Falconer-2010-Identification of sister chromatid.pdfVIP

LETTERS Identification of sister chromatids by DNA template strand sequences Ester Falconer1, Elizabeth A. Chavez1, Alexander Henderson1, Steven S. S. Poon1,2, Steven McKinney2, Lindsay Brown3, David G. Huntsman3 Peter M. Lansdorp1,4 It is generally assumed that sister chromatids are genetically and functionally identical and that segregation to daughter cells is a random process. However, functional differences between sister chromatids regulate daughter cell fate in yeast1 and sister chromatid segregation is not random in Escherichia coli 2. Differentiated sister chromatids, coupled with non-random segregation, have been pro- posed to regulate cell fate during the development of multicellular organisms3. This hypothesis has not been tested because molecular features to reliably distinguish between sister chromatids are not obvious. Here we show that parental ‘Watson’ and ‘Crick’ DNA template strands can be identified in sister chromatids of murine metaphase chromosomes using CO-FISH (chromosome orientation fluorescence in situ hybridization4) with unidirectional probes specific for centromeric and telomeric repeats. All chromosomes were found to have a uniform orientation with the 59 end of the short arm on the same strand as T-rich major satellite repeats. The invariable orientation of repetitive DNA was used to differentially label sister chromatids and directly study mitotic segregation patterns in different cell types. Whereas sister chromatids appeared to be randomly distributed between daughter cells in cultured lung fibroblasts and embryonic stem cells, significant non-random sister chromatid segregation was observed in a subset of colon crypt epi- thelial cells, including cells outside positions reported for colon stem cells5. Our results establish that DNA template sequences can be used to distinguish sister chromatids and follow their mitotic segregation in vivo. Major satellite repeats have a uniform head-to-tail orientation on mouse chromosomes relati