Combined effects of aberrant MEK1 activity and BCL2 overexpression on relieving the cytokine dependency of human and murine hematopoietic cells

W. L. Blalock, P. W. Moye, Fumin Chang, M. Pearce, L. S. Steelman, M. McMahon, J. A. McCubrey

Research output: Contribution to journalArticle

Abstract

The MEK1 oncoprotein plays a critical role in Ras/Raf/MEK/MAPK-mediated transmission of mitogenic signals from cell surface receptors to the nucleus. In order to examine this pathway's role in leukemic transformation, a conditionally active (β-estradiol-inducible) form of the MEK1 protein was created by ligating a cDNA encoding an N-terminal truncated form of MEK1 to the hormone-binding domain of the estrogen receptor (ER). We introduced this chimeric ΔMEK1:ER oncoprotein into cytokine-dependent human TF-1 and murine FDC-P1 hematopoietic cell lines. Two different types of cells were recovered after drug selection in medium containing either cytokine or β-estradiol: (1) cells that expressed the ΔMEK1:ER oncoprotein but remained cytokine-dependent and (2) MEK1-responsive cells that grew in response to ΔMEK1:ER activation. Cytokine-dependent cells were recovered 102 to 104 times more frequently than MEK1-responsive cells depending upon the particular cell line. To determine whether BCL2 overexpression could synergize with the ΔMEK1:ER oncoprotein in relieving cytokine dependence, the cytokine-dependent ΔMEK1:ER-expressing cells were infected with a BCL2-containing retrovirus, and the frequency of MEK1-responsive cells determined. BCL2 overexpression, by itself, did not relieve cytokine dependency of the parental cells, however, it did increase the frequency at which MEK1-responsive cells were recovered approximately 10-fold. ΔMEK1:ER+BCL2 cells remained viable for at least 3 days after estradiol deprivation, whereas viability was readily lost upon withdrawal of β-estradiol in the MEK1-responsive cells which lacked BCL2 overexpression. The MAP kinases, ERK1 and ERK2 were activated in response to ΔMEK1:ER stimulation in both ΔMEK1:ER and ΔMEK1:ER+BCL2 cells. As compared to the cytokine-dependent ΔMEK1:ER and BCL2 infected cells, MEK1-responsive BCL2 infected cells expressed higher levels of BCL2. While both MEK1-responsive ΔMEK1:ER and ΔMEK1:ER+BCL2 infected cells expressed cDNAs encoding the autocrine cytokine GM-CSF, more GM-CSF cDNAs and bioactivity were detected in the MEK1-responsive ΔMEK1:ER+BCL2 cells than in the MEK1-responsive cells lacking BCL2 or cytokine-dependent cells. These conditionally transformed cells will be useful in furthering our understanding of the roles MEK1 and BCL2 play in the prevention of apoptosis in hematopoietic cells.

Original languageEnglish (US)
Pages (from-to)1080-1096
Number of pages17
JournalLeukemia
Volume14
Issue number6
DOIs
StatePublished - Jan 1 2000
Externally publishedYes

Fingerprint

Cytokines
Estrogen Receptors
Oncogene Proteins
Estradiol
Granulocyte-Macrophage Colony-Stimulating Factor
Complementary DNA
Cell Line
Mitogen-Activated Protein Kinase 3
Mitogen-Activated Protein Kinase Kinases
Cell Surface Receptors
Retroviridae
Hormones
Apoptosis

Keywords

  • Apoptosis
  • BCL2
  • Cytokine dependency
  • MEK1

ASJC Scopus subject areas

  • Hematology
  • Oncology
  • Cancer Research

Cite this

Combined effects of aberrant MEK1 activity and BCL2 overexpression on relieving the cytokine dependency of human and murine hematopoietic cells. / Blalock, W. L.; Moye, P. W.; Chang, Fumin; Pearce, M.; Steelman, L. S.; McMahon, M.; McCubrey, J. A.

In: Leukemia, Vol. 14, No. 6, 01.01.2000, p. 1080-1096.

Research output: Contribution to journalArticle

Blalock, W. L. ; Moye, P. W. ; Chang, Fumin ; Pearce, M. ; Steelman, L. S. ; McMahon, M. ; McCubrey, J. A. / Combined effects of aberrant MEK1 activity and BCL2 overexpression on relieving the cytokine dependency of human and murine hematopoietic cells. In: Leukemia. 2000 ; Vol. 14, No. 6. pp. 1080-1096.
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