Granulocyte-macrophage colony-stimulating factor mRNA stabilization enhances transgenic expression in normal cells and tissues

L. E. Rajagopalan, J. K. Burkholder, J. Turner, J. Culp, N. S. Yang, J. S. Malter

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

To increase transgenic production of granulocyte-macrophage colony- stimulating factor (GM-CSF), we mutated the mRNA's 3'-untranslated region, AUUUA instability elements. Expression vectors containing human or murine GM- CSF cDNAs coding for wild-type (GM-AUUUA) or mutant versions with reiterated AUGUA repeats (GM-AUGUA) were transfected into cells in culture or animals using particle-mediated gene-transfer technology. Normal peripheral blood mononuclear cells accumulated 20-fold greater levels of GM-CSF mRNA and secreted comparably greater amounts of cytokine after transfection with hGM- AUGUA expression vectors versus hGM-AUUUA. hGM-AUGUA mRNA was fivefold more stable (t( 1/2 ) = 95 minutes) than hGM-AUUUA mRNA (t( 1/2 ) = 20 minutes), accounting for elevated steady-state levels. Transfection site extracts and serum samples obtained 24 hours after gene transfer of hGM-AUGUA cDNA into mouse skin contained greater than 32 ng/mL and 650 pg/mL of GM-CSF protein, respectively, compared with 0.33 ng/mL and less than 8 pg/mL for hGM-AUUUA cDNA. GM-CSF produced from mGM-AUGUA cDNA transfected into rat abdominal epidermis induced a profound neutrophil infiltrate. These data suggest a novel strategy for enhanced production of biologically active cytokines by normal cells after in vivo gene transfer.

Original languageEnglish (US)
Pages (from-to)2551-2558
Number of pages8
JournalBlood
Volume86
Issue number7
StatePublished - 1995

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Granulocyte-Macrophage Colony-Stimulating Factor
Gene transfer
Stabilization
Tissue
Messenger RNA
Complementary DNA
Transfection
Cytokines
Genes
Technology Transfer
3' Untranslated Regions
Epidermis
Rats
Blood Cells
Skin
Animals
Neutrophils
Blood
Cell Culture Techniques
Serum

ASJC Scopus subject areas

  • Hematology

Cite this

Rajagopalan, L. E., Burkholder, J. K., Turner, J., Culp, J., Yang, N. S., & Malter, J. S. (1995). Granulocyte-macrophage colony-stimulating factor mRNA stabilization enhances transgenic expression in normal cells and tissues. Blood, 86(7), 2551-2558.

Granulocyte-macrophage colony-stimulating factor mRNA stabilization enhances transgenic expression in normal cells and tissues. / Rajagopalan, L. E.; Burkholder, J. K.; Turner, J.; Culp, J.; Yang, N. S.; Malter, J. S.

In: Blood, Vol. 86, No. 7, 1995, p. 2551-2558.

Research output: Contribution to journalArticle

Rajagopalan, LE, Burkholder, JK, Turner, J, Culp, J, Yang, NS & Malter, JS 1995, 'Granulocyte-macrophage colony-stimulating factor mRNA stabilization enhances transgenic expression in normal cells and tissues', Blood, vol. 86, no. 7, pp. 2551-2558.
Rajagopalan, L. E. ; Burkholder, J. K. ; Turner, J. ; Culp, J. ; Yang, N. S. ; Malter, J. S. / Granulocyte-macrophage colony-stimulating factor mRNA stabilization enhances transgenic expression in normal cells and tissues. In: Blood. 1995 ; Vol. 86, No. 7. pp. 2551-2558.
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