Abstract
Retroviral overexpression of NF-Ya, the regulatory subunit of the transcription factor NF-Y, activates the transcription of multiple genes implicated in hematopoietic stem cell (HSC) self-renewal and differentiation and directs HSCs toward self-renewal. We asked whether TAT-NF-Ya fusion protein could be used to transduce human CD34(+) cells as a safer, more regulated alternative approach to gene therapy. Here we show that externally added recombinant protein was able to enter the cell nucleus and activate HOXB4, a target gene of NF-Ya, using real-time polymerase chain reaction RNA and luciferase-based protein assays. After TAT-NF-Ya transduction, the proliferation of human CD34(+) cells in the presence of myeloid cytokines was increased 4-fold. Moreover, TAT-NF-Ya-treated human primary bone marrow cells showed a 4-fold increase in the percentage of huCD45(+) cells recovered from the bone marrow of sublethally irradiated, transplanted NOD-Scid IL2Rγ(null) mice. These data demonstrate that TAT-peptide therapies are an alternative approach to retroviral stem cell therapies and suggest that NF-Ya peptide delivery should be further evaluated as a tool for HSC/progenitors ex vivo expansion and therapy.
Publication types
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Research Support, N.I.H., Extramural
MeSH terms
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Animals
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CCAAT-Binding Factor / genetics*
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CCAAT-Binding Factor / metabolism*
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Cell Proliferation
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Genetic Therapy
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Hematopoietic Stem Cell Transplantation
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Hematopoietic Stem Cells / cytology*
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Hematopoietic Stem Cells / metabolism*
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Homeodomain Proteins / genetics
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Humans
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In Vitro Techniques
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Interleukin Receptor Common gamma Subunit / deficiency
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Interleukin Receptor Common gamma Subunit / genetics
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Mice
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Mice, Inbred NOD
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Mice, Knockout
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Mice, SCID
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Peptide Fragments / genetics*
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Peptide Fragments / metabolism*
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Promoter Regions, Genetic
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Protein Binding
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Transcription Factors / genetics
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Transcriptional Activation
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Transduction, Genetic*
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Transplantation, Heterologous
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tat Gene Products, Human Immunodeficiency Virus / genetics*
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tat Gene Products, Human Immunodeficiency Virus / metabolism*
Substances
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CCAAT-Binding Factor
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HOXB4 protein, human
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Homeodomain Proteins
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Interleukin Receptor Common gamma Subunit
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NFYA protein, human
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Peptide Fragments
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Recombinant Fusion Proteins
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Transcription Factors
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tat Gene Products, Human Immunodeficiency Virus
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tat peptide (47-57), Human immunodeficiency virus 1