NB2a Neuroblastoma Cells Research Articles

Cell type-specific suppression of mechanosensitive genes by audible sound stimulation.

Audible sound is a ubiquitous environmental factor in nature that transmits oscillatory compressional pressure through the substances. To investigate the property of the sound as a mechanical stimulus for cells, an experimental system was set up using 94.0 dB sound which transmits approximately 10 mPa pressure to the cultured cells. Based on research on mechanotransduction and ultrasound effects on cells, gene responses to the audible sound stimulation were analyzed by varying several sound parameters: frequency, wave form, composition, and exposure time. Real-time quantitative PCR analyses revealed a distinct suppressive effect for several mechanosensitive and ultrasound-sensitive genes that were triggered by sounds. The effect was clearly observed in a wave form- and pressure level-specific manner, rather than the frequency, and persisted for several hours. At least two mechanisms are likely to be involved in this sound response: transcriptional control and RNA degradation. ST2 stromal cells and C2C12 myoblasts exhibited a robust response, whereas NIH3T3 cells were partially and NB2a neuroblastoma cells were completely insensitive, suggesting a cell type-specific response to sound. These findings reveal a cell-level systematic response to audible sound and uncover novel relationships between life and sound.

Distinct effect of CacyBP/SIP on the ERK1/2-CREB-BDNF pathway in undifferentiated and differentiated neuroblastoma NB2a cells

CacyBP/SIP, a protein expressed to high extent in the brain, has been shown to act as ERK1/2 phosphatase in vitro and in cultured cells. It has been demonstrated recently that CacyBP/SIP can modulate the activity of some transcription factors in neurons and glioma cells. In the present work we have examined the effect of CacyBP/SIP overexpression and silencing on the phosphorylation/activity of ERK1/2 (pERK1/2) and CREB (pCREB) and on the level of BDNF mRNA in differentiated and undifferentiated neuroblastoma NB2a cells. We have shown that in undifferentiated cells the amount of pERK1/2 decreased upon CacyBP/SIP overexpression. Further studies have shown that the activity of CREB and the level of BDNF mRNA, downstream effectors of the ERK1/2 signaling pathway, also depended on the CacyBP/SIP level and strictly matched the level of pERK1/2. Interestingly, in differentiated NB2a cells, overexpression of CacyBP/SIP appeared to have a distinct effect on the pERK1/2 level from that observed in undifferentiated cells. Subsequent studies have revealed that distinct function of CacyBP/SIP in undifferentiated and differentiated NB2a cells might be due to changes in its posttranslational modifications and protein ligands. Altogether, our studies suggest that CacyBP/SIP is involved in the ERK1/2-CREB-BDNF pathway and that it might regulate this pathway depending on the stage of NB2a cell differentiation.

Influence of S100A6 on CacyBP/SIP Phosphorylation and Elk‐1 Transcriptional Activity in Neuroblastoma NB2a Cells

In this work, we have found that casein kinase II (CKII) phosphorylates the CacyBP/SIP protein under in vitro conditions and have mapped the phosphorylation site to threonine 184. Moreover, we present evidence that S100A6, a CacyBP/SIP interacting protein, inhibits this phosphorylation in the presence of Ca(2+). CacyBP/SIP phosphorylation by CKII was also observed in neuroblastoma NB2a cells. Interestingly, we have found that the effect of DRB, a CKII inhibitor, on CacyBP/SIP phosphorylation state is similar to that of S100A6 overexpression. Phosphorylation at threonine 184 seems to have an effect on CacyBP/SIP phosphatase activity since the T184E phosphorylation mimic mutant overexpressed in NB2a cells has lower phosphatase activity toward p-ERK1/2 when compared to the non-phosphorylable T184A mutant or to the wild-type protein. In conclusion, our data suggest that S100A6 and Ca(2+), through inhibiting CacyBP/SIP phosphorylation on threonine 184, are important regulators of CacyBP/SIP phosphatase activity and of ERK1/2-Elk-1 signaling pathway.

Splicing changes in SMA mouse motoneurons and SMN-depleted neuroblastoma cells: Evidence for involvement of splicing regulatory proteins

Spinal Muscular Atrophy (SMA) is caused by deletions or mutations in the Survival Motor Neuron 1 (SMN1) gene. The second gene copy, SMN2, produces some, but not enough, functional SMN protein. SMN is essential to assemble small nuclear ribonucleoproteins (snRNPs) that form the spliceosome. However, it is not clear whether SMA is caused by defects in this function that could lead to splicing changes in all tissues, or by the impairment of an additional, less well characterized, but motoneuron-specific SMN function. We addressed the first possibility by exon junction microarray analysis of motoneurons (MNs) isolated by laser capture microdissection from a severe SMA mouse model. This revealed changes in multiple U2-dependent splicing events. Moreover, splicing appeared to be more strongly affected in MNs than in other cells. By testing mutiple genes in a model of progressive SMN depletion in NB2a neuroblastoma cells, we obtained evidence that U2-dependent splicing changes occur earlier than U12-dependent ones. As several of these changes affect genes coding for splicing regulators, this may acerbate the splicing response induced by low SMN levels and induce secondary waves of splicing alterations.

The CacyBP/SIP Protein is Sumoylated in Neuroblastoma NB2a Cells

The Calcyclin binding protein and Siah-1 interacting protein (CacyBP/SIP) protein is highly expressed in mammalian brain as well as in neuroblastoma NB2a cells and pheochromocytoma PC12 cells. This protein interacts with several targets such as cytoskeletal proteins or ERK1/2 kinase and seems to be involved in many cellular processes. In this work we examined a post-translational modification of CacyBP/SIP which might have an effect on its function. Since theoretical analysis of the amino acid sequence of CacyBP/SIP indicated several lysine residues which could potentially be sumoylated we checked experimentally whether this protein might be modified by SUMO attachment. We have shown that indeed CacyBP/SIP bound the E2 SUMO ligase, Ubc9, in neuroblastoma NB2a cell extract and was sumoylated in these cells. By fractionation of NB2a cell extract we have found that, contrary to the majority of SUMO-modified proteins, sumoylated CacyBP/SIP is present in the cytoplasmic and not in the nuclear fraction. We have also established that lysine 16 is the residue which undergoes sumoylation in the CacyBP/SIP protein.

Erratum to: Palmitoylcarnitine Affects Localization of Growth Associated Protein GAP-43 in Plasma Membrane Subdomains and its Interaction with Gαo in Neuroblastoma NB-2a Cells

Erratum to: Palmitoylcarnitine Affects Localization of Growth Associated Protein GAP-43 in Plasma Membrane Subdomains and its Interaction with Gαo in Neuroblastoma NB-2a Cells

Quantitative Analysis of How RET Receptor Activation is Coupled to ERK and Akt Signaling

The receptor tyrosine kinase RET is activated by growth factors of the GDNF family in conjunction with one of four glycosylphosphatidylinositol‐linked co‐receptors, GFRα1–4. Like other growth factor receptors, our quantitative knowledge of RET function, and particularly how RET activation is quantitatively and temporally coupled to downstream signaling events, remains very rudimentary. We have addressed this knowledge gap by using ELISA assays to precisely measure the phosphorylation levels of RET receptor, ERK1/2 and Akt that are observed upon stimulating mouse NB41A3 neuroblastoma cells expressing RET and GFRα3 with various concentrations of the growth factor artemin for different periods of time. We also use small molecule inhibitors of different steps in these signaling pathways to gain additional information by orthogonally perturbing the system, and to probe crosstalk between the ERK and Akt pathways. We extracted key features of the RET, ERK1/2 and Akt activation processes, such as initial activation rate, maximum and steady‐state signal amplitude, and signal duration, and tested the resulting relationships against models reflecting different connection topologies between these events. The results shed important new light on how activation of a growth factor receptor is quantitatively and temporally coupled to key events in divergent downstream signaling pathways. (GM087469)

Palmitoylcarnitine Affects Localization of Growth Associated Protein GAP-43 in Plasma Membrane Subdomains and its Interaction with Gαo in Neuroblastoma NB-2a Cells

Palmitoylcarnitine was observed previously to promote differentiation of neuroblastoma NB-2a cells, and to affect protein kinase C (PKC). Palmitoylcarnitine was also observed to increase palmitoylation of several proteins, including a PKC substrate, whose expression augments during differentiation of neural cells-a growth associated protein GAP-43, known to bind phosphatidylinositol 4,5-bisphosphate [PI(4,5)P(2)]. Since palmitoylated proteins are preferentially localized in sphingolipid- and cholesterol-rich microdomains of plasma membrane, the present study has been focused on a possible effect of palmitoylcarnitine on GAP-43 localization in these microdomains. Palmitoylcarnitine treatment resulted in GAP-43 appearance in floating fractions (rafts) in sucrose gradient and increased co-localization with cholesterol and with PI(4,5)P(2), although co-localization of both lipids decreased. GAP-43 disappeared from raft fraction upon treatment with 2-bromopalmitate (an inhibitor of palmitoylating enzymes) and after treatment with etomoxir (carnitine palmitoyltransferase I inhibitor). Raft localization of GAP-43 was completely abolished by treatment with methyl-β-cyclodextrin, a cholesterol binding agent, while there was no change upon sequestration of PI(4,5)P(2) with neomycin. GAP-43 co-precipitated with a monomeric form of Gα(o), a phenomenon diminished after palmitoylcarnitine treatment and paralleled by a decrease of Gα(o) in the raft fraction. These observations point to palmitoylation of GAP-43 as a mechanism leading to an increased localization of this protein in microdomains of plasma membrane rich in cholesterol, in majority different, however, from microdomains in which PI(4,5)P(2) is present. This localization correlates with decreased interaction with Gα(o) and suppression of its activity-an important step regulating neural cell differentiation.

CacyBP/SIP phosphatase activity in neuroblastoma NB2a and colon cancer HCT116 cells

Recently, we have reported that CacyBP/SIP could be a novel phosphatase for ERK1/2 kinase. In this work, we analyzed the CacyBP/SIP phosphatase activity toward ERK1/2 in 2 cell lines of different origin. We showed that overexpression of CacyBP/SIP in NB2a cells resulted in a lower level of phosphorylated ERK1/2 (P-ERK1/2) in the nuclear fraction while such overexpression in HCT116 cells had no effect on the level of P-ERK1/2. Moreover, we found that overexpression of CacyBP/SIP resulted in higher phosphatase activity in the nuclear fraction obtained from NB2a cells when compared with HCT116 cells. Using 2-D electrophoresis we showed that the pattern of spots representing CacyBP/SIP differed in these 2 cell lines and was probably due to a different phosphorylation state of this protein. We also established that after overexpression of CacyBP/SIP in NB2a cells, the amount of nuclear β-catenin was low, while it remained high in HCT116 cells. Since NB2a cells have differentiation potential and HCT116 cells do not, our data suggest that different activity of CacyBP/SIP in these 2 cell lines might affect the ERK1/2 pathway in the differentiation or proliferation processes.

CacyBP/SIP interacts with tubulin in neuroblastoma NB2a cells and induces formation of globular tubulin assemblies

CacyBP/SIP, originally identified as a S100A6 (calcyclin) target, was later shown to interact with some other members of the S100 family as well as with Siah-1 and Skp1 proteins. Recently, it has been shown that CacyBP/SIP is up-regulated during differentiation of cardiomyocytes. In this work we show that the level of CacyBP/SIP is higher in differentiated neuroblastoma NB2a cells than in undifferentiated ones and that in cells overexpressing CacyBP/SIP the level of GAP-43, a marker of differentiation, was increased. Since the process of differentiation is accompanied by an extensive rearrangement of microtubules, we examined whether CacyBP/SIP interacted with tubulin. By applying cross-linking experiments we found that these two proteins bind directly. The dissociation constant of the tubulin–CacyBP/SIP complex determined by the surface plasmon resonance technique is 1.57 × 10 − 7 M which suggests that the interaction is tight. The interaction and co-localization of CacyBP/SIP and tubulin was also demonstrated by co-immunoprecipitation, affinity chromatography and immunofluorescence methods. Light scattering measurements and electron microscopy studies revealed that CacyBP/SIP, but not its homologue, Sgt1, increased tubulin oligomerization. Altogether, our results suggest that CacyBP/SIP, via its interaction with tubulin, might contribute to the differentiation of neuroblastoma NB2a cells.

Palmitoylcarnitine regulates estrification of lipids and promotes palmitoylation of GAP-43

Palmitoylcarnitine was previously shown to promote differentiation of neuroblastoma NB-2a cells. It was also observed to increase palmitoylation of several proteins and to diminish incorporation of palmitic acid to phospholipids, as well as to affect growth associated protein GAP-43 by decreasing its phosphorylation and interaction with protein kinase C. The present study was focused on influence of palmitoylcarnitine on palmitoylation of GAP-43 and lipid metabolism. Althought palmitoylcarnitine did not significantly affect the total phospholipids and fatty acid content, it increased incorporation of palmitate moiety to triacylglicerides and cholesterol esters, with a decrease of free cholesterol content. The presence of palmitoylcarnitine significantly increased the amount of covalently bound palmitate to GAP-43, which can regulate the signal transduction pathways.

Characterization of apoptotic pathway associated with caspase-independent excision of DNA loop domains

Excision of chromatin loop domains and internucleosomal DNA fragmentation are widely considered as consecutive stages of chromatin disassembly during apoptosis. We report here on apoptosis induced by staurosporine in NB-2a neuroblastoma cells, which was accompanied by excision of chromatin loop domains, but proceeded without internucleosomal DNA cleavage. In contrast to apoptosis associated with internucleosomal DNA fragmentation, the apoptotic pathway associated with excision of chromatin loop domains was largely caspase independent. We identify here MAPK family member, p38/JNK, mitochondria, and topoisomerase II as the components of this caspase-independent apoptotic pathway. While caspase-independent excision of chromatin loop domains was a predominant mechanism of DNA disintegration in staurosporine-treated neuroblastoma, both caspase-dependent internucleosomal DNA fragmentation and caspase-independent excision of chromatin loop domains accompanied staurosporine-induced apoptosis of promyelocytic leukemia cells. Our results suggest that caspase-independent excision of chromatin loop domains represents a separate cell death pathway, which operates either in parallel or independently from caspase-dependent internucleosomal DNA fragmentation.

Food Safety: Adding Up to No Good?

The safety of food additives is usually examined by varying the dose of a single additive administered to animal models or cell cultures. However, British researchers at the University of Liverpool and the University of Ulster report in the March 2006 issue of Toxicological Sciences that combinations of additives can produce neurotoxic effects at dosages that are safe when each additive is tested alone. The authors examined four common food additives: Quinoline Yellow (FDC Brilliant Blue was banned from foods in most European countries but has since been reapproved. Coauthor Karen Lau, a doctoral student, says these additives were tested because they are commonly used in foods targeted for consumption by children. Neurotoxicity was measured by the relative growth of neurites from mouse NB2a neuroblastoma cells after exposure to the additives. Two combinations of additives stunted neurite growth: Quinoline Yellow paired with aspartame, and Brilliant Blue paired with L-glutamatic acid. Other pairings showed no effect. Lau hypothesizes that ingestion of the well-established neurotoxicants aspartic acid and L-glutamic acid as additives could lead to a high enough body burden to kill neurons by a mechanism called excitotoxicity. Lau says young children may be especially at risk for the type of toxicity observed in the nerve-cell cultures, because effects were seen at concentrations of additives she says are theoretically achievable in plasma by eating foods and drinks typically consumed by children—for example, a snack of corn chips, which may contain MSG, and a fruit juice drink, which may contain aspartame. Scientists at the U.K. Food Standards Agency (FSA) question whether these results are relevant to the human consumption of these additives. “[The Lau study assumes] that both MSG and aspartame are absorbed one hundred percent in the gut, but [other studies] show that this does not seem to be the case,” says FSA senior press officer Shaun Whelan. “It is . . . extremely unlikely that the plasma levels predicted by the authors of this study accurately reflect the actual situations in vivo.” Whelan says glutamic acid and aspartic acid occur naturally in many foods, and there is no evidence that they are treated differently in the body when they are ingested as food additives. Lau counters, however, that consumption of glutamic acid in its free form or as MSG has a more dramatic effect on plasma levels than that of glutamatic acid in protein, and can lead to high concentrations in the body. More research is needed to clarify these effects. Other studies have suggested that non-nutritive food additives are associated with behavioral disorders such as attention deficit/hyperactivity disorder. These effects are controversial, but Lau’s team believes their results warrant further investigation of such possibilities. Whelan says the FSA is currently funding research on the effects of ingested chemical mixtures, including color additives, on the behavior of young children.

Palmitoylcarnitine modulates interaction between protein kinase C βII and its receptor RACK1

Palmitoylcarnitine, known to promote differentiation of neuroblastoma NB-2a cells as well as to inhibit protein kinase C (PKC) activity and to decrease phorbol ester binding, was shown previously to diminish the amount of complex formed between PKCdelta and its substrate GAP-43. In the present work we studied the effect of palmitoylcarnitine on the interaction between PKCbetaII and its receptor RACK1. Palmitoylcarnitine was found to decrease autophosphorylation of PKCbetaII on serine in a concentration-dependent manner and to decrease the amount of PKCbetaII/RACK1 complex. The effect of palmitoylcarnitine on cellular localization was found to be dependent on the presence of ATP; palmitoylcarnitine lowered the amount of PKCbetaII in cytosol and decreased the amount of PKCbetaII-RACK1 complex in membrane in the absence of ATP. Palmitoylcarnitine also reversed the effect of phorbol ester on the increase in the amount of PKCbetaII in membrane. Palmitoylcarnitine binds to PKCbetaII through hydrophobic interactions, although acylation of PKCbetaII by the palmitate moiety has been excluded. The presence of palmitoylcarnitine did not have any additive effect on the diminution of PKCbetaII-RACK1 complex formation in the presence of a RACK1-binding peptide from within the C2 region of PKCbetaII. These results rather exclude a possibility of interaction of palmitoylcarnitine with the C2 domain and suggest a possible interaction with the V5 domain and a conformational change affecting the C1 region.

Synergistic Interactions between Commonly Used Food Additives in a Developmental Neurotoxicity Test

Exposure to non-nutritional food additives during the critical development window has been implicated in the induction and severity of behavioral disorders such as attention deficit hyperactivity disorder (ADHD). Although the use of single food additives at their regulated concentrations is believed to be relatively safe in terms of neuronal development, their combined effects remain unclear. We therefore examined the neurotoxic effects of four common food additives in combinations of two (Brilliant Blue and L-glutamic acid, Quinoline Yellow and aspartame) to assess potential interactions. Mouse NB2a neuroblastoma cells were induced to differentiate and grow neurites in the presence of additives. After 24 h, cells were fixed and stained and neurite length measured by light microscopy with computerized image analysis. Neurotoxicity was measured as an inhibition of neurite outgrowth. Two independent models were used to analyze combination effects: effect additivity and dose additivity. Significant synergy was observed between combinations of Brilliant Blue with L-glutamic acid, and Quinoline Yellow with aspartame, in both models. Involvement of N-methyl-D-aspartate (NMDA) receptors in food additive-induced neurite inhibition was assessed with a NMDA antagonist, CNS-1102. L-glutamic acid- and aspartame-induced neurotoxicity was reduced in the presence of CNS-1102; however, the antagonist did not prevent food color-induced neurotoxicity. Theoretical exposure to additives was calculated based on analysis of content in foodstuff, and estimated percentage absorption from the gut. Inhibition of neurite outgrowth was found at concentrations of additives theoretically achievable in plasma by ingestion of a typical snack and drink. In addition, Trypan Blue dye exclusion was used to evaluate the cellular toxicity of food additives on cell viability of NB2a cells; both combinations had a straightforward additive effect on cytotoxicity. These data have implications for the cellular effects of common chemical entities ingested individually and in combination.

Methylmercury inhibits type II 5′-deiodinase activity in NB41A3 neuroblastoma cells

Methylmercury (MeHg) is a well-known neurotoxicant and prenatal exposure to MeHg results in severe brain damage. Since MeHg has a high affinity for thiol groups, we sought to determine whether MeHg inhibited type II iodothyronine deiodinase (D2) activity, by which prohormone thyroxine (T4) is converted to active thyroid hormone, 3,5,3′-triiodothyronine (T3) in the brain, using NB41A3 mouse neuroblastoma cells. In MeHg-treated cells, D2 activity was inhibited in a dose- and time-dependent manner; relatively low concentrations of MeHg (30 nM) inhibited D2. Kinetic analysis using a double reciplocal plot of D2 activity revealed competitive inhibition by MeHg. DTT protected D2 from MeHg when cells were incubated with both MeHg and DTT or when MeHg was added to the assay buffer containing DTT and cell sonicates from untreated cells. Removal of MeHg from culture medium did not recover D2 activity. These results demonstrate that MeHg inhibited D2 activity in NB41A3 cells and the selenocysteine in the catalytic subunit of D2 may be involved in the inhibitory action of MeHg. Further our results suggest that T3 deficiency due to D2 inhibition in the brain may be involved in the neurotoxicity of MeHg.

VSV replication in neurons is inhibited by type I IFN at multiple stages of infection

Vesicular stomatitis virus (VSV) is a rhabdovirus which causes acute encephalitis in mice after intranasal infection. Because type I interferon (IFN) has been shown to be a potent inhibitor of VSV, we investigated the role of type I IFN in viral replication in neurons in culture. Pre-treatment of NB41A3 neuroblastoma cells or primary neuron cultures with IFN-β or IFN-α strongly inhibits virus replication, with 1000-fold inhibition of infectious virus release occurring at 7 h post-infection, and maximum inhibition of 14,000-fold occurring at 14 h. Type I IFN inhibited both viral protein and RNA synthesis, but not enough to account for the inhibition of infectious virus yield. The influenza virus protein NS1 binds dsRNA and antagonizes induction of PKR activity, an IFN-inducible antiviral protein which phosphorylates and inactivates the elongation factor eIF-2α, resulting in cessation of translation. In NS1-expressing neuroblastoma cells, VSV replication was inhibited by IFN-β as well as in control NB41A3 cells, and eIF-2α phosphorylation was blocked, suggesting that PKR activity was not involved in inhibition of viral protein synthesis. Similarly, inhibition of VSV by IFN-β was not affected by addition of inhibitors of nitric oxide synthase, indicating that IFN-β activity is not mediated by nitric oxide or superoxide. This contrasts with the essential role of NOS-1 in inhibition of VSV replication when neurons are treated with IFN-γ. Analysis of cell culture supernatants revealed suppression of release of VSV particles from both NB41A3 cells and primary neurons treated with IFN. The inhibition of virion release closely matched the overall suppression of infectious VSV particle release, suggesting that type I IFN plays a role in inhibition of VSV assembly.

Peptide-modified alginate surfaces as a growth permissive substrate for neurite outgrowth.

Different strategies are being investigated for treatment of spinal cord injuries, one of the most promising being application of neurotrophic factors, which have been shown to prevent neuronal death and stimulate regeneration of injured axons. Ex vivo gene therapy has emerged as the leading delivery method at the site of the injury, and we have shown previously that encapsulating genetically engineered fibroblasts in an immunoprotective alginate capsule can permit implantation of the factor-secreting cells without need for immunosuppression. This strategy could be greatly enhanced by providing the sprouting neurons with a permissive substrate upon which to attach and grow. We report here studies on the modification of an alginate gel surface by either coating it with laminin or by covalent attachment of YIGSR peptide. Using NB2a neuroblastoma cells, we found that native alginate elicited minimal cell attachment ( approximately 1.5%); however, YIGSR-alginate conjugate elicited a fivefold increase in numbers of cells attached using peptide ratios of 0.5 and 1 mg/g alginate, ranging from 9.5% of the cells at the lower ratio, to about 44% at the higher. Only a further 19% increase was obtained at an increased peptide density of 2 mg/g alginate ( approximately 63% over control). Laminin-coated gels showed approximately 60% cell attachment. However, laminin coating did not stimulate differentiation and neurite growth, whereas both numbers and lengths of outgrowths increased with increasing peptide density on peptide-modified alginate. We demonstrate here the ability of the peptide-modified alginate gels to allow adhesion of NB2a neuroblastoma cells and to promote neurite outgrowth from these cells when attached to the peptide-modified alginate surface. Also, we show that the adhesion of NB2a neuroblastoma cells and neurite outgrowth from the attached cells is a function of the peptide density on the gel surface.

PKR is not required for interferon-gamma inhibition of VSV replication in neurons.

In this report, the contribution of PKR to the IFN-gamma mediated inhibition of VSV replication in neurons was examined. IFN-gamma treatment of NB41A3 murine neuroblastoma cells resulted in the reduced expression of VSV protein during infection. PKR was found to be modestly upregulated in NB41A3 cells following IFN-gamma treatment. The phosphorylation state of PKR and its downstream target, eIF2alpha, were unaffected by either IFN-gamma or VSV infection. Inhibition of PKR through the use of 2-aminopurine or the expression of the Influenza A NS1 gene had no effect on the ability of IFN-gamma to inhibit the replication of VSV in vitro. These data indicate that endogenously expressed PKR is not required for the IFN-gamma mediated inhibition of VSV replication in NB41A3 neuroblastoma cells.

Palmitoylcarnitine modulates palmitoylation of proteins: implication for differentiation of neural cells.

[3H]Palmitic acid accumulates in neuroblastoma NB-2a cells, being incorporated in lipids (90%) and proteins (10%) fractions. Addition of palmitoylcarnitine, known to modulate activity of protein kinase C and to promote differentiation of neurons, was observed to decrease incorporation of palmitic acid to sphingomyelin, phosphatidylserine, and phosphatidylcholine, with a parallel increase of palmitic acid bound to proteins through a thioester bond (palmitoylation). In the presence of palmitoylcarnitine, one of the palmitoylated proteins expressed at growing neural cones, GAP-43, was observed to co-localize with caveolin-1, what was correlated with the beginning of differentiation. A new function of palmitoylcarnitine in controlling palmitoylation of proteins and their targeting to cholesterol-rich domains has been proposed.