Search PubMed⌕ Search

Biomedical subjects

J E Eriksson

Publications and source records attributed to J E Eriksson.

At least 37 records · Page 2Linked to original sources
PubMed2000

Phosphorylation-Based signaling in Fas receptor-mediated apoptosis.

Apoptosis or programmed cell death plays an essential role during development of the immune system, in immune responses, and in the control of tissue homeostasis in the adult. An important physiological mediator of apoptosis is the Fas/APO-1/CD95 receptor (FasR), a surface receptor belonging to the tumor necrosis factor receptor family. Apoptosis consists of a series of characteristic features that occur following activation of caspases, a collective term for apoptosis-specific proteases. The focus in FasR research has been on determining the mechanisms resulting in caspase activation. However, the role of phosphorylation-based signaling has received increasing attention both as an outcome of FasR activation and as a factor regulating FasR responses. Tyrosine-directed phosphorylation has been implicated to be induced and required during FasR stimulation. The FasR also activates all major signaling pathways that belong to the family of mitogen-activated protein kinase (MAPK) pathways, by either caspase-independent or -dependent mechanisms. Furthermore, phosphorylation-based signaling serves as a potent modifier of FasR responses. In this respect, especially the extracellular signal-regulated kinase and the phosphoinositide 3-kinase signaling pathways have been established as important regulators. This type of control seems to be directly phosphorylation-mediated without the requirement of newly synthesized proteins. Signaling through phosphorylation also regulates the expression of the Fas ligand (FasL), the FasR, as well as various other proteins that affect the outcome of receptor stimulation. While the involvement of phosphorylation has been established in FasR responses, the targets, molecular mechanisms, and biological significance of this aspect of the FasR signaling machinery still require further elucidation.

Apoptosis↗
PubMed1999

Intermediate filament protein partnership in astrocytes.

Intermediate filaments are general constituents of the cytoskeleton. The function of these structures and the requirement for different types of intermediate filament proteins by individual cells are only partly understood. Here we have addressed the role of specific intermediate filament protein partnerships in the formation of intermediate filaments in astrocytes. Astrocytes may express three types of intermediate filament proteins: glial fibrillary acidic protein (GFAP), vimentin, and nestin. We used mice with targeted mutations in the GFAP or vimentin genes, or both, to study the impact of loss of either or both of these proteins on intermediate filament formation in cultured astrocytes and in normal or reactive astrocytes in vivo. We report that nestin cannot form intermediate filaments on its own, that vimentin may form intermediate filaments with either nestin or GFAP as obligatory partners, and that GFAP is the only intermediate filament protein of the three that may form filaments on its own. However, such filaments show abnormal organization. Aberrant intermediate filament formation is linked to diseases affecting epithelial, neuronal, and muscle cells. Here we present models by which the normal and pathogenic functions of intermediate filaments may be elucidated in astrocytes.

Animals↗
PubMed1999

Specific and innervation-regulated expression of the intermediate filament protein nestin at neuromuscular and myotendinous junctions in skeletal muscle.

The intermediate filament proteins nestin, vimentin, and desmin show a specific temporal expression pattern during the development of myofibers from myogenic precursor cells. Nestin and vimentin are actively expressed during early developmental stages to be later down-regulated, vimentin completely and nestin to minimal levels, whereas desmin expression begins later and is maintained in mature myofibers, in which desmin participates in maintaining structural integrity. In this study we have analyzed the expression levels and distribution pattern of nestin in intact and denervated muscle in rat and in human. Nestin immunoreactivity was specifically and focally localized in the sarcoplasm underneath neuromuscular junctions (NMJs) and in the vicinity of the myotendinous junctions (MTJs), ie, in regions associated with acetylcholine receptors (AChRs). This association prompted us to analyze nestin in neurogenically and myogenically denervated muscle. Immunoblot analysis disclosed a marked overall increase of accumulated nestin protein. Similar to the extrajunctional redistribution of AChRs in denervated myofibers, nestin immunoreactivity extended widely beyond the NMJ region. Re-innervation caused complete reversion of these changes. Our study demonstrates that the expression levels and distribution pattern of nestin are regulated by innervation, ie, signal transduction into myofibers.

Adult↗
PubMed1999

Mitotic aberrations induced by carbaryl reflect tyrosine kinase inhibition with coincident up-regulation of serine/threonine protein phosphatase activity: implications for coordination of karyokinesis and cytokinesis.

The insecticide carbaryl and its metabolite 1-naphthol cause partial uncoupling of karyokinesis and cytokinesis in V79 Chinese hamster fibroblasts; karyokinesis is blocked in metaphase, the microtubules of the spindle depolymerize and the chromosomes and spindle remnants become displaced to the periphery of the cell. A high frequency of these disturbed cells elongate and a smaller fraction initiate a cleavage furrow. Here, we attempt to determine the potential targets for carbaryl and 1-naphthol in cytokinesis-specific signalling, led by the fact that the potential protein phosphatase inhibitor 1-naphthyl phosphate was previously identified in treated cells. We found that the typical cytological pattern induced by carbaryl and 1-naphthol could be obtained with tyrphostins, specific tyrosine kinase inhibitors, indicating that the carbaryl-induced effects could be due to tyrosine kinase inhibition. This was confirmed by tyrosine kinase assays showing that carbaryl, 1-naphthol and 2-naphthol were equally efficient at inhibiting tyrosine kinase activity as tyrphostin B44(-). As tyrosine kinases can act as regulatory factors in determining dephosphorylation rates, the activities of type-1 (PP1) and type-2A (PP2A) serine/threonine protein phosphatases were also determined. There was a clear up-regulation of the overall PP1/PP2A activities in cells treated with carbaryl, 1-naphthol or tyrphostin B44(-). This stimulation was shown to be indirect because these compounds had no effect on the activity of purified human PP1 in the test tube. 2-Naphthol, which has been found to be less efficient with regard to displacement of chromatin, did not cause up-regulation, but a significant decrease in PP1/PP2A activity. We suggest that a net decrease in tyrosine kinase activity in combination with a net increase in PP1/PP2A activity is a precondition for cell elongation and cytokinesis in mammalian cells and that the corresponding enzymes are targets in the network of activities serving to coordinate karyokinesis and cytokinesis.

Adenosine Triphosphate↗
PubMed1999

Inhibition of mitogen-activated kinase signaling sensitizes HeLa cells to Fas receptor-mediated apoptosis.

The Fas receptor (FasR) is an important physiological mediator of apoptosis in various tissues and cells. However, there are also many FasR-expressing cell types that are normally resistant to apoptotic signaling through this receptor. The mitogen-activated protein kinase (MAPK) signaling cascade has, apart from being a growth-stimulating factor, lately received attention as an inhibitory factor in apoptosis. In this study, we examined whether MAPK signaling could be involved in protecting FasR-insensitive cells. To this end, we used different approaches to inhibit MAPK signaling in HeLa cells, including treatment with the MAPK kinase inhibitor PD 98059, serum withdrawal, and expression of dominant-interfering MAPK kinase mutant protein. All of these treatments were effective in sensitizing the cells to FasR-induced apoptosis, demonstrating that MAPK indeed is involved in the control of FasR responses. The MAPK-mediated control seemed to occur at or upstream of caspase 8, the initiator caspase in apoptotic FasR responses. Transfection with the constitutively active MAPK kinase abrogated FasR-induced apoptosis also in the presence of cycloheximide, indicating that the MAPK-generated suppression of FasR-mediated apoptotic signaling is protein synthesis independent. In cells insensitive to FasR-induced apoptosis, stimulation of the FasR with an agonistic antibody resulted in significant MAPK activation, which was inhibited by PD 98059. When different cell types were compared, the FasR-mediated MAPK activation seemed proportional to the degree of FasR insensitivity. These results suggest that the FasR insensitivity is likely to be a consequence of FasR-induced MAPK activation, which in turn interferes with caspase activation.

Apoptosis↗
PubMed1998

Suppression of Fas/APO-1-mediated apoptosis by mitogen-activated kinase signaling.

Jurkat T cells undergo rapid apoptosis upon stimulation of the Fas/APO-1 (CD95) receptor. We examined the role of the mitogen-activated protein kinase (MAPK) cascade as a negative regulator of Fas-mediated apoptosis. To this end, we used both physiologic and artificial activators of MAPK, all of which activate MAPK by distinct routes. MAPK activity could be efficiently elevated by two T cell mitogens, the lectin PHA and an agonistic Ab to the T cell receptor complex as well as by the type 1 and 2A phosphatase inhibitor, calyculin A, and the protein kinase C-activating phorbol ester, tetradecanoyl phorbol acetate. All these treatments were effective in preventing the characteristic early and late features of Fas-mediated apoptosis, including activation of caspases. Our results indicate that the elevated MAPK activities intervene upstream of caspase activation. The degree of MAPK activation by the different stimuli used in our study corresponds well to their potency to inhibit apoptosis, indicating that MAPK activation serves as an efficient modulator of Fas-mediated apoptosis. The role of MAPK in modulation of Fas-mediated apoptosis was further corroborated by transient transfection with constitutively active MAPK kinase, resulting in complete inhibition of the Fas response, whereas transfection with a dominant negative form of MAPK kinase had no effect. Furthermore, the apoptosis inhibitory effect of the MAPK activators could be abolished by the specific MAPK kinase inhibitor PD 098059. Modulation of Fas responses by MAPK signaling may determine the persistence of an immune response and may explain the insensitivity of recently activated T cells to Fas receptor stimulation.

Apoptosis↗
PubMed1998

Hsp70 accumulation in chondrocytic cells exposed to high continuous hydrostatic pressure coincides with mRNA stabilization rather than transcriptional activation.

In response to various stress stimuli, heat shock genes are induced to express heat shock proteins (Hsps). Previous studies have revealed that expression of heat shock genes is regulated both at transcriptional and posttranscriptional level, and the rapid transcriptional induction of heat shock genes involves activation of the specific transcription factor, heat shock factor 1 (HSF1). Furthermore, the transcriptional induction can vary in intensity and kinetics in a signal- and cell-type-dependent manner. In this study, we demonstrate that mechanical loading in the form of hydrostatic pressure increases heat shock gene expression in human chondrocyte-like cells. The response to continuous high hydrostatic pressure was characterized by elevated mRNA and protein levels of Hsp70, without activation of HSF1 and transcriptional induction of hsp70 gene. The increased expression of Hsp70 was mediated through stabilization of hsp70 mRNA molecules. Interestingly, in contrast to static pressurization, cyclic hydrostatic loading did not result in the induction of heat shock genes. Our findings show that hsp70 gene expression is regulated posttranscriptionally without transcriptional induction in chondrocyte-like cells upon exposure to high continuous hydrostatic pressure. We suggest that the posttranscriptional regulation in the form of hsp70 mRNA stabilization provides an additional mode of heat shock gene regulation that is likely to be of significant importance in certain forms of stress.

Cartilage↗
PubMed1998

Enhancement of fibroblast collagenase (matrix metalloproteinase-1) gene expression by ceramide is mediated by extracellular signal-regulated and stress-activated protein kinase pathways.

Inflammatory cytokines tumor necrosis factor-alpha and interleukin-1 trigger the ceramide signaling pathway, initiated by neutral sphingomyelinase-elicited hydrolysis of cell membrane phospholipid sphingomyelin to ceramide, a new lipid second messenger. Here, we show that triggering the ceramide pathway by sphingomyelinase or C2- and C6-ceramide enhances collagenase-1 (matrix metalloproteinase-1; MMP-1) gene expression by fibroblasts. C2-ceramide activates three distinct mitogen-activated protein kinases (MAPKs) in dermal fibroblasts, i.e. extracellular signal-regulated kinase 1/2 (ERK1/2), stress-activated protein kinase/Jun N-terminal-kinase (SAPK/JNK), and p38. Stimulation of MMP-1 promoter activity by C2-ceramide is dependent on the presence of a functional AP-1 cis-element and is entirely inhibited by overexpression of MAPK inhibitor, dual specificity phosphatase CL100 (MAPK phosphatase-1). Activation of MMP-1 promoter by C2-ceramide is also effectively inhibited by kinase-deficient forms of ERK1/2 kinase (MEK1/2) activator Raf-1, ERK1 and ERK2, SAPK/JNK activator SEK1, or SAPKbeta. In addition, ceramide-dependent induction of MMP-1 expression is potently prevented by PD 98059, a selective inhibitor of MEK1 activation, and by specific p38 inhibitor SB 203580. These results show that triggering the ceramide signaling pathway activates MMP-1 gene expression via three distinct MAPK pathways, i.e. ERK1/2, SAPK/JNK, and p38, and suggest that targeted modulation of the ceramide signaling pathway may offer a novel therapeutic approach for inhibiting collagenolytic activity, e.g. in inflammatory disorders.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed1998

Protein phosphatase inhibition in normal and keratin 8/18 assembly-incompetent mouse strains supports a functional role of keratin intermediate filaments in preserving hepatocyte integrity.

The function and regulation of keratin 8 (K8) and 18 (K18), intermediate filament (IF) proteins of the liver, are not fully understood. We employed the liver damage induced by microcystin-LR (MC-LR), a liver-specific inhibitor of type-1 and type-2A protein phosphatases, in normal and in keratin assembly-incompetent mouse strains as a model to elucidate the roles of IF phosphorylation in situ. The mouse strains used were wild-type (wt) mice and mice with abnormal filament assembly, caused by a targeted null mutation of the K8 gene or caused by expression of a point-mutated dominant negative human K18. In vivo 32P-labeled wt mice, subsequently injected with a lethal dose of MC-LR, showed hyperphosphorylation, disassembly, and reorganization of K8/K18, in particular K18, indicating high phosphate turnover on liver keratins in situ. At lethal doses, the keratin assembly-incompetent mice displayed liver lesions faster than wt mice, as indicated histopathologically and by liver-specific plasma enzyme elevations. The histological changes included centrilobular hemorrhage in all mouse strains. The assembly-incompetent mice showed a marked vacuolization of periportal hepatocytes. Indistinguishable MC-LR-induced reorganization of microfilaments was observed in all mice, indicating that this effect on microfilaments is not dependent on the presence of functional K8/K18 networks. At sublethal doses of MC-LR, all animals had the same potential to recover from the liver damage. Our study shows that K8/K18 filament assembly is regulated in vivo by serine phosphorylation. The absence or occurrence of defective K8/K18 filaments render animals more prone to liver damage, which supports the previously suggested roles of keratin IFs in maintenance of structural integrity.

Animals↗
PubMed1998

Enhancement of fibroblast collagenase-1 (MMP-1) gene expression by tumor promoter okadaic acid is mediated by stress-activated protein kinases Jun N-terminal kinase and p38.

Collagenase-1 (matrix metalloproteinase-1, MMP-1) is expressed by several types of cells, including fibroblasts, and apparently plays an important role in the remodeling of collagenous extracellular matrix in various physiologic and pathologic situations. Here, we have examined the molecular mechanisms of the activation of fibroblast MMP-1 gene expression by a naturally occurring non-phorbol ester type tumor promoter okadaic acid (OA), a potent inhibitor of serine/threonine protein phosphatase 2A. We show that in fibroblasts OA activates three distinct subgroups of mitogen activated protein kinases (MAPKs): extracellular signal-regulated kinase1,2 (ERK1,2), c-Jun N-terminal-kinase/stress-activated protein kinase (JNK/SAPK) and p38. Activation of MMP-1 promoter by OA is entirely blocked by overexpression of dual-specificity MAPK phosphatase CL100. In addition, expression of kinase-deficient forms of ERK1,2, SAPKbeta, p38, or JNK/SAPK kinase SEK1 strongly inhibited OA-elicited activation of MMP-1 promoter. OA-elicited enhancement of MMP-1 mRNA abundance was also strongly prevented by two chemical MAPK inhibitors: PD 98059, a specific inhibitor of the activation of ERK1,2 kinases MEK1,2; and SB 203580, a selective inhibitor of p38 activity. Results of this study show that MMP-1 gene expression in fibroblasts is coordinately regulated by ERK1,2, JNK/SAPK, and p38 MAPKs and suggest an important role for the stress-activated MAPKs JNK/SAPK and p38 in the activation of MMP-1 gene expression. Based on these observations, it is conceivable that specific inhibition of stress-activated MAPK pathways may serve as a novel therapeutic target for inhibiting degradation of collagenous extracellular matrix.

3T3 Cells↗
PubMed1998

Conventional and novel PKC isoenzymes modify the heat-induced stress response but are not activated by heat shock.

In mammalian cells, the heat-induced stress response is mediated by the constitutively expressed heat shock transcription factor 1 (HSF1). Upon exposure to elevated temperatures, HSF1 undergoes several post-translational modifications, including inducible phosphorylation or hyperphosphorylation. To date, neither the role of HSF1 hyperphosphorylation in regulation of the transcriptional activity of HSF1 nor the signaling pathways involved have been characterized. We have previously shown that the protein kinase C (PKC) activator, 12-O-tetradecanoylphorbol 13-acetate (TPA), markedly enhances the heat-induced stress response, and in the present study we elucidate the mechanism by which PKC activation affects the heat shock response in human cells. Our results show that several conventional and novel PKC isoenzymes are activated during the TPA-mediated enhancement of the heat shock response and that the enhancement can be inhibited by the specific PKC inhibitor bisindolylmaleimide I. Furthermore, the potentiating effect of TPA on the heat-induced stress response requires an intact heat shock element in the hsp70 promoter, indicating that PKC-responsive pathways are able to modulate the activity of HSF1. We also demonstrate that PKC is not activated by heat stress per se. These results reveal that PKC exhibits a significant modulatory role of the heat-induced stress response, but is not directly involved in regulation of the heat shock response.

Carcinogens↗
PubMed1997

Thioredoxin is transcriptionally induced upon activation of heat shock factor 2.

Heat shock gene expression is differentially regulated in cells exposed to stress stimuli and in cells undergoing processes of differentiation and development. Regulation of the classical heat shock response is mediated by heat shock factor 1 (HSF1), whereas heat shock factor 2 (HSF2) is activated in certain differentiating cells, for example during hemin-mediated differentiation of human K562 erythroleukemia cells. Hence, the signaling pathways leading to induction of heat shock gene expression upon different stimuli are likely to be distinct. We have used RNA arbitrarily primed polymerase chain reaction to identify genes that are differentially regulated upon activation of HSF1 and HSF2. In this study, we report that thioredoxin (TRX) expression is induced in K562 cells in response to hemin in an HSF2-dependent manner. Increased TRX expression was primarily detected on the transcriptional level, subsequently leading to elevated TRX mRNA and protein levels. Hemin treatment caused no reduction in cellular glutathione concentrations, indicating that the increased TRX expression was not due to oxidative stress. Studies using cell lines where overexpression of the HSF2-beta isoform represses HSF2 activation implied that active HSF2 is required for transcriptional induction of TRX. Unlike HSF2, activation of HSF1 did not induce TRX expression. Taken together, our results suggest that HSF1 and HSF2 may regulate distinct target genes, and activation of HSF2 could be involved in the regulation of TRX expression during hemin-mediated differentiation of K562 cells.

Cell Differentiation↗
PubMed1997

The phorbol ester 12-O-tetradecanoylphorbol 13-acetate enhances the heat-induced stress response.

Induction of heat shock gene expression is mediated by specific heat shock transcription factors (HSFs), but the signaling pathways leading to activation of HSFs are poorly understood. To elucidate whether protein kinase C-responsive signaling pathways could be involved in the regulation of heat shock gene expression, we have examined the effects of the protein kinase C activator 12-O-tetradecanoylphorbol 13-acetate (TPA) on the heat-induced stress response in K562 cells. We demonstrate that TPA treatment markedly enhances heat shock gene expression during heat stress, although TPA alone does not induce the heat shock response. This TPA-mediated enhancement can initially be detected as an accelerated acquisition of DNA binding and transcriptional activity of HSF1 resulting in elevated Hsp70 protein concentrations. In the presence of TPA, the attenuation of HSF1 DNA binding activity during continuous exposure to heat shock occurs more rapidly and in concert with the appearance of newly synthesized Hsp70, which supports earlier studies on the autoregulatory role of Hsp70 in deactivation of HSF1. During heat stress, a correlation between the hyperphosphorylation of HSF1 and its transcriptional activity was observed, in both the presence and the absence of TPA. Our results show that the heat-induced stress response can be significantly modulated by activation of protein kinase C-responsive signaling pathways.

DNA↗
PubMed1997

The intermediate filament protein nestin occurs transiently in differentiating testis of rat and mouse.

Nestin is an intermediate filament (IF) protein (IFP) which occurs during early developmental stages and during regenerative processes in muscle and neuronal cells. The spatial and temporal localization of nestin in the developing testis of rat and mouse was studied by immunolabeling light and electron microscopy and by immunoblotting. Nestin localization was related to the localization of the other major IFPs specific for this tissue, i.e. cytokeratins, vimentin and desmin. Laminin immunocytochemistry and conventional microscopy were used to identify tissues and cells. With the incipient differentiation of the gonadal anlage, the reaction for nestin was weak in the gonadal ridge, whereas the cells of the mesonephric mesenchyme showed a prominent reaction for this IFP. The nestin-specific reaction in the epithelial mesonephric duct and tubules was weak and disappeared at an early phase of differentiation. With the development of the testis proper, nestin was transiently found in several cell types. Nestin was found as well as vimentin and cytokeratins in the Sertoli cells. In the interstitial cells nestin was found together with vimentin and desmin IFPs, and was most prominent in the differentiating myoid cells. After birth, nestin gradually disappeared from the testicular cells and in the rat at puberty was found only in the endothelial cells of some blood vessels. The abolished nestin synthesis in the testis was confirmed by immunoblotting. These results suggest that nestin is required transiently during the development of the testis and mesonephros. The temporary presence of nestin, and several other IFPs during these phases, coincides with key phases of urogenital sex differentiation. This may imply that the orchestrated synthesis of the IFPs nestin, cytokeratins, vimentin and desmin is likely to be linked with the genes regulating sex differentiation.

Animals↗
PubMed1997

Protein phosphatases maintain the organization and structural interactions of hepatic keratin intermediate filaments.

The importance of protein phosphatases in the maintenance of cytoskeletal structure is supported by the serious liver injury caused by microcystin-LR, a hepatotoxic inhibitor of type-1 and type-2A serine/threonine protein phosphatases. We used the microcystin-LR-induced cell injury as a model to study the roles of protein dephosphorylation in maintaining cytoskeletal structure and cellular interactions in primary rat hepatocyte cultures. Confocal microscopy revealed that the first visible effect of microcystin-LR is disruption of desmoplakin organization at the cell surface, indicating dissociation of desmosomes. This effect is followed by a dramatic reorganization of both the intermediate filament (keratins 8 and 18) and microfilament networks, resulting in a merged structure in which the intermediate filaments are organized around a condensed actin core. Keratin 8, keratin 18 and desmoplakin I/II are the major cytoskeleton-associated targets for microcystin-LR-induced phosphorylation. Hyperphosphorylation of keratin 8 and 18 is accompanied by an increased keratin solubility, which correlates with the observed morphological effects. Phosphopeptide mapping shows that four specific tryptic phosphopeptides are highly phosphorylated predominantly in the soluble pool of keratin 18, whereas keratin 8 shows no indications of such assembly state-specific sites. Phosphopeptide maps of keratins phosphorylated in vivo and in vitro indicate that Ca2+/calmodulin-dependent kinase may be involved in regulating the serine-specific phosphorylation of both keratin 8 and keratin 18, while cAMP-dependent protein kinase does not seem to play a major role in this context. Taken together, our results show that the interactions between keratin intermediate filaments and desmosomes as well as the assembly states of their main constituent proteins, are directly regulated by serine/threonine kinase/phosphatase equilibria.

Actin Cytoskeleton↗
PubMed1996

Lamin and beta-tubulin fragmentation precede chromatin degradation in glutamate-induced neuronal apoptosis.

During ischaemic brain injury, glutamate accumulation with overstimulation of postsynaptic glutamate receptors and intracellular Ca2+ overload lead to neuronal death. We have shown previously that delayed neuronal death in cultures of cerebellar granule cells (CGCs) exposed to glutamate occurs by apoptosis. Here, we report that lamin cleavage and dissolution of the microtubule network precede chromatin fragmentation in glutamate-induced CGC apoptosis. Like other events that characterize excitotoxic cell death, degradation of lamins, beta-tubulin and disruption of microtubule architecture is inhibited by the NMDA-receptor antagonist MK-801. Our findings suggest that cleavage of key cytoskeletal elements is an important step in glutamate-induced neuronal apoptosis.

Animals↗
PubMed1996

Chromatin condensation during apoptosis requires ATP.

The processes leading to morphological changes of the chromatin in cells that undergo apoptosis are presently unclear. We have recently shown that chromatin fragmentation and the nuclear morphological changes typically seen in apoptosis were reproduced in an in vitro system comprised of isolated rat thymocyte nuclei incubated in the presence of a lysate from Fas/APO-1-stimulated JURKAT cells [Chow, Weis, Kass, Holmström, Eriksson and Orrenius (1995) FEBS Lett. 364, 134-138]. Using this in vitro system, we now report that the presence of ATP is necessary for chromatin condensation, its movement to the nuclear periphery and apoptotic body formation. In clear contrast, chromatin cleavage into high-molecular-mass and oligonucleosomal-length DNA fragments induced by lysates derived from Fas/APO-1-activated JURKAT cells did not require the presence of ATP. The induction of these morphological changes by ATP could not be substituted by the analogues, adenosine 5'-[beta, gamma-methylene]triphosphate and adenosine 5'-[alpha, beta-methylene]-triphosphate, AMP, cAMP and UTP. However, adenosine 5'-[gamma-thio]triphosphate, and to a lesser degree GTP and ADP, could partially replace ATP in inducing nuclear apoptotic morphological changes. It is concluded that ATP is essential for the morphological changes occurring in nuclei during apoptosis, but not for DNA fragmentation.

Adenosine Triphosphate↗