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Proliferation and Differentiation of Murine Myeloid Precursor 32D/G-CSF-R Cells
Published on: February 21, 2018
Differences in expression of transcription factor AP-1 in human promyelocytic HL-60 cells during differentiation
F Mollinedo1, C Gajate, A Tugores
1Centro de Investigaciones Biológicas, C.S.I.C., Veláquez, Madrid, Spain.
Abstract:
Commitment of HL-60 cells to macrophage or granulocytic differentiation was achieved by incubation with 4 beta-phorbol 12-myristate 13-acetate (PMA) for 30-60 min or with dimethyl sulphoxide (DMSO) for 24 h respectively. The commitment stage towards PMA-induced macrophage differentiation was associated with increases in jun B and c-fos mRNA levels, as well as with an increase in the binding activity of transcription factor AP-1. Nevertheless, gel retardation analysis indicated that the AP-1 activity detected in untreated cells was drastically reduced during the commitment stage of DMSO-induced HL-60 differentiation towards granulocytes. When HL-60 cells were treated with sodium butyrate, which induced monocytic differentiation, a remarkable increase in AP-1 binding activity was detected. Treatment of HL-60 cells with 1 alpha,25-dihydroxyvitamin D3, another monocytic differentiation agent, induced a weak, but appreciable, increase in AP-1 activity. Furthermore, addition of sodium butyrate or 1 alpha,25-dihydroxyvitamin D3 to HL-60 cells induced the expression of c-fos, c-jun, jun B and jun D proto-oncogenes. In contrast, when HL-60 cells were treated with retinoic acid, a granulocytic differentiation inducer, no enhanced AP-1 binding activity was observed, and only a weak increase in jun D mRNA level was detected. These data indicate that formation of AP-1 is not required for the induction of HL-60 differentiation towards granulocytes, whereas induction of monocytic differentiation is correlated with an increase in AP-1 activity. The differential expression of AP-1 activity may be critical in the differentiation of HL-60 cells towards monocytic or granulocytic lineages.
Insights
AP-1 transcription factor activity is crucial for monocytic differentiation of HL-60 cells but not for granulocytic differentiation. This highlights AP-1
Area of Science:
- Cellular and Molecular Biology
- Hematopoiesis
- Transcription Factors
Background:
- HL-60 cells are a human promyelocytic leukemia cell line widely used to study myeloid differentiation.
- The transcription factor Activator Protein-1 (AP-1) plays a role in various cellular processes, including differentiation.
- Understanding the role of AP-1 in HL-60 cell differentiation can provide insights into myeloid lineage commitment.
Purpose of the Study:
- To investigate the role of AP-1 activity during the differentiation of HL-60 cells induced by different agents.
- To determine if AP-1 is differentially regulated during monocytic versus granulocytic differentiation of HL-60 cells.
Main Methods:
- HL-60 cells were induced to differentiate into macrophages using 4 beta-phorbol 12-myristate 13-acetate (PMA).
- Granulocytic differentiation was induced using dimethyl sulfoxide (DMSO).
- Monocytic differentiation was induced using sodium butyrate or 1 alpha,25-dihydroxyvitamin D3.
- AP-1 binding activity was assessed using gel retardation analysis.
- Proto-oncogene mRNA levels (c-fos, c-jun, jun B, jun D) were analyzed.
Main Results:
- PMA-induced macrophage differentiation increased jun B, c-fos mRNA, and AP-1 binding activity.
- DMSO-induced granulocytic differentiation showed reduced AP-1 activity during commitment.
- Sodium butyrate and 1 alpha,25-dihydroxyvitamin D3 induced monocytic differentiation with increased AP-1 binding activity and proto-oncogene expression.
- Retinoic acid-induced granulocytic differentiation did not enhance AP-1 binding activity.
Conclusions:
- AP-1 formation is not essential for granulocytic differentiation of HL-60 cells.
- Increased AP-1 activity is correlated with the induction of monocytic differentiation in HL-60 cells.
- Differential regulation of AP-1 activity is critical for determining the lineage commitment of HL-60 cells.
