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Title: In vivo models for elucidation of role of MLL-AF9 chromosomal translocations in leukaemia
Author: Collins, E. C. J.
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2002
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Chromosomal translocations involving the MLL and AF9 genes in human acute myeloid leukaemia were investigated using two complementary in vivo models. LoxP sites were introduced into the murine Mll and Af9 genes by homologous recombination in embryonic stem (ES) cells, at positions corresponding to the breakpoints observed in human patients. Generation of de novo inter-chromosomal recombination events was evaluated by transient expression of Cre recombinase in doubly transfected ES cells. The presence and correct expression of the expected fusion gene were verified by polymerase chain reaction and sequence analysis. Subsequently, mouse strains were generated that carried the modified Mll and Af9 alleles. After inter-breeding with a mouse strain ubiquitously expressing a cre transgene, Mll-Af9 fusion genes were detected in tissues of adult animals, indicating that recombination - presumably translocation - had occurred during mouse development. This constitutes the first example of Cre-loxP-dependent inter-chromosomal rearrangement in mammals, and should provide a valuable tool to study translocation fusion genes associated with human cancers as well as potential therapeutic regimes. Furthermore, with a view to elucidation of the role of the AF9 gene, its murine homologue was inactivated by homologous recombination in ES cells and subsequent generation of Af9 null mutant mice. Homozygous gene disruption was lethal shortly after birth, but the mutation was found to have no discernible effect on haematopoiesis as determined by histology and FACS analysis. However, knock-out mice exhibited anomalies of the axial skeleton, indicating a role of Af9 in embryo patterning. Both the cervical and thoracic regions appeared affected by anterior homeotic transformation. Strikingly, mice lacking functional Af9 exhibited a grossly deformed atlas, as well as an extra cervical vertebra, an effect not hitherto observed in any known mouse mutants. This phenotype suggests that the MLL-AF9 oncogene affects key regulators of embryo patterning processes also involved in leukaemogenesis.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available