Secretory TAT-peptide-mediated protein transduction of LIF receptor α-chain distal cytoplasmic motifs into human myeloid HL-60 cells

Secretory TAT-peptide-mediated protein transduction of LIF receptor α-chain distal cytoplasmic motifs into human myeloid HL-60 cells

The distal cytoplasmic motifs of leukemia inhibitory factor receptor α-chain (LIFRα-CT3) can independently induce intracellular myeloid differentiation in acute myeloid leukemia (AML) cells by gene transfection; however, there are significant limitations in the potential clinical u...

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Bibliographic Details
Main Authors: Q. Sun, J. Xiong, J. Lu, S. Xu, Y. Li, X.P. Zhong, G.K. Gao, H.Q. Liu
Format: Article
Language:English
Published: Associação Brasileira de Divulgação Científica 2012-10-01
Series:Brazilian Journal of Medical and Biological Research
Subjects:
Leukemia inhibitory factor
TAT-HIV1
Protein transduction domain
Acute myeloid leukemia
LIF receptor
Online Access:http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2012001000005
Description
Summary:The distal cytoplasmic motifs of leukemia inhibitory factor receptor &#945;-chain (LIFR&#945;-CT3) can independently induce intracellular myeloid differentiation in acute myeloid leukemia (AML) cells by gene transfection; however, there are significant limitations in the potential clinical use of these motifs due to liposome-derived genetic modifications. To produce a potentially therapeutic LIFR&#945;-CT3 with cell-permeable activity, we constructed a eukaryotic expression pcDNA3.0-TAT-CT3-cMyc plasmid with a signal peptide (ss) inserted into the N-terminal that codes for an ss-TAT-CT3-cMyc fusion protein. The stable transfection of Chinese hamster ovary (CHO) cells via this vector and subsequent selection by Geneticin resulted in cell lines that express and secrete TAT-CT3-cMyc. The spent medium of pcDNA3.0-TAT-CT3-cMyc-transfected CHO cells could be purified using a cMyc-epitope-tag agarose affinity chromatography column and could be detected via SDS-PAGE, with antibodies against cMyc-tag. The direct administration of TAT-CT3-cMyc to HL-60 cell culture media caused the enrichment of CT3-cMyc in the cytoplasm and nucleus within 30 min and led to a significant reduction of viable cells (P < 0.05) 8 h after exposure. The advantages of using this mammalian expression system include the ease of generating TAT fusion proteins that are adequately transcripted and the potential for a sustained production of such proteins in vitro for future AML therapy.
ISSN:0100-879X
1414-431X

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