Relative biological effectiveness and dose rate effect of tritiated water on chromosomes in human lymphocytes and bone marrow cells

Tritriated water (HTO) is a major toxic effluent from the nuclear power industry, that is released into the environment in large quantities. The low dose radiation effect and dose rate effect of HTO on human lymphocytes and bone marrow cells have not been well studied. The present study was therefore undertaken to investigate the HTO dose-response relationship for chromosomal aberrations in human lymphocytes and bone marrow cells at low in vitro radiation doses ranging from 0.1 to 1 Gy. Lymphocytes (G₀ stage) and bone marrow cells were incubated for 10–150 min with HTO at a dose rate of 2cGy/min (555 MBq/ml). The relative biological effectiveness (RBE) of HTO was calculated with respect to ₆₀Co γ-rays for the induction of dicentric and centric ring chromosomes at low radiation doses. The RBE value for HTO β-rays relative to ⁶⁰Co γ-rays was 2.7 for lymphocytes and 3.1 for chromatid aberrations in bone marrow cells. Lymphocytes were also chronically exposed to HTO for 6.7–80 h at dose rates of 0.5 cGy/min (138.5 MBq/ml) and 0.02 cGy/min (5.6 MBq/ml). There was a 71.5% decrease in the yield of dicentrics and centric rings at the dose rate of 0.02 cGy/min, indicating a clear dose rate effect of HTO. The RBE value for HTO relative to ¹³⁷Cs γ-rays was 2.0 at a dose rate of 0.02 cGy/min, suggesting that low HTO dose rates produce no increase of the RBE values and that the values may be constant between 2 and 3 within these dose rates. These results should prove useful in assessment of the health risk for humans exposed to low levels of HTO.     

Coupling of histone mRNA levels to radioresistant DNA synthesis in ataxia-telangiectasia cells

Cloned genomic DNA for human histone H1, H3 and H4 genes has been used to determine the effects of -radiation on histone mRNA levels and synthesis in ataxia-telangiectasia cells. Synthesis of histone mRNA was determined in cells synchronized with aphidicolin. Effects of irradiation on DNA synthesis and passage through S phase were also monitored. Irradiation was found to slow the passage of control cells through the cell cycle but had no effect on progression of ataxia-telangiectasia cells. H1 and core histone mRNA synthesis was inhibited by radiation in two control cell lines after release from aphidicolin block. No inhibition was observed in one ataxia-telangiectasia cell line and a small degree of inhibition in a second. An increased level of mRNA was observed in both irradiated control and ataxia-telangiectasia cells at 5–7 h post-irradiation compared to unirradiated cells. Similar results were obtained in log phase cells. These results demonstrate that histone mRNA synthesis is radioresistant in ataxia-telangiectasia cells and is coupled to radioresistant DNA synthesis in these cells.     

Uptake and Growth-promoting Activity of Indoleacetic Acid in Segments of Cold-hardened Wheat Coleoptiles

Seedings of winter wheat (Triticum aestivum L. cv. Kharkov MC 22) were grown at 24 C (unhardened) and 4 C (hardened). Indoleacetic acid (IAA) was added to excised coleoptile segments after lengthy incubation and their responses were determined by photometric auxanometry at both 25 C and 5 C. The segments' rates of uptake of ¹⁴CIAA were also compared at both temperatures. Cold hardening had no significant effect on the rates of elongation and uptake in a saturating concentration of IAA (2 to 10 μM) at either temperature. Elongation was more sensitive to temperature of measurement than was uptake. At suboptimal concentrations of IAA and 25 C, hardened coleoptiles took up [2-¹⁴C]-IAA twice as fast but elongated half as fast as unhardened coleoptiles. This and the lack of effect of cold hardening on apparent uptake of [1-¹⁴C]-IAA raised the possibility that a higher rate of IAA-decarboxylation was coupled with the higher rate of uptake of IAA by hardened coleoptiles. Homeostatic hormonal regulation was also evident in the same endogenous rates of elongation of segments of cold-hardened and unhardened coleoptiles.     

Caffeine inhibition of postreplication repair of UV-damaged DNA in mouse epidermal cells

The effect of caffeine (0.25–1.5 mM) on UV-irradiated (5 and 10 J/m²) primary cultures of mouse epidermal cells (EPD) and an in vitro transformed cell line (PDV) was studied at the cellular and molecular levels. A synergistic reduction in cell survival induced by caffeine with UV-irradiation was found in the PDV cells at 10 J/m² but not at 5 J/m². When conversion of low molecular weight newly-synthesized DNA to high molecular weight DNA was studied in both cell types, caffeine at 1.5 mM had no effect on this conversion in unirradiated cultures. At 5 J/m², caffeine had a transitory inhibitory effect on this conversion. However, at 10 J/m² caffeine had a strong permanent inhibitory effect on this conversion at doses higher than 0.5 mM in PDV cells and higher than 0.25 mM in EPD cells. This apparent inhibition of elongation by caffeine in irradiated cells could not be accounted for by an effect on the rate of DNA synthesis. In PDV cells there was a direct correlation in terms of effective caffeine dose level between synergistic reduction in cell survival after UV and the effect on DNA elongation. Irradiated EPD cells were more sensitive to the inhibitory effect of caffeine on DNA elongation.     

Effects of alkylating agents on the DNA replication of cultured Yoshida sarcoma cells

Logarithmically growing Yoshida sarcoma cells were treated for 1 h with low (2 decades cell kill) or high (more than 6 decades cell kill) doses of alkylating agents. Pulse and chase labelled DNA from treated cells were studied by alkaline sucrose gradient centrifugation. Nitrogen mustard (HN-2), 4-hydroperoxycyclophosphamide (CY-OOH), melphalan (L-PAM) and chlorambucil (CA) had no effect on the elongation rate of newly replicated DNA, both at low and high doses, although per cell the rate of DNA synthesis declined as inferred from the rates of [³H] thymidine incorporation compared to the increase in numbers of S phase cells in the treated populations. It is concluded that these drugs act specifically on the initiation step of the DNA replication, leaving chain elongation undisturbed. At low doses the chemically related sulphur mustard (SM) had also no effect on the maturation of new DNA but at high doses a decreased elongation rate was observed. A transient inhibition of chain growth was observed following treatment with a low dose of 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). In contrast, the intercalating agent adriamycin showed a severe but delayed effect resulting in an almost complete block of the maturation.     

Decay of γ-H2AX foci correlates with potentially lethal damage repair and P53 status in human colorectal carcinoma cells

The influence of p53 status on potentially lethal damage repair (PLDR) and DNA double-strand break (DSB) repair was studied in two isogenic human colorectal carcinoma cell lines: RKO (p53 wild-type) and RC10.1 (p53 null). They were treated with different doses of ionizing radiation, and survival and the induction of DNA-DSB were studied. PLDR was determined by using clonogenic assays and then comparing the survival of cells plated immediately with the survival of cells plated 24 h after irradiation. Doses varied from 0 to 8 Gy. Survival curves were analyzed using the linear-quadratic formula: S(D)/S(0) = exp-(αD+βD²). The γ-H2AX foci assay was used to study DNA DSB kinetics. Cells were irradiated with single doses of 0, 0.5, 1 and 2 Gy. Foci levels were studied in non-irradiated control cells and 30 min and 24 h after irradiation. Irradiation was performed with gamma rays from a ¹³⁷Cs source, with a dose rate of 0.5 Gy/min. The RKO cells show higher survival rates after delayed plating than after immediate plating, while no such difference was found for the RC10.1 cells. Functional p53 seems to be a relevant characteristic regarding PLDR for cell survival. Decay of γ-H2AX foci after exposure to ionizing radiation is associated with DSB repair. More residual foci are observed in RC10.1 than in RKO, indicating that decay of γ-H2AX foci correlates with p53 functionality and PLDR in RKO cells.     

γ-Aminobutyric acid-stimulated chloride permeability in crayfish muscle

γ-Aminobutyric acid selectively increased Cl^? permeability in isolated strips of crayfish abdominal muscle. Muscle fibers incubated in VAn Harreveld's solution at room temperature took up ³⁶Cl^? to the extent of 700 ml/kg wet weight with a halftime of 2.5 min. During 15-s incubations, the control ³⁶Cl^? uptake space was 131 ± 4 ml/kg (n = 60) and this was significantly increased by γ-aminobutyric acid at 200 μM or higher concentrations to 177 ± 4 ml/kg (n = 48, P < 0.05). This effect was specific for chloride since γ-aminobutyric acid did not increase the uptake by crayfish muscle of radioactive sucrose, inositol, or propionate. γ-Aminobutyric acid stimulation of ³⁶Cl^? uptake is mediated by receptor-ionophore function since the process shows pharmacological properties virtually identical to those observed by electrophysiological techniques. The γ-aminobutyric acid stimulation of Cl^? permeability is dose dependent with 50% of the maximal effect at 40 μM γ-aminobutyric acid and the dose vs. response curve is somewhat sigmoid. The γ-aminobutyric acid agonist muscimol causes the same maximal effect on Cl^? uptake as γ-aminobutyric acid, but acts at 5-fold lower concentrations, i.e. is more potent. However, the partial agonist γ-amino, β-hydroxybutyric acid produced little or no stimulation of ³⁶Cl^? flux. The response to γ-aminobutyric acid was blocked by 2 mM β-guanidinopropionate or γ-guanidinobutyrate, 0.5 mM bicuculline, and 10 μM picrotoxinin. Picrotoxinin inhibition was dose dependent with 50% inhibition occurring at 4 μM. Antagonists did not affect control ³⁶Cl^? uptake. These results confirm electrophysiological observations that the postsynaptic response to the inhibitory neurotransmitter γ-aminobutyric acid involves a rapid increase in membrane permeability to Cl^?     

The initiation and elongation steps in protein synthesis: Relative rates in chinese hamster ovary cells during and after hyperthermic and hypothermic shocks

The relative rates of the initiation and elongation phases of protein synthesis have been determined in heat- and cold-shocked CHO cells from measurements of the incorporation of ³⁵S-methionine into N-terminal and internal positions of growing peptides by a modified Edman degradation. When the cells are shifted from 37°C to temperatures between 10°C and 34°C, the rate of initiation is at first reduced more extensively than that of elongation. After 20 to 30 minutes at the lower temperature, however, the cells undergo a metabolic adjustment which includes increasing the rate of initiation until it corresponds to the rate of elongation at that temperature. Calculated apparent energies of activation for initiation and elongation are in reasonable agreement with those determined in other mammalian cells. When the cooled cells are returned to 37°C, the rates of initiation and elongation recover immediately but do not exceed the control values. Exposure to elevated temperature (43°C) causes an immediate cessation of initiation and thus a delayed inhibition of elongation; upon return to 37°C, the rate of initiation is transiently elevated above the control rate, and the rate of elongation returns to the control rate after a 2- to 3-minute delay. Hence, a factor which leads to supranormal rates of initiation may accumulate at high but not at low temperatures.     

Excess processing of oxidative damaged bases causes hypersensitivity to oxidative stress and low dose rate irradiation

Abstract

Ionizing radiations such as X-ray and γ-ray can directly or indirectly produce clustered or multiple damages in DNA. Previous studies have reported that overexpression of DNA glycosylases in Escherichia coli (E. coli) and human lymphoblast cells caused increased sensitivity to γ-ray and X-ray irradiation. However, the effects and the mechanisms of other radiation, such as low dose rate radiation, heavy-ion beams, or hydrogen peroxide (H₂O₂), are still poorly understood. In the present study, we constructed a stable HeLaS3 cell line overexpressing human 8-oxoguanine DNA N-glycosylase 1 (hOGG1) protein. We determined the survival of HeLaS3 and HeLaS3/hOGG1 cells exposed to UV, heavy-ion beams, γ-rays, and H₂O₂. The results showed that HeLaS3 cells overexpressing hOGG1 were more sensitive to γ-rays, OH^?, and H₂O₂, but not to UV or heavy-ion beams, than control HeLaS3. We further determined the levels of 8-oxoG foci and of chromosomal double-strand breaks (DSBs) by detecting γ-H2AX foci formation in DNA. The results demonstrated that both γ-rays and H₂O₂ induced 8-oxoguanine (8-oxoG) foci formation in HeLaS3 cells. hOGG1-overexpressing cells had increased amounts of γ-H2AX foci and decreased amounts of 8-oxoG foci compared with HeLaS3 control cells. These results suggest that excess hOGG1 removes the oxidatively damaged 8-oxoG in DNA more efficiently and therefore generates more DSBs. Micronucleus formation also supported this conclusion. Low dose-rate γ-ray effects were also investigated. We first found that overexpression of hOGG1 also caused increased sensitivity to low dose rate γ-ray irradiation. The rate of micronucleus formation supported the notion that low dose rate irradiation increased genome instability.     

Gamma-ray-enhanced reactivation of UV-irradiated adenovirus in normal human fibroblasts.

An enhanced reactivation of UV-irradiated adenovirus type 2 (Ad 2) was detected following irradiation of the host cells with γ-rays prior to infection. Non-irradiated and γ-irradiated normal human fibroblasts were infected immediately after irradiation with either non-irradiated or UV-irradiated Ad 2. At 48h after infection, cultures were examined by indirect immunofluorescence to determine the number cells in which the viral function of viral structural antigen (Vag) was expressed. Pre-irradiation of cells with 1 krad resulted in a 2–3-fold increase in the survival of this viral function following different UV doses to the virus up to 1.75 × 10³ J/m². For a fixed UV dose of 1.0 × 10³ J/m² to the virus this enhancement increased with preirradiation dose to the cells up to a maximum factor of 2–3 for a dose of 1 krad. An examination of Vag expression at various times after infection indicates that pre-irradiation of the cells with γ-rays prior to infection with UV-irradiated virus leads to an earlier onset and/or increased rate of Vag synthesis. This enhancement of Vag production from a UV-damaged template may result from an inducible DNA-repair mechanism in human fibroblasts which may or may not be error-prone.     

Radioresistant DNA synthesis in cells of patients showing increased chromosomal sensitivity to ionizing radiation

The rate of DNA synthesis after γ-irradiation was studied either by analysis of the steady-state distribution of daughter [³H]DNA in alkaline sucrose gradients or by direct assay of the amount of [³H]thymidine incorporated into DNA of fibroblasts derived from a normal donor (LCH882) and from Down's syndrome (LCH944), Werner's syndrome (WS1LE) and xeroderma pigmentosum (XP2LE) patients with chromosomal sensitivity to ionizing radiation. Doses of γ-irradiation that markedly inhibited the rate of DNA synthesis in normal human cells caused almost no inhibition of DNA synthesis in the cells from the affected individuals. The radioresistant DNA synthesis in Down's syndrome cells was mainly due to a much lower inhibition of replicon initiation than that in normal cells; these cells were also more resistant to damage that inhibited replicon elongation. Our data suggest that radioresistant DNA synthesis may be an intrinsic feature of all genetic disorders showing increased radiosensitivity in terms of chromosome aberrations.     

Nitrogen Deprivation Induces Changes in the Leaf Elongation Zone of Maize Seedlings

The influence of nitrogen deprivation on leaf development and the biomechanics of leaf growth were studied using maize (Zea mays L.) seedlings grown under low irradiance. Although the nitrogen deprivation had no significant effect on photosynthesis, the leaf length, the leaf area, and the total assimilation area of plants decreased. The mature size of the epidermal cells was not altered, while the cells of nitrogen-deprived plants reached their final length closer to the leaf base than the epidermal cells of control plants. Decreases in the length of the growing zone (from 50 to 30 mm) and in the maximum value of relative elemental growth rate (from 0.08 to 0.06 mm mm^–1 h^–1) were observed in the nitrogen deprived plants. The maximal value of growth velocity in the control treatment was higher along the elongation zone, except for the basal 20 mm, where there was no significant difference between the control and the N-deprived plants. The net deposition rates of water and dry matter were also affected by nitrogen deprivation: the values of these features decreased and the spatial position of the maximum of the deposition rates shifted towards the leaf base.     

Effect of cytosine arabinoside on replicon initiation in human lymphoblasts

Cytosine arabinoside inhibited DNA synthesis in human lymphoblasts by inhibiting the initiation of DNA replication units. This effect was observed by a decrease in the incorporation of (³H) thymidine into low molecular weight DNA analyzed by velocity sedimentation in alkaline sucrose gradients. In contrast, there was no detectable effect on chain elongation and joining of those molecules that initiated replication before addition of the drug. These data indicate that cytosine arabinoside acts preferentially at the level of initiation of DNA replication rather than chain elongation.     

Sedimentation behavior of solubilized gonadotropin receptor from plasma membranes of bovine corpus luteum

The gonadotropin receptors associated with plasma membrane fractions were solubilized by detergents, including Triton X-100, Lubrol WX, Lubrol PX and sodium deoxycholate before and after equilibration with ¹²⁵I-labelled human chorionic gonadotropin. The binding activity remained in solution even after centrifugation at 300 000 × g for 3 h. The solubilized gonadotropin receptor or gonadotropin receptor complex was characterized by gel filtration and sucrose density gradient centrifugation. Sucrose density gradient centrifugation of solubilized gonadotropin-receptor complex in the presence of Triton X-100 had a sedimentation coefficient of 6.5 S whereas the solubilized uncomplexed receptor had a sedimentation coefficient of 5.1 S. In the absence of the detergent, solubilized hormone receptor complex from plasma membrane fractions I and II sedimented with a apparent sedimentation coefficient of 6.6 S and 7.4 S, respectively. Similary, the free receptor also showed higher sedimentation profile with a apparent sedimentation coefficient of 6.7 S for fraction I and 7.2 S for fraction II. Treatment of plasma membranes with phospholipase A and C inhibited the binding of ¹²⁵I-labelled human chorionic gonadotropin in a dose dependent manner, whereas phospholipase D was without any effect. Doses of 1.4 mI.U. of phospholipase A or 0.6 mI.U. of phospholipase C were required to produce 50% inhibition of the binding activity. These phospholipases had no effect on the performed ¹²⁵I-labelled human chorionic gonadotropin-receptor complex nor on the sedimentation profile of solubilized gonadotropin receptor complex.     

Methylated DNA Causes a Physical Block to Replication Forks Independently of Damage Signalling, O-Methylguanine or DNA Single-Strand Breaks and Results in DNA Damage

Even though DNA alkylating agents have been used for many decades in the treatment of cancer, it remains unclear what happens when replication forks encounter alkylated DNA. Here, we used the DNA fibre assay to study the impact of alkylating agents on replication fork progression. We found that the alkylator methyl methanesulfonate (MMS) inhibits replication elongation in a manner that is dose dependent and related to the overall alkylation grade. Replication forks seem to be completely blocked as no nucleotide incorporation can be detected following 1 h of MMS treatment. A high dose of 5 mM caffeine, inhibiting most DNA damage signalling, decreases replication rates overall but does not reverse MMS-induced replication inhibition, showing that the replication block is independent of DNA damage signalling. Furthermore, the block of replication fork progression does not correlate with the level of DNA single-strand breaks. Overexpression of O⁶-methylguanine (O6meG)-DNA methyltransferase protein, responsible for removing the most toxic alkylation, O6meG, did not affect replication elongation following exposure to N-methyl-N′-nitro-N-nitrosoguanidine. This demonstrates that O6meG lesions are efficiently bypassed in mammalian cells. In addition, we find that MMS-induced γH2AX foci co-localise with 53BP1 foci and newly replicated areas, suggesting that DNA double-strand breaks are formed at MMS-blocked replication forks. Altogether, our data suggest that N-alkylations formed during exposure to alkylating agents physically block replication fork elongation in mammalian cells, causing formation of replication-associated DNA lesions, likely double-strand breaks.     

Control of leaf cell elongation in barley. Generation rates of osmotic pressure and turgor, and growth-associated water potential gradients

In a previous study on the effects of N-supply on leaf cell elongation, the spatial distribution of relative cell elongation rates (RCER), epidermal cell turgor, osmotic pressure (OP) and water potential (Ψ) along the elongation zone of the third leaf of barley was determined (W. Fricke et al. 1997, Planta 202: 522–530). The results suggested that in plants receiving N at fixed relative addition rates (N-supply limitation of growth), cell elongation was rate-limited by the rate of solute provision, whereas in plants growing on complete nutrient solution containing excessive amounts of N (N-demand limitation), cell elongation was rate-limited by the rate of water supply or wall yielding. In the present paper, these suggestions were tested further. The generation rates of cell OP, turgor and Ψ along the elongation zone were calculated by applying the continuity equation of fluid dynamics to the previous data. To allow a more conclusive interpretation of results, anatomical data were collected and bulk solute concentrations determined. The rate of OP generation generally exceeded the rate of turgor generation. As a result, negative values of cell Ψ were created, particularly in demand-limited plants. These plants showed highest RCER along the elongation zone and a Ψ gradient of at least −0.15 MPa between water source (xylem) and expanding epidermal cells. The latter was similar to a theoretically predicted value (−0.18 MPa). Highest rates of OP generation were observed in demand-limited plants, with a maximum rate of 0.112 MPa · h⁻¹ at 16–20 mm from the leaf base. This was almost twice the rate in N-supply-limited plants and implied that the cells in the leaf elongation zone were capable of importing (or synthesising) every minute almost 1 mM of osmolytes. Potassium, Cl⁻ and NO₃ ⁻ were the main inorganic osmolytes (only determined for demand-limited plants). Their concentrations suggest that, unlike the situation in fully expanded epidermal cells, sugars are used to generate OP and turgor. Anatomical data revealed that the zone of lateral cell expansion extended distally beyond the zone of cell elongation. It is concluded that leaf cell expansion in barley relies on high rates of water and solute supply, rates that may not be sustainable during periods of sufficient N-supply (limitation by water supply: Ψ gradients) or limiting N-supply (limitation by solute provision: reduced OP-generation rates). To minimise the possibility of growth limitation by water and osmolyte provision, longitudinal and lateral cell expansion peak at different locations along the growth zone. Received: 15 October 1997 / Accepted: 12 March 1998     

Repair of DNA strand breaks after gamma-irradiation of protoplasts isolated from cultured wild carrot cells

We have isolated protoplasts of cultured wild carrot (Daucus carota L.) cells, lysed them directly on top of alkaline sucrose gradients, and measured single-stranded DNA of molecular weight 1·10⁸ by velocity sedimentation. DNA sedimentation studies on γ-irradiated protoplasts indicate that the energy absorbed in DNA per strand break is 85 eV in air and 260 eV in nitrogen. Isolated wild carrot protoplasts can repair 50% of the DNA strand breaks within 5 min after a dose of 20 krad, and by 1 h none can be detected.     

ANATOMICAL AND PHYSIOLOGICAL STUDIES OF FLORAL TUBE ELONGATION OF CROCUS VERNUS (IRIDACEAE)

Intact floral tubes of Crocus vernus grown under controlled conditions elongated 50 mm in 8 days. Mitoses of the epidermal cells did not occur during the growth of the intact tube; however, cells did elongate from 50 μm to 150 μm, a three-fold increase in cell length. When the floral buds were excised and maintained in distilled water, elongation of floral tubes was inhibited by 46%. The presence of the ovary or the addition of nutrients had no significant effect upon the elongation of the floral tubes of excised buds. When the excised floral buds were placed in 10^–6 m indoleacetic acid, the final tube lengths exceeded that of the water controls by 30% and achieved 91% of the elongation of the intact tubes. Gibberellic acid and kinetin had no effect on floral tube elongation. As with the intact floral tubes, mitoses of the epidermal cells did not occur during the elongation of the excised floral tubes.     

Study of chromatin structure in ataxia-telangiectasia cells

Micrococcal nuclease was used as a probe to study chromatin structure in control and ataxia-telangiectasia cells. The rate and extent of release of acid-soluble nucleotide was similar in both cell types. Production of mono- and oligonucleosomes by micrococcal nuclease as determined by gel electrophoresis also failed to reveal differences in chromatin structure between control and ataxia-telangiectasia cells. Radiation exposure did not significantly alter the kinetics of digestion. These results indicate that there are no gross alterations in chromatin structure in ataxia-telangiectasia cells.     

Effects of Boron Deficiency on Rubidium Uptake and Photosynthesis in the Diatom Cylindrotheca fusiformis

Culturing the diatom Cylindrotheca fusiformis under boron-deficient conditions leads to changes in ⁸⁶Rb uptake and photosynthesis prior to any effect on the rate of cell division. The influx rate of ⁸⁶Rb into boron-deficient cells was 79% of the control rate after 5 to 5.5 hours culture. Despite lowered ⁸⁶Rb influx, however, boron-deficient diatoms accumulated more ⁸⁶Rb than did control cells; this was due to the deficient cells' lower efflux rate. After 24 hours culture, boron-deficient cells had accumulated 30% more ⁸⁶Rb than had control cells, while releasing ⁸⁶Rb at only one-half the control rate. Increased photosynthetic rates were another effect of boron deficiency during this early stage of culture. Prior to 20 hours boron-deficient culture, diatoms had photosynthetic rates 37% greater than those of control cells. Corresponding to the increase in photosynthesis, boron-deficient diatoms had 12% more carbohydrate than control cells after 16 hours culture.