Activation of protein kinase C induces nuclear translocation of RFX1 and down-regulates c-myc via an intron 1 X box in undifferentiated leukemia HL-60 cells

Lei Chen, Lucinda Smith, Martin R. Johnson, Kangsheng Wang, Robert B Diasio, Jeffrey Bingham Smith

Research output: Contribution to journalArticle

33 Citations (Scopus)

Abstract

Treatment of human promyelocytic leukemia ceils (HL-60) with phorbol 12-myristate 13-acetate (PMA) is known to decrease c-myc mRNA by blocking transcription elongation at sites near the first exon/intron border. Treatment of HL-60 cells with either PMA or bryostatin 1, which acutely activates protein kinase C (PKC), decreased the levels of myc mRNA and Myc protein. The inhibition of Myc synthesis accounted for the drop in Myc protein, because PMA treatment had no effect on Myc turnover. Treatment with PMA or bryostatin 1 increased nuclear protein binding to MIE1, a c-myc intron 1 element that defines an RFX1-binding X box. RFX1 antiserum supershifted MIE1-protein complexes. Increased MIE1 binding was independent of protein synthesis and abolished by a selective PKC inhibitor, which also prevented the effect of PMA on myc mRNA and protein levels and Myc synthesis. PMA treatment increased RFX1 in the nuclear fraction and decreased it in the cytosol without affecting total RFX1. Transfection of HL-60 cells with myc reporter gene constructs showed that the RFX1-binding X box was required for the down-regulation of reporter gene expression by PMA. These findings suggest that nuclear translocation and binding of RFX1 to the X box cause the down-regulation of myc expression, which follows acute PKC activation in undifferentiated HL-60 cells.

Original languageEnglish (US)
Pages (from-to)32227-32233
Number of pages7
JournalJournal of Biological Chemistry
Volume275
Issue number41
StatePublished - Oct 13 2000
Externally publishedYes

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HL-60 Cells
Introns
Protein Kinase C
Leukemia
Acetates
Down-Regulation
Chemical activation
Proteins
Reporter Genes
Messenger RNA
myc Genes
Protein C Inhibitor
Gene Expression Regulation
Transcription
Protein Kinase Inhibitors
Nuclear Proteins
phorbol-12-myristate
Protein Binding
Gene expression
Cytosol

ASJC Scopus subject areas

  • Biochemistry

Cite this

Activation of protein kinase C induces nuclear translocation of RFX1 and down-regulates c-myc via an intron 1 X box in undifferentiated leukemia HL-60 cells. / Chen, Lei; Smith, Lucinda; Johnson, Martin R.; Wang, Kangsheng; Diasio, Robert B; Smith, Jeffrey Bingham.

In: Journal of Biological Chemistry, Vol. 275, No. 41, 13.10.2000, p. 32227-32233.

Research output: Contribution to journalArticle

Chen, Lei ; Smith, Lucinda ; Johnson, Martin R. ; Wang, Kangsheng ; Diasio, Robert B ; Smith, Jeffrey Bingham. / Activation of protein kinase C induces nuclear translocation of RFX1 and down-regulates c-myc via an intron 1 X box in undifferentiated leukemia HL-60 cells. In: Journal of Biological Chemistry. 2000 ; Vol. 275, No. 41. pp. 32227-32233.
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abstract = "Treatment of human promyelocytic leukemia ceils (HL-60) with phorbol 12-myristate 13-acetate (PMA) is known to decrease c-myc mRNA by blocking transcription elongation at sites near the first exon/intron border. Treatment of HL-60 cells with either PMA or bryostatin 1, which acutely activates protein kinase C (PKC), decreased the levels of myc mRNA and Myc protein. The inhibition of Myc synthesis accounted for the drop in Myc protein, because PMA treatment had no effect on Myc turnover. Treatment with PMA or bryostatin 1 increased nuclear protein binding to MIE1, a c-myc intron 1 element that defines an RFX1-binding X box. RFX1 antiserum supershifted MIE1-protein complexes. Increased MIE1 binding was independent of protein synthesis and abolished by a selective PKC inhibitor, which also prevented the effect of PMA on myc mRNA and protein levels and Myc synthesis. PMA treatment increased RFX1 in the nuclear fraction and decreased it in the cytosol without affecting total RFX1. Transfection of HL-60 cells with myc reporter gene constructs showed that the RFX1-binding X box was required for the down-regulation of reporter gene expression by PMA. These findings suggest that nuclear translocation and binding of RFX1 to the X box cause the down-regulation of myc expression, which follows acute PKC activation in undifferentiated HL-60 cells.",
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