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 coli2. Differentiated sister chromatids, coupled with non-random segregation, have been proposed 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 5′ 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 epithelial 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.
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K. Lisaingo helped acquire images for this study, J. Tan helped with image analysis and C. J. S. Smith assisted with in vivo studies. We thank S. Selig, T. de Lange and R. C. Wang for help with CO-FISH protocols. S. Aparicio, L. Lefebvre, L. F. Lansdorp, B. M. Lansdorp and members of the Lansdorp laboratory are thanked for discussions and for critically reading the manuscript. This study was funded in part by grants from the Canadian Institutes of Health Research (RMF-92093), the Michael Smith Foundation for Health Research, the Canadian Cancer Society Research Institute and the Terry Fox Foundation. We are grateful to the British Columbia Cancer Agency, the Canada Foundation for Innovation, the British Columbia Knowledge Development Fund, the British Columbia Cancer Foundation, the Blossom Fund of the University of British Columbia and the Mahon family for funding the infrastructure that enabled this work.
Author Contributions E.F. helped with the design of the experiments, image acquisition, data analysis, interpretation of results and writing of the paper. E.A.C. performed most of the CO-FISH experiments. A.H. performed most of the mouse work. L.B. acquired some images for this study. S.S.S.P. performed analysis of digital data and helped with statistical analysis that was performed by S.M. D.G.H. helped with the design of the study and interpretation of results. P.M.L. conceived the study, helped with image acquisition, interpretation of results and writing of the paper.
This file contains Supplementary Figures 1-11 with Legends, Supplementary Notes, Supplementary Video Legends 1-4, Supplementary Data Legend and Supplementary Data Image Gallery Legend. (PDF 3547 kb)
This movie file shows the distribution of defined DNA template strands in paired daughter cells in mouse colon analyzed by CO-FISH (see Supplementary file for full Legend). (MOV 1067 kb)
This movie file shows the distribution of defined DNA template strands in paired metaphase chromosomes in the mouse colon analyzed by CO-FISH (see Supplementary file for full Legend). (MOV 1692 kb)
This movie file shows the mirror asymmetry of defined DNA template strands in a pair of post-mitotic mouse colon cells (see Supplementary Information file for full Legend). (MOV 2248 kb)
This movie file shows mirror asymmetry of defined DNA template strands in a pair of post-mitotic mouse colon cells (see Supplementary Information file for full Legend). (MOV 1347 kb)
This file contains an excel spreadsheet which shows the relative fluorescence measurements extracted from two-colour digital CO-FISH images of cell pairs present in colon tissue sections, colon suspension cells, fibroblast and ES cells (see Supplementary Information file for full Legend). (XLS 133 kb)
This file contains an image gallery which shows cell pairs and surrounding tissue from statistically significant asymmetric segregation events (see Supplementary Information file for full Legend). (PDF 589 kb)
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Falconer, E., Chavez, E., Henderson, A. et al. Identification of sister chromatids by DNA template strand sequences. Nature 463, 93–97 (2010). https://doi.org/10.1038/nature08644
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