Upstream conserved sequences of mouse leukemia viruses are important for high transgene expression in lymphoid and hematopoietic cells
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Upstream conserved sequences of mouse leukemia viruses are important for high transgene expression in lymphoid and hematopoietic cells. / Wahlers, Anke; Kustikova, Olga; Zipfel, Peter F; Itoh, Katsuhiko; Koester, Markus; Heberlein, Christoph; Li, Zhixiong; Schiedlmeier, Bernd; Skerka, Christine; Fehse, Boris; Baum, Christopher.
in: MOL THER, Jahrgang 6, Nr. 3, 09.2002, S. 313-320.Publikationen: SCORING: Beitrag in Fachzeitschrift/Zeitung › SCORING: Zeitschriftenaufsatz › Forschung › Begutachtung
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TY - JOUR
T1 - Upstream conserved sequences of mouse leukemia viruses are important for high transgene expression in lymphoid and hematopoietic cells
AU - Wahlers, Anke
AU - Kustikova, Olga
AU - Zipfel, Peter F
AU - Itoh, Katsuhiko
AU - Koester, Markus
AU - Heberlein, Christoph
AU - Li, Zhixiong
AU - Schiedlmeier, Bernd
AU - Skerka, Christine
AU - Fehse, Boris
AU - Baum, Christopher
PY - 2002/9
Y1 - 2002/9
N2 - Highly conserved enhancer sequences located in the upstream part of the long terminal repeat (LTR) of murine leukemia retroviruses (MLV) were reported to compromise viral gene expression in multipotent embryonic cells in vitro and to reduce the likelihood for maintenance of retroviral gene expression in hematopoietic cells in vivo. We show that deletion of these sequences (nucleotides +37 to +95) attenuates rather than increases the transcriptional activity of retroviral vectors in hematopoietic cells almost independently of the developmental lineage (erythroid, myeloid, or lymphoid). Expression rates of modified vectors were reduced by as much as 34-65%, although the strong enhancer array located in the direct repeat of the LTR was preserved. Sequence analysis and electrophoretic mobility shift assays revealed the presence of a highly conserved binding site for NFAT (nuclear factor of activated T cells) proteins that immediately neighbors a known binding site for the transcription factor Yin-Yang1 (YY1) [corrected]. Specific inactivation of the NFAT site reduced transgene expression in all cell types investigated and had a similar effect as the destruction of a neighboring SP1 motif. Combined destruction of individual motifs for NFAT, SP1, and E twenty-six transcription factors (ETS) resulted in a severe attenuation (by 40-60%) of the retroviral enhancer. These results provide novel clues for the manipulation of retrovirus replication and vector tropism.
AB - Highly conserved enhancer sequences located in the upstream part of the long terminal repeat (LTR) of murine leukemia retroviruses (MLV) were reported to compromise viral gene expression in multipotent embryonic cells in vitro and to reduce the likelihood for maintenance of retroviral gene expression in hematopoietic cells in vivo. We show that deletion of these sequences (nucleotides +37 to +95) attenuates rather than increases the transcriptional activity of retroviral vectors in hematopoietic cells almost independently of the developmental lineage (erythroid, myeloid, or lymphoid). Expression rates of modified vectors were reduced by as much as 34-65%, although the strong enhancer array located in the direct repeat of the LTR was preserved. Sequence analysis and electrophoretic mobility shift assays revealed the presence of a highly conserved binding site for NFAT (nuclear factor of activated T cells) proteins that immediately neighbors a known binding site for the transcription factor Yin-Yang1 (YY1) [corrected]. Specific inactivation of the NFAT site reduced transgene expression in all cell types investigated and had a similar effect as the destruction of a neighboring SP1 motif. Combined destruction of individual motifs for NFAT, SP1, and E twenty-six transcription factors (ETS) resulted in a severe attenuation (by 40-60%) of the retroviral enhancer. These results provide novel clues for the manipulation of retrovirus replication and vector tropism.
KW - 5' Flanking Region
KW - Animals
KW - Base Sequence
KW - Binding Sites/genetics
KW - Conserved Sequence
KW - DNA-Binding Proteins/metabolism
KW - Enhancer Elements, Genetic
KW - Fibroblasts/physiology
KW - Genetic Vectors
KW - Hematopoietic Stem Cells/physiology
KW - Leukemia Virus, Murine
KW - Lymphocytes/physiology
KW - Mice
KW - Molecular Sequence Data
KW - NFATC Transcription Factors
KW - Nuclear Proteins
KW - Retroviridae/genetics
KW - Transcription Factors/metabolism
KW - Transgenes/physiology
U2 - 10.1006/mthe.2002.0671
DO - 10.1006/mthe.2002.0671
M3 - SCORING: Journal article
C2 - 12231166
VL - 6
SP - 313
EP - 320
JO - MOL THER
JF - MOL THER
SN - 1525-0016
IS - 3
ER -
