|
|
 Previous Article | Table of Contents | Next Article 
Function of normal and mutated gamma-globin gene promoters in
electroporated K562 erythroleukemia cells
MJ Ulrich and TJ Ley
Department of Medicine, Jewish Hospital at Washington University Medical
Center, St Louis, MO 63110.
We examined the importance of cis-acting regulatory elements of the human
gamma-globin gene promoter in a cell line (K562) where this gene normally
functions. A gamma-Globin promoter fragments were fused to the neomycin
phosphotransferase (neoR) gene in a plasmid-based vector and transiently
transfected by electroporation into K562 cells. Correctly initiated "A
gamma-neo" transcripts were detected with an S1 nuclease protection assay
that was internally controlled for transfection efficiency and RNA content.
We first optimized the conditions for electroporation, and then determined
input DNA concentrations that permitted study of gamma-promoter function in
the linear range of the assay. We discovered that a gamma-globin promoter
fragment extending from -299 to +36 (with respect to the transcription
initiation site) was active in this transient transfection assay, and that
the expression of this promoter was increased by the SV40 enhancer.
Deletion of the gamma-globin promoter to position -199 did not
significantly affect gamma-globin promoter function. However, deletion to
-160 reduced gamma promoter strength to 70% that of control, deletion to
position -130 to 19% that of control, and deletion to position -61 to 8.7%
that of control. Three gamma-globin promoters containing mutations
associated with hereditary persistence of fetal hemoglobin (-202 C----G,
-196 C----T, and -117 G----A) were not overexpressed in the K562 cell
environment, consistent with the hypothesis that these promoters are not
overexpressed in fetal erythroblasts, only adult erythroid cells. This
system will allow us to further dissect the roles of regulatory globin
cis-acting DNA elements in fetal erythroid cells.
Volume 75,
Issue 4,
pp. 990-999,
02/15/1990
Copyright © 1990 by The American Society of Hematology
 CiteULike  Connotea  Del.icio.us  Digg  Reddit  Technorati What's this?
This article has been cited by other articles:
|
|
|
|
 
Q. Li, X. Fang, I. Olave, H. Han, M. Yu, P. Xiang, and G. Stamatoyannopoulos
Transcriptional potential of the {gamma}-globin gene is dependent on the CACCC box in a developmental stage-specific manner
Nucleic Acids Res.,
September 1, 2006;
34(14):
3909 - 3916.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
L.-R. Liu, Z.-W. Du, H.-L. Zhao, X.-L. Liu, X.-D. Huang, J. Shen, L.-M. Ju, F.-D. Fang, and J.-W. Zhang
T to C Substitution at -175 or -173 of the {gamma}-Globin Promoter Affects GATA-1 and Oct-1 Binding in Vitro Differently but Can Independently Reproduce the Hereditary Persistence of Fetal Hemoglobin Phenotype in Transgenic Mice
J. Biol. Chem.,
March 4, 2005;
280(9):
7452 - 7459.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
X. Fang, H. Han, G. Stamatoyannopoulos, and Q. Li
Developmentally Specific Role of the CCAAT Box in Regulation of Human {gamma}-Globin Gene Expression
J. Biol. Chem.,
February 13, 2004;
279(7):
5444 - 5449.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
P. A. Navas, Q. Li, K. R. Peterson, R. A. Swank, A. Rohde, J. Roy, and G. Stamatoyannopoulos
Activation of the {beta}-like globin genes in transgenic mice is dependent on the presence of the {beta}-locus control region
Hum. Mol. Genet.,
April 15, 2002;
11(8):
893 - 903.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
H. Asano, X. S. Li, and G. Stamatoyannopoulos
FKLF, a Novel Kruppel-Like Factor That Activates Human Embryonic and Fetal beta -Like Globin Genes
Mol. Cell. Biol.,
May 1, 1999;
19(5):
3571 - 3579.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
M. M. Zutter, A. D. Painter, and X. Yang
The Megakaryocyte/Platelet-Specific Enhancer of the alpha 2beta 1 Integrin Gene: Two Tandem AP1 Sites and the Mitogen-Activated Protein Kinase Signaling Cascade
Blood,
March 1, 1999;
93(5):
1600 - 1611.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
T. Ikuta, Y. W. Kan, P. S. Swerdlow, D. V. Faller, and S. P. Perrine
Alterations in Protein-DNA Interactions in the gamma -Globin Gene Promoter in Response to Butyrate Therapy
Blood,
October 15, 1998;
92(8):
2924 - 2933.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K. W. Makar, C. T. N. Pham, M. H. Dehoff, S. M. O'Connor, S. M. Jacobi, and V. M. Holers
An Intronic Silencer Regulates B Lymphocyte Cell- and Stage-Specific Expression of the Human Complement Receptor Type 2 (CR2, CD21) Gene
J. Immunol.,
February 1, 1998;
160(3):
1268 - 1278.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
H. A. a. G. Stamatoyannopoulos
Activation of beta -Globin Promoter by Erythroid Kruppel-Like Factor
Mol. Cell. Biol.,
January 1, 1998;
18(1):
102 - 109.
[Abstract]
[Full Text]
|
|
|
|
|
|
|
S. D. Langdon and R. E. Kaufman
Gamma-Globin Gene Promoter Elements Required for Interaction With Globin Enhancers
Blood,
January 1, 1998;
91(1):
309 - 318.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
P. G. McCaffrey, D. A. Newsome, E. Fibach, M. Yoshida, and M. S.-S. Su
Induction of gamma -Globin by Histone Deacetylase Inhibitors
Blood,
September 1, 1997;
90(5):
2075 - 2083.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S. Lehmann and D. A. Harris
Blockade of Glycosylation Promotes Acquistion of Scrapie-like Properties by the Prion Protein in Cultured Cells
J. Biol. Chem.,
August 22, 1997;
272(34):
21479 - 21487.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
M. M. Zutter, E. E. Ryan, and A. D. Painter
Binding of Phosphorylated Sp1 Protein to Tandem Sp1 Binding Sites Regulates alpha 2 Integrin Gene Core Promoter Activity
Blood,
July 15, 1997;
90(2):
678 - 689.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. TomHon, W. Zhu, D. Millinoff, K. Hayasaka, J. L. Slightom, M. Goodman, and D. L. Gumucio
Evolution of a Fetal Expression Pattern via cis Changes near the gamma Globin Gene
J. Biol. Chem.,
May 30, 1997;
272(22):
14062 - 14066.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
T. Ikuta, T. Papayannopoulou, G. Stamatoyannopoulos, and Y. W. Kan
Globin Gene Switching. IN VIVO PROTEIN-DNA INTERACTIONS OF THE HUMAN beta -GLOBIN LOCUS IN ERYTHROID CELLS EXPRESSING THE FETAL OR THE ADULT GLOBIN GENE PROGRAM
J. Biol. Chem.,
June 14, 1996;
271(24):
14082 - 14091.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
W. M. Nauseef, M. Cogley, and S. McCormick
Effect of the R569W Missense Mutation on the Biosynthesis of Myeloperoxidase
J. Biol. Chem.,
April 19, 1996;
271(16):
9546 - 9549.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S. Lehmann and D. A. Harris
A Mutant Prion Protein Displays an Aberrant Membrane Association When Expressed in Cultured Cells
J. Biol. Chem.,
October 13, 1995;
270(41):
24589 - 24597.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S.-L. Shyng, K. L. Moulder, A. Lesko, and D. A. Harris
The N-terminal Domain of a Glycolipid-anchored Prion Protein Is Essential for Its Endocytosis via Clathrin-coated Pits
J. Biol. Chem.,
June 16, 1995;
270(24):
14793 - 14800.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
L. H. Shapiro and L. H. Shapiro
Myb and Ets Proteins Cooperate to Transactivate an Early Myeloid Gene
J. Biol. Chem.,
April 14, 1995;
270(15):
8763 - 8771.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
