Role of phospholipase D signaling in ethanol-induced inhibition of carbachol-stimulated DNA synthesis of 1321N1 astrocytoma cells

Inhibition of astrocyte proliferation has been suggested to be an important event in the developmental neurotoxicity associated with ethanol. We have previously shown that the acetylcholine analog carbachol induces astroglial cell proliferation through activation of muscarinic M3 receptors, and that ethanol strongly inhibits this effect by inhibiting activation of protein kinase C (PKC) zeta and its down-stream effector 70-kDa ribosomal S6 kinase (p70S6K). In this study, we investigated whether inhibition by ethanol of this signal transduction pathway in 1321N1 human astrocytoma cells may be due, at least in part, to inhibition of the formation of the PKC zeta activator phosphatidic acid (PA), which is formed by hydrolysis of phosphatidylcholine by phospholipase D (PLD). 1-Butanol, which is a substrate for PLD and inhibits PA formation, inhibited carbachol-induced cell proliferation and the underlying intracellular signaling, whereas its analog tert-butanol, which is a poor substrate for PLD, was much less effective. In addition, exogenous PAs were able to increase DNA synthesis and to activate PKC zeta and p70S6K. Furthermore, in carbachol-stimulated cells, ethanol increased the formation of phosphatidylethanol and inhibited the formation of PA. Taken together, these results indicate that PLD activation plays an important role in carbachol-induced astroglial cell proliferation by generating the second messenger PA, which activates PKC zeta. Moreover, the effect of ethanol on carbachol-induced proliferation appears to be mediated, at least in part, by its ability to interact with PLD leading to a decreased synthesis of PA.     

Decrease in thromboxane A2 receptor expression by differentiation with dibutyryl cyclic AMP in 1321N1 human astrocytoma cells.

Thromboxane A2 (TXA2) receptor expression with its signaling was investigated in 1321N1 human astrocytoma cells differentiated with dibutyryl cyclic AMP (dbcAMP). The cells cultured in 0.5% fetal calf serum containing 0.5 mM dbcAMP for 3 days showed the star-shaped morphology, accompanied with the reduction of a TXA2 mimetic U46619-induced phosphoinositide hydrolysis and Ca2+ mobilization. Immunoblotting analysis revealed that human astrocytoma cells expressed phospholipase C (PLC)-beta1 and -beta3, but not PLC-beta2. The contents of PLC-beta1 and beta3 were not changed by the differentiation. The alpha subunit of Gq/ll bound to TXA2-receptor was reduced by the differentiation, determined by immunoblotting after immunoprecipitation with an anti-TXA2-receptor antibody. Scatchard analysis of the binding of [3H]SQ29548, a TXA2 receptor antagonist, to the membranes revealed that the maximum binding site was reduced by the differentiation. The expression of TXA2 receptor mRNA also was reduced by the differentiation, determined by reverse-transcribed-polymerase chain reaction. Although placental type of TXA2 receptor mRNA expression increased after the differentiation, endothelial type of TXA2 receptor mRNA expression slightly decreased. The results suggest that 1321N1 human astrocytoma cells differentiated with dbcAMP show impaired TXA2 receptor-mediated phosphoinositide hydrolysis and Ca2+ mobilization, due to the decrease in TXA2 receptor number.     

Oxidative stress and heat shock stimulate RGS2 expression in 1321N1 astrocytoma cells

RGS2, a regulators of G-protein signaling family member, regulates G-protein signaling and is itself controlled in part by regulated expression. We tested if cell stress regulates RGS2 expression in human astrocytoma 1321N1 cells. Treatment with H2O2 increased RGS2 mRNA levels time- and concentration-dependently, with 200 microM H2O2 causing an approximately eightfold increase after 2 h. Peroxynitrite and heat shock also increased RGS2 mRNA levels. H2O2-induced RGS2 expression was negatively regulated by phosphoinositide-3-kinase and extracellular signal-regulated kinases. H2O2 also concentration-dependently increased RGS2 protein levels, and the RGS2 appeared to be predominantly in the nucleus. These results demonstrate that RGS2 expression is up-regulated by cell stress.     

Nicotinic acetylcholine receptor antagonists alter the function and expression of serine racemase in PC-12 and 1321N1 cells

Western blot analysis demonstrated that PC-12 cells express monomeric and dimeric forms of serine racemase (m-SR, d-SR) and that 1321N1 cells express m-SR. Quantitative RT-PCR and functional studies demonstrated that PC-12 cells express homomeric and heteromeric forms of nicotinic acetylcholine receptors (nAChR) while 1321N1 cells primarily express the α₇-nAChR subtype. The effect of nAChR agonists and antagonists on SR activity and expression was examined by following concentration-dependent changes in intracellular d-Ser levels and SR protein expression. Incubation with (S)-nicotine increased d-Ser levels, which were attenuated by the α₇-nAChR antagonist methyllycaconitine (MLA). Treatment of PC-12 cells with mecamylamine (MEC) produced a bimodal reduction of d-Ser reflecting MEC inhibition of homomeric and heteromeric nAChRs, while a unimodal curve was observed with 1321N1 cells, reflecting predominant expression of α₇-nAChR. The nAChR subtype selectivity was probed using α₇-nAChR selective inhibitors MLA and (R,S)-dehydronorketamine and α₃β₄-nAChR specific inhibitor AT-1001. The compounds reduced d-Ser in PC-12 cells, but only MLA and (R,S)-dehydronorketamine were effective in 1321N1 cells. Incubation of PC-12 and 1321N1 cells with (S)-nicotine, MEC and AT-1001 did not affect m-SR or d-SR expression, while MLA and (R,S)-dehydronorketamine increased m-SR expression but not SR mRNA levels. Treatment with cycloheximide indicated that increased m-SR was due to de novo protein synthesis associated with phospho-active forms of ERK1/2, MARCKS, Akt and rapamycin-sensitive mTOR. This effect was attenuated by treatment with the pharmacological inhibitors U0126, LY294002 and rapamycin, which selectively block the activation of ERK1/2, Akt and mTOR, respectively, and siRNAs directed against ERK1/2, Akt and mTOR. We propose that nAChR-associated changes in Ca^2 + flux affect SR activity, but not expression, and that MLA and (R,S)-dehydronorketamine bind to allosteric sites on the α₇-nAChR and promote multiple signaling cascades that converge at mTOR to increase m-SR levels.     

P2Y6 nucleotide receptor activates PKC to protect 1321N1 astrocytoma cells against tumor necrosis factor-induced apoptosis

1. We recently reported that the activation by UDP of rat P2Y₆ nucleotide receptors expressed in 1321N1 astrocytoma cells protected them from TNF-induced apoptosis by suppressing activation of caspase 3 and 8. This study aims to characterize the involvement of intracellular signaling pathways, including kinases, involved in the antiapoptotic effect of UDP.2. Cell death was induced in 1321N1 astrocytoma cells permanently expressing the rat P2Y₆ receptor by exposure to TNF in the presence of cycloheximide. The apoptotic fraction was analyzed using flow cytometry.3. The activation of P2Y₆ receptors by UDP both protected the astrocytes from TNF- induced apoptosis and activated protein kinase C (PKC) isotypes. The phorbol ester PMA also activated PKC and protected the cells from TNF-induced cell death. The - and -isotypes of PKC were both activated in a persistent fashion upon 5-min exposure to either UDP (10 M) or the phorbol ester PMA (100 nM). The PKC isotype was markedly activated upon UDP treatment.4. The addition of PKC inhibitors, GF109203X or Gö6976, partially antagonized the protective effect of UDP and reduced the UDP-induced phosphorylation of extracellular signal-regulated protein kinases (Erk). The inhibitors of Erk, PD98,059 or U0126, antagonized UDP-induced protection.5. The antiapoptotic protein, Akt, was not affected by P2Y₆ receptor activation. Incubation of the astrocytes with calcium modifiers, BAPTA-AM or dantrolene, did not affect the UDP-induced protection from apoptosis.6. The addition of phospholipase C (PLC) inhibitors, D609 or U73122, partially antagonized both UDP-induced protection and PKC activation.7. Therefore, it is suggested that P2Y₆ receptors in 1321N1 cells, through coupling to PC-PLC and PI-PLC, activate PKC to protect against TNF -induced apoptosis, in which the activation of Erk is involved in part.     

Desensitization and recovery of muscarinic and histaminergic Ca2+ mobilization in 1321N1 astrocytoma cells.

Intracellular free Ca2+ was monitored in suspensions of 1321N1 astrocytoma cells by using the Ca2+ indicator fura-2. The cytoplasmic Ca2+ concentration increased from 237 +/- 6 nM to 1580 +/- 170 nM within 3-5 s of addition of 300 microM-carbachol. After the peak in response, the Ca2+ concentration diminished, establishing a new steady state in about 1 min that was approx. 150 nM above the previous baseline. Histamine increased cytoplasmic Ca2+ to about 40% of the maximal value seen with carbachol. In Ca2+-free buffer each agonist elicited a normal initial increase in cytoplasmic Ca2+, but the sustained portion of the response was abolished. The increase in Ca2+ in response to either carbachol or histamine could be completely inhibited by pretreating the cells with carbachol; the response to carbachol could be partially inhibited by pretreating the cells with histamine. The Ca2+ responses did not recover in the continued presence of carbachol. However, if the carbachol was washed out or if atropine was added after carbachol, the responses to agonist recovered in a time-dependent manner (half-time 3-4 min), and recovery depended on the presence of extracellular calcium. The results indicate that carbachol and histamine stimulate release of Ca2+ from the same intracellular Ca2+ store, that depletion of this store is responsible for heterologous desensitization between these two agonists, and that repletion of the agonist-sensitive Ca2+ pool does not occur in the continued presence of agonist or in the absence of extracellular Ca2+.     

Further evidence that muscarinic cholinergic receptors of 1321N1 astrocytoma cells couple to a guanine nucleotide regulatory protein that is not Ni.

Inhibitory coupling of receptors to adenylate cyclase previously has been shown to be relatively sensitive to inactivation by alkylation with N-ethylmaleimide (NEM). Modification of the inhibitory guanine nucleotide regulatory protein, Ni, has been proposed to be responsible for this effect. The effects of NEM on GTP-sensitive binding of carbachol to muscarinic cholinergic receptors has been compared in a cell line (1321N1 human astrocytoma cells) in which these receptors stimulate phosphoinositide breakdown and in a cell line (NG108-15 neuroblastoma X glioma cells) in which activation of these receptors results in inhibition of adenylate cyclase. Pretreatment of membrane preparations from 1321N1 cells with NEM resulted in a concentration-dependent decrease in the extent of pertussis toxin-catalysed [32P]ADP-ribosylation of a 41 000 Da protein previously proposed to be the alpha subunit of Ni. Under conditions where 32P-labelling of Ni in 1321N1 membranes was reduced by NEM by 90%, no effect was observed on the extent of guanine nucleotide-sensitive high-affinity binding of carbachol to muscarinic cholinergic receptors. In contrast, treatment of NG108-15 membranes with NEM under the same conditions resulted in complete loss of high-affinity guanine nucleotide sensitive binding of carbachol. These results illustrate another difference between the muscarinic receptor population of these two cell lines, and support the previous proposal that muscarinic receptors of 1321N1 cells couple to a guanine nucleotide regulatory protein that is not Ni.     

Increases in intracellular Ca2+ regulate the binding of [3H]phorbol 12,13-dibutyrate to intact 1321N1 astrocytoma cells

The redistribution of protein kinase C (Ca2+/phospholipid-dependent protein kinase) from a cytosolic or a loosely associated membrane compartment to a more integral membrane compartment is stimulated by Ca2+ in vitro. This event is thought to be necessary for activation of the enzyme. To determine whether such a redistribution of protein kinase C occurs following hormonally stimulated increases in cytoplasmic Ca2+, we measured [3H]phorbol 12,13-dibutyrate ([3H]PDB) binding to protein kinase C in intact 1321N1 astrocytoma cells. The muscarinic agonist carbachol causes a 2-fold increase in [3H]PDB binding. This increase is transient, peaking at 1 min and returning toward control levels by 5 min. Scatchard analysis of [3H]PDB binding in the presence of carbachol reveals a 2-fold increase in the Bmax and no change in the KD compared to control values. This increase in Bmax likely represents a redistribution of protein kinase C to the membrane because [3H]PDB binding in intact cells is predominantly to membrane-associated enzyme. The Ca2+ ionophore ionomycin, and two other Ca2+-mobilizing hormones, bradykinin and histamine, mimic the effects of carbachol. Furthermore, when hormone-sensitive Ca2+ stores are depleted by prior agonist treatment, the carbachol-induced increases in intracellular [Ca2+] and [3H]PDB binding are completely blocked. Under these conditions, phosphoinositide hydrolysis and diacylglycerol (DAG) formation are not inhibited. We also examined the time course of DAG accumulation in response to carbachol. DAG is not yet significantly elevated when the increase in [3H]PDB binding is maximal. Furthermore, [3H]PDB binding has returned to control levels when DAG concentrations are maximally elevated. These data suggest that hormone-stimulated increases in cytoplasmic Ca2+ cause a marked and rapid redistribution of protein kinase C which precedes any significant increase in DAG. Our findings also demonstrate that [3H]PDB binding to intact cells may be a useful measure of the ability of Ca2+-mobilizing hormones to affect protein kinase C.     

Characterization of the subtype of muscarinic receptor coupled to the stimulation of phosphoinositide hydrolysis in 132-1N1 human astrocytoma cells.

Stimulation of muscarinic receptors increases phosphoinositide (PI) hydrolysis in 132-1N1 human astrocytoma cells. To evaluate the subtype of receptors which mediate PI hydrolysis in 132-1N1 cells, the effects of: a) the nonselective M1 agonist, carbachol; b) the selective M1 agonist, 4-hydroxy-2-butynyl-trimethylammonium chloride-m-chlorocarbinilate (McN-343); c) the nonselective antagonists, atropine and scopolamine; d) the relatively selective M1 antagonist, pirenzepine; e) the relatively selective M2 antagonists, AF-DX 116 (11-2-diethylaminomethyl-1-piperidinylacetyl-5, 11-dihydro-6H-pyrido-2,3-b-1,4-benzodiazepine-6-one) and methoctramine and f) the relatively selective M3 antagonist, hexahydrosila-difenidol (HHSiD) on PI hydrolysis in 132-1N1 cells were studied. The cell pools of inositol-phospholipids were prelabelled by incubating 132-1N1 cells in a low inositol containing medium (CMRL-1066) supplemented with [3H]inositol (2 microCi/ml) for 20-24 hours at 37 degrees C. The cells were washed and resuspended in a physiological salt solution, and PI hydrolysis was measured by accumulation of [3H]inositol-1-phosphate (IP) in the presence of 10 mM LiCl. Carbachol produced time and concentration dependent PI hydrolysis (EC50, 37 microM). McN-A343 did not cause significant hydrolysis of PI in 132-1N1 cells indicating that the receptor was not of M1 type. All the above muscarinic antagonists caused a concentration dependent decrease in the level of IP in response to carbachol (100 microM). The rank order of their affinities (pA2 values) was: atropine (8.8) > HHSiD (7.6) > pirenzepine (6.8) > methoctramine (6.0) > AF-DX 116 (5.8). This rank order supports the concept that M3 (other names, M2 beta, glandular M2) receptors are linked to PI hydrolysis in 132-1N1 cells. HHSiD, which is selective for M3 receptors of the smooth muscle has higher affinity for muscarinic receptors in 132-1N1 cells than AF-DX 116 which is selective for M2 receptors in cardiac tissue. If the receptor in 132-1N1 cells had been M2, part of the rank order for affinities would have been methoctramine > AF-DX 116 > HHSiD > pirenzepine. From all of these observations, the muscarinic receptor for PI hydrolysis in 132-1N1 cells is tentatively characterized as of M3 type.     

Myometrial effects of selective estrogen receptor modulators on estradiol-responsive gene expression are gene and cell-specific

We examined in vivo effects of selective estrogen receptor modulators (SERMs) 4-OH-tamoxifen (Tam), GW 5638 (GW) and EM-800 (EM) on myometrial gene expression. The uteri of ovariectomized ewes were infused with 10⁻⁷ M of one SERM via indwelling catheters for 24 h preceding hysterectomy. Half of the ewes in each SERM group received an intramuscular injection of 50 μg 17β-estradiol (E2) 18 h prior to hysterectomy. Northern blot analysis and in situ hybridization demonstrated that E2 increased estrogen receptor (ER), progesterone receptor (PR) and cyclophilin (CYC) gene expression in the cells of both inner layer of myometrium (IM) and outer layer of myometrium (OM) as well as glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene expression in OM. Tam also increased ER mRNA levels in OM. EM appeared to increase ER gene expression, but antagonized E2’s up-regulation of PR and CYC gene expression in both IM and OM. Tam and GW also antagonized E2 up-regulation of PR gene expression in OM but not IM. No SERM affected GAPDH gene expression with or without E2. Immunohistochemistry indicated that E2 increased nuclear ER and PR protein levels in both IM and OM. EM was unique in up-regulating ER protein levels, opposite to its effects in endometrial cells. All SERMs tested antagonized this increase in PR immunostaining preferentially in OM compared to the IM layer. These results illustrate gene and cell layer-specific effects of SERMs in sheep myometrium.     

Aflatoxin B1, a selective inhibitor of DNA synthesis in mammalian cells.

Aflatoxin B1 in concentrations between 0.01 and 0.1 microgram/ml inhibits DNA synthesis in African green monkey cells in culture, but has little effect on RNA synthesis and no effect on protein synthesis. The drug even at concentrations up to 1.0 microgram/ml does not interfere with DNA repair promoted by ultraviolet irradiation nor does it induce DNA repair. The inhibition of DNA synthesis attains maximum values 3 h after addition of aflatoxin B1 and is irreversible upon removal of the drug. Profiles of pulselabeled DNA sedimented in alkaline sucrose gradients indicate that aflatoxin B1 blocks initiation of replication rather than elongation.     

Mechanism of killing Chinese hamster ovary cells heated in G1: effects on DNA synthesis and blocking in G2

To determine where in the cell cycle Chinese hamster ovary cells die following heating in G1, a mild hyperthermia treatment, i.e., 10 or 11.5 min at 45.5 degrees C, resulting in 40-50% cell kill was used. After a 7-14-h delay in G1, the cells heated in G1 eventually entered S phase and replicated all their DNA. Both an autoradiographic analysis with tritiated thymidine and a bromodeoxyuridine-propidium iodide bivariate analysis by flow cytometry revealed that both clonogenic and nonclonogenic cells were delayed in progression through S phase for at least 4 h. Then they completed replication of all their DNA and entered G2. Alkaline sucrose gradient sedimentation analysis revealed that these heated cells could complete replicon elongation into cluster-sized molecules of 120-160 S which persisted for 2-12 h after heating. However, further replicon elongation into multicluster-sized molecules greater than 160 S required an additional 12 h in heated cells compared to the 4 h needed in unheated control cells. Our results when compared with the literature suggest that when G1 cells are heated to a survival level of about 50%, the nonclonogenic cells recover from a long delay in G1, traverse S at a reduced rate, and then die either in G2 or as multinucleated cells after an aberrant division.     

Initiation of DNA synthesis in the prematurely condensed chromosomes of G1 cells

The objective of this study was to investigate whether G1 cells could enter S phase after premature chromosome condensation resulting from fusion with mitotic cells. HeLa cell synchronized in early G1, mid-G1, late G1, and G2 and human diploid fibroblasts synchronized in G0 and G1 phases were separately fused by use of UV-inactivated Sendai virus with mitotic HeLa cells. After cell fusion and premature chromosome condensation, the fused cells were incubated in culture medium containing Colcemid (0.05 micrograms/ml) and [3H]thymidine ([3H]ThdR) (0.5 microCi/ml; sp act, 6.7 Ci/mM). At 0, 2, 4, and 6 h after fusion, cell samples were taken to determine the initation of DNA synthesis in the prematurely condensed chromosomes (PCC) on the basis of their morphology and labeling index. The results of this study indicate that PCC from G0, G1, and G2 cells reach the maximum degree of compaction or condensation at 2 h after PCC induction. In addition, the G1-PCC from normal and transformed cells initiated DNA synthesis, as indicated by their "pulverized" appearance and incorporation of [3H]ThdR. Further, the initiation of DNA synthesis in G1-PCC occurred significantly earlier than in the mononucleate G1 cells. Neither pulverization nor incorporation of label was observed in the PCC of G0 and G2 cells. These findings suggest that chromosome decondensation, although not controlling the timing of a cell's entry into S phase, is an important step for the initiation of DNA synthesis. These data also suggest that the entry of a S phase may be regulated by cell cycle phase-specific changes in the permeability of the nuclear envelope to the inducers of DNA synthesis present in the cytoplasm.     

Muscarinic receptors mediate phospholipase C-dependent activation of protein kinase B via Ca2+, ErbB3, and phosphoinositide 3-kinase in 1321N1 astrocytoma cells.

In 1321N1 astrocytoma cells, heterotrimeric G-protein-coupled receptors that activate phosphoinositide-specific phospholipase Cbeta (PLCbeta) isoforms via G(q), induced a prolonged activation of protein kinase B (PKB) after a short delay. For example, the effect of carbachol acting on M3 muscarinic receptors is blocked by wortmannin, suggesting it is mediated via a phosphoinositide 3-kinase (PI 3-kinase). In support of this, carbachol increased PI 3-kinase activity in PI 3-kinase (p85) immunoprecipitates. The pathway linking PLC-coupled receptors to PI 3-kinase was deduced to involve phosphoinositide hydrolysis and Ca2+-dependent ErbB3 transactivation but not protein kinase C on the basis of the following evidence: (i) inhibition of carbachol stimulated PLC by pretreatment with the phorbol ester phorbol 12-myristate 13-acetate concomitantly reduced PKB activity, whereas stimulation of other PLC-coupled receptors also activated PKB; (ii) Ca2+ ionophores and thapsigargin stimulated PKB activity in a wortmannin-sensitive manner, whereas bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid blocked carbachol-stimulated PKB activity; (iii) phorbol 12-myristate 13-acetate alone did not activate PKB, whereas a protein kinase C inhibitor did not prevent the activation of PKB by carbachol; and (iv) carbachol stimulated ErbB3-tyrosine phosphorylation and association with p85, and both these and PKB activity were blocked by tyrphostin AG1478, an epidermal growth factor receptor-tyrosine kinase inhibitor. These experiments define a novel pathway linking G(q)-coupled G-protein-coupled receptors to the activation of PI 3-kinase and PKB.     

Involvement of aquaporin in thromboxane A2 receptor-mediated, G12/13/RhoA/NHE-sensitive cell swelling in 1321N1 human astrocytoma cells

The physiological role of the thromboxane A₂ (TXA₂) receptor expressed on glial cells remains unclear. We previously reported that 1321N1 human astrocytoma cells pretreated with dibutyryl cyclic AMP (dbcAMP) became swollen in response to U46619, a TXA₂ analogue. In the present study, we examined the detailed mechanisms of TXA₂ receptor-mediated cell swelling in 1321N1 cells. The cell swelling caused by U46619 was suppressed by expression of p115-RGS, an inhibitory peptide of Gα_12/13 pathway and C3 toxin, an inhibitory protein for RhoA. The swelling was also inhibited by treatment with Y27632, a Rho kinase inhibitor and 5-(ethyl-N-isopropyl)amiloride (EIPA), a Na⁺/H⁺-exchanger inhibitor. Furthermore, cell swelling was suppressed by the pretreatment with aquaporin inhibitors mercury chloride or phloretin in a concentration-dependent manner, suggesting that aquaporins are involved in U46619-induced 1321N1 cell swelling. In fact, U46619 caused [³H]H₂O influx into the cells, which was inhibited by p115-RGS, C3 toxin, EIPA, mercury chloride and phloretin. This is the first report that the TXA₂ receptor mediates water influx through aquaporins in astrocytoma cells via TXA₂ receptor-mediated activation of Gα_12/13, Rho A, Rho kinase and Na⁺/H⁺-exchanger.     

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.