Selective translocation of βII-protein kinase C to the nucleus of human promyelocytic (HL60) leukemia cells

Barbara A. Hocevar, Alan P Fields

Research output: Contribution to journalArticle

220 Citations (Scopus)

Abstract

The promyelocytic leukemia (HL60) cell line differentiates into monocyte-like cells after treatment with phorbol dibutyrate (PBt2). In contrast, bryostatin 1 (bryo), a structurally distinct protein kinase C (PKC) activator, does not induce differentiation and blocks the cytostatic effect of PBt2. The divergent responses to these agents correlate with activation of a PKC-like activity at the nucleus in response to bryo but not PBt2 (Fields, A. P., Pettit, G. R., and May, W. S. (1988) J. Biol. Chem. 263, 8253-8260). In the present study, this nuclear PKC-like activity (termed PKCn) was isolated from HL60 cells and shown to phosphorylate its known nuclear substrate, lamin B. PKCn-mediated phosphorylation of nuclear envelope-associated lamin B in vitro is calcium-dependent and is stimulated by bryo and 1,2-dioctanoylglycerol (DiC8), but not PBt2. In contrast, PKCn-mediated phosphorylation of histone IIIS is stimulated equally by all three activators. PKCn mediates calcium- and phosphatidylserine-dependent phosphorylation of both histone IIIS and partially purified lamin B. PKCn activity can be inhibited by an anti-PKC monoclonal antibody which specifically inhibits PKC. Isotype-specific PKC antibodies identify PKCn as βII-PKC. Immunoblot analysis indicates that HL60 cells express both α- and βII-PKC but no βI- or γ-PKC. Treatment of intact cells with bryo for 30 min leads to complete translocation of both α- and βII-PKC from the cytosol to the membrane fractions. Approximately 8-10% of the total βII-PKC (and 2 treatment leads to complete translocation of α-PKC, but only partial translocation of βII-PKC to the plasma membrane fraction. Neither PKC isotype is found associated with the nuclear membrane of PBt2-treated cells. These data demonstrate that α- and βII-PKC differ with respect to activator responsiveness, intracellular distribution, and substrate specificity and indicate that their selective activation at distinct intracellular sites, including the nucleus, can have a dramatic effect on resulting cellular responses.

Original languageEnglish (US)
Pages (from-to)28-33
Number of pages6
JournalJournal of Biological Chemistry
Volume266
Issue number1
StatePublished - Jan 5 1991
Externally publishedYes

Fingerprint

HL-60 Cells
Protein Kinase C
Leukemia
Lamin Type B
Phosphorylation
Nuclear Envelope
Cells
Histones
Chemical activation
Calcium
Membranes
Phosphatidylserines
Cytostatic Agents
Substrates
Cell membranes
Nuclear Proteins
Substrate Specificity
Cytosol
Monocytes

ASJC Scopus subject areas

  • Biochemistry

Cite this

Selective translocation of βII-protein kinase C to the nucleus of human promyelocytic (HL60) leukemia cells. / Hocevar, Barbara A.; Fields, Alan P.

In: Journal of Biological Chemistry, Vol. 266, No. 1, 05.01.1991, p. 28-33.

Research output: Contribution to journalArticle

@article{db5654ada2744fe7ab36aa64c1129360,
title = "Selective translocation of βII-protein kinase C to the nucleus of human promyelocytic (HL60) leukemia cells",
abstract = "The promyelocytic leukemia (HL60) cell line differentiates into monocyte-like cells after treatment with phorbol dibutyrate (PBt2). In contrast, bryostatin 1 (bryo), a structurally distinct protein kinase C (PKC) activator, does not induce differentiation and blocks the cytostatic effect of PBt2. The divergent responses to these agents correlate with activation of a PKC-like activity at the nucleus in response to bryo but not PBt2 (Fields, A. P., Pettit, G. R., and May, W. S. (1988) J. Biol. Chem. 263, 8253-8260). In the present study, this nuclear PKC-like activity (termed PKCn) was isolated from HL60 cells and shown to phosphorylate its known nuclear substrate, lamin B. PKCn-mediated phosphorylation of nuclear envelope-associated lamin B in vitro is calcium-dependent and is stimulated by bryo and 1,2-dioctanoylglycerol (DiC8), but not PBt2. In contrast, PKCn-mediated phosphorylation of histone IIIS is stimulated equally by all three activators. PKCn mediates calcium- and phosphatidylserine-dependent phosphorylation of both histone IIIS and partially purified lamin B. PKCn activity can be inhibited by an anti-PKC monoclonal antibody which specifically inhibits PKC. Isotype-specific PKC antibodies identify PKCn as βII-PKC. Immunoblot analysis indicates that HL60 cells express both α- and βII-PKC but no βI- or γ-PKC. Treatment of intact cells with bryo for 30 min leads to complete translocation of both α- and βII-PKC from the cytosol to the membrane fractions. Approximately 8-10{\%} of the total βII-PKC (and 2 treatment leads to complete translocation of α-PKC, but only partial translocation of βII-PKC to the plasma membrane fraction. Neither PKC isotype is found associated with the nuclear membrane of PBt2-treated cells. These data demonstrate that α- and βII-PKC differ with respect to activator responsiveness, intracellular distribution, and substrate specificity and indicate that their selective activation at distinct intracellular sites, including the nucleus, can have a dramatic effect on resulting cellular responses.",
author = "Hocevar, {Barbara A.} and Fields, {Alan P}",
year = "1991",
month = "1",
day = "5",
language = "English (US)",
volume = "266",
pages = "28--33",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "1",

}

TY - JOUR

T1 - Selective translocation of βII-protein kinase C to the nucleus of human promyelocytic (HL60) leukemia cells

AU - Hocevar, Barbara A.

AU - Fields, Alan P

PY - 1991/1/5

Y1 - 1991/1/5

N2 - The promyelocytic leukemia (HL60) cell line differentiates into monocyte-like cells after treatment with phorbol dibutyrate (PBt2). In contrast, bryostatin 1 (bryo), a structurally distinct protein kinase C (PKC) activator, does not induce differentiation and blocks the cytostatic effect of PBt2. The divergent responses to these agents correlate with activation of a PKC-like activity at the nucleus in response to bryo but not PBt2 (Fields, A. P., Pettit, G. R., and May, W. S. (1988) J. Biol. Chem. 263, 8253-8260). In the present study, this nuclear PKC-like activity (termed PKCn) was isolated from HL60 cells and shown to phosphorylate its known nuclear substrate, lamin B. PKCn-mediated phosphorylation of nuclear envelope-associated lamin B in vitro is calcium-dependent and is stimulated by bryo and 1,2-dioctanoylglycerol (DiC8), but not PBt2. In contrast, PKCn-mediated phosphorylation of histone IIIS is stimulated equally by all three activators. PKCn mediates calcium- and phosphatidylserine-dependent phosphorylation of both histone IIIS and partially purified lamin B. PKCn activity can be inhibited by an anti-PKC monoclonal antibody which specifically inhibits PKC. Isotype-specific PKC antibodies identify PKCn as βII-PKC. Immunoblot analysis indicates that HL60 cells express both α- and βII-PKC but no βI- or γ-PKC. Treatment of intact cells with bryo for 30 min leads to complete translocation of both α- and βII-PKC from the cytosol to the membrane fractions. Approximately 8-10% of the total βII-PKC (and 2 treatment leads to complete translocation of α-PKC, but only partial translocation of βII-PKC to the plasma membrane fraction. Neither PKC isotype is found associated with the nuclear membrane of PBt2-treated cells. These data demonstrate that α- and βII-PKC differ with respect to activator responsiveness, intracellular distribution, and substrate specificity and indicate that their selective activation at distinct intracellular sites, including the nucleus, can have a dramatic effect on resulting cellular responses.

AB - The promyelocytic leukemia (HL60) cell line differentiates into monocyte-like cells after treatment with phorbol dibutyrate (PBt2). In contrast, bryostatin 1 (bryo), a structurally distinct protein kinase C (PKC) activator, does not induce differentiation and blocks the cytostatic effect of PBt2. The divergent responses to these agents correlate with activation of a PKC-like activity at the nucleus in response to bryo but not PBt2 (Fields, A. P., Pettit, G. R., and May, W. S. (1988) J. Biol. Chem. 263, 8253-8260). In the present study, this nuclear PKC-like activity (termed PKCn) was isolated from HL60 cells and shown to phosphorylate its known nuclear substrate, lamin B. PKCn-mediated phosphorylation of nuclear envelope-associated lamin B in vitro is calcium-dependent and is stimulated by bryo and 1,2-dioctanoylglycerol (DiC8), but not PBt2. In contrast, PKCn-mediated phosphorylation of histone IIIS is stimulated equally by all three activators. PKCn mediates calcium- and phosphatidylserine-dependent phosphorylation of both histone IIIS and partially purified lamin B. PKCn activity can be inhibited by an anti-PKC monoclonal antibody which specifically inhibits PKC. Isotype-specific PKC antibodies identify PKCn as βII-PKC. Immunoblot analysis indicates that HL60 cells express both α- and βII-PKC but no βI- or γ-PKC. Treatment of intact cells with bryo for 30 min leads to complete translocation of both α- and βII-PKC from the cytosol to the membrane fractions. Approximately 8-10% of the total βII-PKC (and 2 treatment leads to complete translocation of α-PKC, but only partial translocation of βII-PKC to the plasma membrane fraction. Neither PKC isotype is found associated with the nuclear membrane of PBt2-treated cells. These data demonstrate that α- and βII-PKC differ with respect to activator responsiveness, intracellular distribution, and substrate specificity and indicate that their selective activation at distinct intracellular sites, including the nucleus, can have a dramatic effect on resulting cellular responses.

UR - http://www.scopus.com/inward/record.url?scp=0026067787&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0026067787&partnerID=8YFLogxK

M3 - Article

VL - 266

SP - 28

EP - 33

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 1

ER -