally, in the absence of PKA, ΔF508-CFTR inactivated over a 1 to 5 min period whereas wt-CFTR remained active. Time-dependent inactivation could be mimicked in wt-CFTR by the intermittent absence of ATP in the cytosolic solution. The effects of 3-isobutyl-1-methyl xanthine (IBMX), a compound reported to stimulate ΔF508-CFTR, were evaluated on wt- and ΔF508-CFTR channels. At concentrations up to 5 mm, IBMX caused a concentration dependent reduction in the observed single channel amplitude (i) of wt-CFTR (maximal observed reduction 35 ± 3%). However, IBMX failed to significantly alter total patch current because of a concomitant 30% increase in P _o . The effects of IBMX on ΔF508-CFTR were similar to effects on wt-CFTR in that i was reduced and P _o was increased by similar magnitudes. Additionally, ΔF508-CFTR channel inactivation was dramatically slowed by IBMX. These results suggest that IBMX interacts with the ATP-bound open state of CFTR to introduce a short-lived nonconducting state which prolongs burst duration and reduces apparent single channel amplitude. A secondary effect observed in ΔF508-CFTR, which may result from this interaction, is a prolongation of the activated state. In light of previously proposed linear kinetic models of CFTR gating, these results suggest that IBMX traps CFTR in an ATP-bound state which may preclude inactivation of ΔF508-CFTR. Received: 5 February 1999/Revised: 25 March 1999     

Isolation and proteomic analysis of the halotolerant alga <Emphasis Type="Italic">Dunaliella salina</Emphasis> flagella using shotgun strategy

Previous studies have demonstrated that flagella/cilia are critical organelles and play diverse roles of motility, sensory perception and development in many eukaryotic cells. However, there is very little information available about flagella composition in Dunaliella salina, a halotolerant, unicellular biflagellate green alga. In the present study, we used strategy of shotgun proteomics to identify flagella proteins after flagella were released and collected from D. salina. A total of 520 groups of proteins were identified under a stringent filter condition (Xcorr ≥1.9, ≥2.2 and ≥3.75; ΔCn ≥ 0.1). In addition to six kinds of known flagella proteins, the putative flagella proteins of D. salina identified by one or more peptides are abundant in signaling, cell division, metabolism, etc. The findings provide guidance for further studies to elucidate the roles of these proteins in the function and assembly of this organelle in microalgae.     

Metabolite profiling studies in Saccharomyces cerevisiae: an assisting tool to prioritize host targets for antiviral drug screening

Background  

The cellular proteins Pat1p, Lsm1p, and Dhh1p are required for the replication of some positive-strand viruses and therefore are potential targets for new antiviral drugs. To prioritize host targets for antiviral drug screening a comparative metabolome analysis in Saccharomyces cerevisiae reference strain BY4742 Matα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 and deletion strains pat1Δ, lsm1Δ and dhh1Δ was performed.     

Comparative Model Analysis of Calcium Exchange between the Cytosol and Stores of Mitochondria or Endoplasmic Reticulum

The objects of the study were single-compartment mathematical models corresponding to a fragment of the dendrite of a cerebellar Purkinje neuron containing the mitochondria (model 1) or a cistern of the endoplasmic reticulum, ER, (model 2) as the calcium stores. We investigated the dependence of the intracellular Ca²⁺ dynamics on geometrical sizes of calcium exchanging parts of the intracellular space and the difference between the kinetic characteristics of storing in two types of stores occupying different portions of the compartment volume. The plasma membrane of the compartment bore the ion channels, particularly those conducting excitatory synaptic current, and the calcium pump typical of this neuron type. The model equations took into account Ca²⁺ exchange between the cytosol, extracellular medium, organelle stores, non-organelle endogenous buffers, and an exogenous buffer (fluorescent dye), and also the diffusion of Са²⁺ into adjacent regions of the dendrite. In model 1, the mitochondria exchanged Са²⁺ with the cytosol via the uniporter and sodium/calcium exchanger; mitochondrial processes, such as the tricarboxylic acid cycle and aerobic cellular respiration, were also taken into account. In model 2, the ER membrane contained the calcium pump, channels of passive leak, and channels of calcium-induced and inositol-3-phosphate-dependent release of Са²⁺. Increases in the portion of the stores in the total volume of the compartment from 1 to 36% led to a proportional increase in the peak values of the cytosolic calcium concentration ([Ca²⁺] _i ); the concentration of Са²⁺ in the mitochondria ([Ca²⁺]_mit) or ER ([Ca²⁺]_ER) increased correspondingly. During generation of bell-shaped cytosolic calcium signals of equal intensity and duration, the ER (due to a greater rate of storing, as compared with that in the mitochondria) was able to uptake several times more Са²⁺ (four times at 36% filling of the volume by the organelles). It is suggested that the revealed different kinetic characteristics of Са²⁺ storing by different organelles are determined by the rates of binding to transport molecules present in the store membrane and, therefore, are defined by concentrations (surface densities) of these molecules and their saturation at certain levels of [Ca²⁺]i. It has been shown that the occupancy of the intracellular volume by organelle stores of any type is a structural factor, which is able to essentially modulate the values of Ca²⁺ concentration.