In vivo analysis of retroviral enhancer mutations in hematopoietic cells: SP1/EGR1 and ETS/GATA motifs contribute to long terminal repeat specificity.
J Virol
; 76(1): 303-12, 2002 Jan.
Article
em En
| MEDLINE
| ID: mdl-11739695
ABSTRACT
The objective of this work was to identify, in the context of chromosomally integrated DNA, the contribution of defined transcription factor binding motifs to the function of a complex retrovirus enhancer in hematopoietic cells in vivo. Repopulating murine hematopoietic cells were transduced with equal gene dosages of replication-incompetent retrovirus vectors encoding enhanced green fluorescent protein. Enhancer sequences were derived from mouse spleen focus-forming virus. Destruction of GC-rich sites representing overlapping targets for SP1 or EGR1 uniformly attenuated gene expression (approximately 25 to 70% of wild-type levels) in all hematopoietic lineages, as shown by multicolor flow cytometry of peripheral blood and bone marrow cells at various time points posttransplantation. In contrast, a point mutation within a dual ETS/GATA motif that abolished transactivation by ETS factors but not by GATA-1 slightly increased activity in erythroid cells and significantly attenuated enhancer function in T lymphocytes. This study shows that controlled gene transfer in transplantable hematopoietic cells allows a functional analysis of distinct cis elements within a complex retrovirus enhancer, as required for the characterization and engineering of various cellular and viral regulatory sequences in basic research and gene therapy.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Retroviridae
/
Células-Tronco Hematopoéticas
/
Elementos Facilitadores Genéticos
/
Sequências Repetidas Terminais
Tipo de estudo:
Prognostic_studies
Limite:
Animals
Idioma:
En
Revista:
J Virol
Ano de publicação:
2002
Tipo de documento:
Article
País de afiliação:
Alemanha