Granulocytic differentiation of HL-60 promyelocytic leukemia cells is associated with increased expression of Cul5

Shaneen S. Baxter, Lauren A. Carlson, Alejandro M.S. Mayer, Mary L. Hall, Michael J. Fay

Research output: Contribution to journalArticle

10 Scopus citations

Abstract

The human HL-60 promyelocytic leukemia cell line has been widely used as a model for studying granulocytic differentiation. All-trans retinoic acid (ATRA) treatment of HL-60 cells promotes granulocytic differentiation and is effective as differentiation therapy for patients with acute promyelocytic leukemia. The identification of genes that are transcriptionally regulated by ATRA has provided insight into granulocytic differentiation and differentiation therapy. The Asb-2 (ankyrin repeat SOCS box 2) gene has previously been identified as a transcriptional target in ATRA-treated HL-60 cells. The ASB-2 protein forms an E3 ubiquitin ligase complex with the proteins, Cul5, regulator of cullin 2 (ROC2), and elongin B and C. The purpose of this study was to determine if there is increased expression of Cul5 during granulocytic differentiation of HL-60 cells. To induce granulocytic differentiation, HL-60 cells were treated for 5 d with ATRA and differentiation was confirmed by examining superoxide anion production, nuclear morphology, and changes in the expression of CD11b, CD13, and CD15. Quantitative realtime RT-PCR was used to measure Cul5 mRNA expression and also the expression of other components of the E3 ubiquitin ligase (ASB-2, ROC2, elongin B and C). Granulocytic differentiation of HL-60 cells was associated with a 1.6-, 1.7-, and 23-fold statistically significant (P=0.05) increase in mRNA expression for Cul5, ROC2, and ASB-2, respectively. No significant change was found in elongin B and C mRNA expression. Using Western blot analysis, the expression of Cul5 protein was increased 6.5-fold with granulocytic differentiation of the HL-60 cells. Increased expression of multiple components of the Cul5-containing E3 ubiquitin ligase complex with ATRA treatment of HL-60 cells indicates that this complex may play an important role in granulocytic differentiation.

Original languageEnglish (US)
Pages (from-to)264-274
Number of pages11
JournalIn Vitro Cellular and Developmental Biology - Animal
Volume45
Issue number5-6
DOIs
StatePublished - 2009
Externally publishedYes

Keywords

  • All-trans retinoic acid
  • ASB-2
  • Cul5
  • Elongin B/C
  • Granulocytic differentiation
  • HL-60
  • ROC2

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