Ee. Bouhassira et al., TRANSCRIPTIONAL BEHAVIOR OF LCR ENHANCER ELEMENTS INTEGRATED AT THE SAME CHROMOSOMAL LOCUS BY RECOMBINASE-MEDIATED CASSETTE EXCHANGE, Blood, 90(9), 1997, pp. 3332-3344
Efficient integration of transgenes at preselected chromosomal locatio
ns was achieved in mammalian cells by recombinase-mediated-cassette-ex
change (RMCE), a novel procedure that makes use of the CRE recombinase
together with Lox sites bearing different spacer regions, We have app
lied RMCE to the study of the human beta-globin gene Locus Control Reg
ion by integrating at the same genetic locus in MEL cells, a LacZ gene
driven by the human beta-globin promoter linked to HS2 and HS3 alone
or in combination with HS4. Expression studies at the cell population
level and in individual cells before and after induction of differenti
ation with hemin or DMSO show that the presence of these enhancers is
associated with variegated patterns of expression, We were able to sho
w that the LCR fragments tested act by controlling both the probabilit
y of expression and the rare of transcription of the linked beta-globi
n promoter, Both of these factors were also dependent on the state of
differentiation of the MELc and on the presence of a second transcript
ion unit located in cis. The ability to manipulate by RMCE constructs
integrated into chromosomes should help in the creation of complex, ra
tionally designed, artificial genetic loci. (C) 1997 by The American S
ociety of Hematology.