International Mammalian Genome Society

The 15th International Mouse Genome Conference (2001)


Dr Melany Jackson
University of Edinburgh
John Hughes Bennett Laboratory
Western General Hospital
Crewe Road
Edinburgh EH4 2XU

Co-Authors: 1) Baird J, 2) Cambray N,  2) Ansell J, 1)Graham G,  2) Forrester L.
Institutions: 1) The Beatson Institute, 2) John Hughes Bennett Laboratory, University of Edinburgh

The differentiation of ES cells as embryoid bodies in vitro is a valuable model for embryonic development and for the functional characterisation of genes involved in this process. To identify novel genes that may be involved in embryogenesis we generated a subtracted cDNA library from RNA isolated from day 5 and day 3 embryoid bodies. One of these cDNAs, (ehox) encodes a novel gene that contains a homeodomain and was  identified as a potentially interesting developmentally-regulated gene.   We used an episomal overexpression system as an initial screen to assess the function of ehox in ES cells. Plasmids carrying the ehox cDNA in sense or anti-sense orientations were transfected into ES cells and colonies were selected in puromycin.  Transfection experiments indicated that  high levels of eHOX  expression was incompatible with an undifferentiated ES cell phenotype.  In addition, we observed that ES cells expressing anti-sense ehox maintained a stem cell phenotype  in low concentrations of LIF whereas ES cells expressing sense ehox, EGFP or the empty vector, differentiated in a comparable manner to parental ES cell lines. ES cells expressing anti-sense ehox are also unable to differentiate into beating cardiomyocytes in contrast to eHOX or GFP-expressing cells.  Our data therefore suggest that eHOX is required either for the initial differentiation of pluripotent ES cells or for the survival of differentiated cell types.

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