Induction of differentiation and <i>c‐jun</i> expression in human leukemic cells by okadaic acid, an inhibitor of protein phosphatases

Adunyah, Samuel E.; Unlap, Tino; Franklin, Christopher C.; Kraft, Andrew S.

doi:10.1002/jcp.1041510223

Cited by 23 publications

(15 citation statements)

References 43 publications

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“…TPA down-regulates c-myc transcripts in HL-60 cells (47) and induces expression of the c-jun gene (49,54,61). Similar findings have been obtained with other inducers of monocytic differentiation (49), including okadaic acid, an inhibitor of phosphoserine/threonine protein phosphatases 1 and 2A (1,25). Activation of Jun/AP-1 contributes to induction of c-jun transcription (2) and monocytic differentiation (54).…”

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confidence: 86%

Functional Role for Protein Kinase Cβ as a Regulator of Stress-Activated Protein Kinase Activation and Monocytic Differentiation of Myeloid Leukemia Cells

Kaneki

Kharbanda

Pandey

et al. 1999

Molecular and Cellular Biology

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The human U-937 and HL-60 myeloid leukemia cell lines proliferate autonomously in the absence of exogenous hematopoietic growth factors (6, 52). These cells, however, have retained the capacity to respond to inducers of differentiation with growth arrest and the appearance of a mature phenotype. In this context, treatment of U-937 and HL-60 cells with agents that activate protein kinase C (PKC), including 12-O-tetradecanoylphorbol-13-acetate (TPA) and phorbol-12,13-dibutyrate (PDBu), induces differentiation along the monocytic lineage. Bryostatin 1, a macrocyclic lactone, also activates PKC and induces monocytic differentiation of myeloid leukemia cells (51). While these findings have indicated that factor-independent growth of myeloid leukemia cells is reversible by activation of PKC-mediated signaling, little is known about the downstream effectors responsible for induction of the differentiated monocytic phenotype.PKC is a family of at least 12 serine/threonine protein kinase isoforms which are involved in diverse cellular responses (24, 43). The ␣, ␤, ␥, ␦, ε, , , , and forms of PKC are responsive to phorbol esters. The available evidence suggests that PKC␤ is involved in TPA-induced differentiation of myeloid leukemia cells. Accordingly, TPA-resistant HL-60 cell variants are deficient in PKC␤ expression (37,42,56,57). Down-regulation of PKC␤ expression (19) and functional defects in PKC␤ (31) have also been found for TPA-resistant U-937 cell variants. In addition, defective translocation of PKC␤ from the cytosol to the cell membrane has been shown for TPA-resistant variants of both U-937 and HL-60 cells (19,64). Importantly, increased expression of PKC␤ resulting from treatment with retinoic acid (64) or from transfection of the PKC␤ gene (56) restores TPA inducibility of growth arrest and a differentiated monocytic phenotype. PKC␤ is expressed as two isoforms, ␤I and ␤II, as a result of an alternative splicing mechanism that produces a PKC␤I protein which is truncated by 50 amino acids at the carboxy terminus (32); the longer PKC␤II isoform is expressed in U-937 and HL-60 cells (22,56).Treatment of myeloid leukemia cells with TPA is associated with changes in the expression of certain early-and late-response genes. TPA down-regulates c-myc transcripts in HL-60 cells (47) and induces expression of the c-jun gene (49,54,61). Similar findings have been obtained with other inducers of monocytic differentiation (49), including okadaic acid, an inhibitor of phosphoserine/threonine protein phosphatases 1 and 2A (1,25). Activation of Jun/AP-1 contributes to induction of c-jun transcription (2) and monocytic differentiation (54). The early growth response 1 (EGR-1) gene is also activated during TPA-and okadaic acid-induced monocytic differentiation (27,29) and is necessary for the appearance of the monocytic phenotype (41). Thus, the induction of early response genes and thereby upstream signals involved in their transcriptional activation may be directly linked to the reversal of the leukemia phenotype.Members of ...

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confidence: 86%

Functional Role for Protein Kinase Cβ as a Regulator of Stress-Activated Protein Kinase Activation and Monocytic Differentiation of Myeloid Leukemia Cells

Kaneki

Kharbanda

Pandey

et al. 1999

Molecular and Cellular Biology

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“…Transient transfections were performed using the DEAE-dextran procedure as described previously (32). Stable transfections were performed by electroporation (26).…”

Section: Methodsmentioning

confidence: 99%

Conditional Expression of the Mitogen-activated Protein Kinase (MAPK) Phosphatase MKP-1 Preferentially Inhibits p38 MAPK and Stress-activated Protein Kinase in U937 Cells

Franklin

Kraft

1997

Journal of Biological Chemistry

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409

361

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Phorbol ester tumor promoters, such as phorbol 12-myristate 13-acetate (PMA), are potent activators of extracellular signal-regulated kinase 2 (ERK2), stress-activated protein kinase (SAPK), and p38 mitogenactivated protein kinase (MAPK) in U937 human leukemic cells. These kinases are regulated by the reversible dual phosphorylation of conserved threonine and tyrosine residues. The dual specificity protein phosphatase MAPK phosphatase-1 (MKP-1) has been shown to dephosphorylate and inactivate ERK2, SAPK, and p38 MAPK in transient transfection studies. Here we demonstrate that PMA treatment induces MKP-1 protein expression in U937 cells, which is detectable within 30 min with maximal levels attained after 4 h. This time course coincides with the rapid inactivation of PMAinduced SAPK activity, but not ERK2 phosphorylation, which remains elevated for up to 6 h. To examine directly the role of MKP-1 in the regulation of these protein kinases in vivo, we established a U937 cell line that conditionally expresses MKP-1 from the human metallothionein IIa promoter. Conditional expression of MKP-1 inhibited PMA-induced ERK2, SAPK, and p38 MAPK activity. By titrating the levels of MKP-1 expression from the human metallothionein IIa promoter, however, it was found that p38 MAPK and SAPK were much more sensitive to inhibition by MKP-1 than ERK2. This differential substrate specificity of MKP-1 can be functionally extended to nuclear transcriptional events in that PMAinduced c-Jun transcriptional activity was more sensitive to inhibition by MKP-1 than either Elk-1 or c-Myc. Conditional expression of MKP-1 also abolished the induction of endogenous MKP-1 protein expression in response to PMA treatment. This negative feedback regulatory mechanism is likely due to MKP-1-mediated inhibition of ERK2, as studies utilizing the MEK1/2 inhibitor PD98059 suggest that ERK2 activation is required for PMA-induced MKP-1 expression. These findings suggest that ERK2-mediated induction of MKP-1 may play an important role in preferentially attenuating signaling through the p38 MAPK and SAPK signal transduction pathways. Mitogen-activated protein (MAP)1 kinases play a key role in transducing various extracellular signals to the nucleus (1). The MAP kinases (MAPKs) consist of three major subgroups that include the ERK, SAPK/JNK, and p38 MAPK families (2). The ERKs, SAPKs, and p38 MAPK are activated by the reversible dual threonine and tyrosine phosphorylation of a conserved TEY, TPY, or TGY motif, respectively (1-5). Although distinct and selective activators of the MAPKs have been cloned and characterized (6 -9), less in known about the negative regulation of these kinases. The reversible nature of MAPK phosphorylation suggests that protein phosphatases play an important role in regulating MAPK activity. An expanding subfamily of dual specificity protein tyrosine phosphatases has been identified which is capable of dephosphorylating and inactivating various members of the MAPK family. This class of phosphatases is characterized by MKP-1 (also k...

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“…Cells were cultured in flasks at 37°C for propagation and in 12 well plates with rattail collagen coated glass cover slips for differentiation and infection. U937 cell differentiation was achieved as described previously (Adunyah et al, 1992). Briefly, cells were counted using a hemocytometer and 1×106 cells were incubated with complete RPMI medium containing 4nM PMA.…”

Section: Mammalian Cell Culturementioning

confidence: 99%

A Short Interfering Rna Molecular Beacon for the Attenuation of Mycobacterial Infection

George¹

2014

American Journal of Biochemistry and Biotechnology

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The ability of the pathogen Mycobacterium Tuberculosis (MTB) to invade and survive within macrophages of granulomas is attributed to the product of the Mammalian Cell Entry (MCE) operon whose gene, mce4A, encodes a cholesterol transporter that transports host lipids into the bacterium that allows the bacterium to survive during chronic infection. Here, we proposed and tested the hypothesis that a mce4A siRNA molecular beacon can be used to attenuate mycobacterial infection in macrophages. Mce4A gene was cloned and expressed in E. coli (E. coli-4A) and differentiated U937 cells were transduced with piLenti-siRNA-GFP phage expressing the mce4A siRNA for 24 h. This was followed by infection with either E. coli-4A or M. smegmatis for 3 h followed by incubation for 0, 3, 6, 24 and 48 h. The cells were lysed and the lysates were plated on LB agar plates containing ampicillin (100 µg mL −1 ) or on 7H11 media and incubated at 37°C overnight. Our results showed that the siRNA treatment attenuated E.coli-4A infection in macrophages at 3, 6, 24 and 48 h by 0, 77, 59.6 and 99.7%, respectively. Our results also showed that the siRNA treatment attenuated M. smegmatis infection in macrophages at 3, 6, 24 and 48 h. by 94.8, 70.3, 98.9 and 93.4%, respectively. In conclusion, a mce4A siRNA molecular beacon was successfully delivered and stably expressed in macrophages which attenuated E. coli expressing mce4A (E. coli-4A) and M. smegmatis infection in macrophages.

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Induction of differentiation and c‐jun expression in human leukemic cells by okadaic acid, an inhibitor of protein phosphatases

Cited by 23 publications

References 43 publications

Functional Role for Protein Kinase Cβ as a Regulator of Stress-Activated Protein Kinase Activation and Monocytic Differentiation of Myeloid Leukemia Cells

Functional Role for Protein Kinase Cβ as a Regulator of Stress-Activated Protein Kinase Activation and Monocytic Differentiation of Myeloid Leukemia Cells

Conditional Expression of the Mitogen-activated Protein Kinase (MAPK) Phosphatase MKP-1 Preferentially Inhibits p38 MAPK and Stress-activated Protein Kinase in U937 Cells

A Short Interfering Rna Molecular Beacon for the Attenuation of Mycobacterial Infection

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