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Transcription-dependent radial distribution of TCF7L2 regulated genes in chromosome territories

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Abstract

Human chromosomes occupy distinct territories in the interphase nucleus. Such chromosome territories (CTs) are positioned according to gene density. Gene-rich CTs are generally located in the center of the nucleus, while gene-poor CTs are positioned more towards the nuclear periphery. However, the association between gene expression levels and the radial positioning of genes within the CT is still under debate. In the present study, we performed three-dimensional fluorescence in situ hybridization experiments in the colorectal cancer cell lines DLD-1 and LoVo using whole chromosome painting probes for chromosomes 8 and 11 and BAC clones targeting four genes with different expression levels assessed by gene expression arrays and RT-PCR. Our results confirmed that the two over-expressed genes, MYC on chromosome 8 and CCND1 on chromosome 11, are located significantly further away from the center of the CT compared to under-expressed genes on the same chromosomes, i.e., DLC1 and SCN3B. When CCND1 expression was reduced after silencing the major transcription factor of the WNT/β-catenin signaling pathway, TCF7L2, the gene was repositioned and mostly detected in the interior of the CT. Thus, we suggest a non-random distribution in which over-expressed genes are located more towards the periphery of the respective CTs.

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Acknowledgements

The authors would like to thank Dr. Subhadra Banerjee from the Flow Cytometry Core Facility at NCI.

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Authors and Affiliations

  1. Unitat de Biologia Cel·lular i Genètica Mèdica, Departament de Biologia Cel·lular, Fisiologia i Immunologia, Facultat de Medicina, Universitat Autònoma de Barcelona, 08193, Bellaterra, Catalonia, Spain

    Keyvan Torabi, Immaculada Ponsa, Rosa Miró & Jordi Camps

  2. Gastrointestinal and Pancreatic Oncology Group, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Rosselló 153, 4th floor, 08036, Barcelona, Catalonia, Spain

    Keyvan Torabi, Maria Vila-Casadesús, Antoni Castells & Jordi Camps

  3. Genetics Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, 20892-8010, USA

    Darawalee Wangsa, Markus Brown & Thomas Ried

  4. Institut de Biotecnologia i Biomedicina, Universitat Autònoma de Barcelona, 08193, Bellaterra, Catalonia, Spain

    Immaculada Ponsa

  5. Advanced Optical Microscopy Unit, Scientific and Technologic Centers, Facultat de Medicina, Universitat de Barcelona, 08036, Barcelona, Catalonia, Spain

    Anna Bosch & Maria Calvo

  6. CCR/LRBGE Optical Microscopy Core, National Cancer Institute, National Institutes of Health, Bethesda, MD, 20892, USA

    Tatiana S. Karpova & Rosa Miró

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  1. Keyvan Torabi
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  2. Darawalee Wangsa
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  8. Maria Calvo
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Correspondence to Thomas Ried or Jordi Camps.

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Funding

This work was supported by grants from the European Commission (COLONGEVA to J.C.), the Instituto de Salud Carlos III and co-funded by the European Regional Development Fund (ERDF) (CP13/00160 to J.C.), the CIBEREHD program, the Agència de Gestió d’Ajuts Universitaris i de Recerca, Generalitat de Catalunya (2014 SGR 135 and 2014 SGR 903), and from the Intramural Research Program of National Institutes of Health/NCI. J.C. has received a travel grant from the Instituto de Salud Carlos III and was co-funded by the European Regional Development Fund (ERDF) to perform experiments at NIH (MV15/00026). K.T. received a PIF fellowship and a mobility grant from the Universitat Autònoma de Barcelona (456-01-02/2013).

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The authors declare that they have no conflict of interest.

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This article does not contain any studies with human participants or animals performed by any of the authors.

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Keyvan Torabi, Darawalee Wangsa, and Immaculada Ponsa are co-first authors.

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Table S1

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Table S2

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Fig. S1

Nuclear radial positioning of chromosome territories. Chromosome territory nuclear radial distribution profiles of a G1-phase and S-phase DLD-1 cells; b G1-phase and S-phase LoVo cells. The Mann-Whitney sum-rank test was used in order to compare CT median radial positioning distributions (n > 50). (GIF 26 kb)

High resolution image (TIFF 29636 kb)

Fig. S2

Gene expression patterns. Gene expression patterns of a,b G-1 phase DLD-1 and LoVo cells and c,d S-phase DLD-1 and LoVo cells. All cases showed an over-expression of MYC and CCND1 compared to DLC1 and SCN3B. Values were normalized by expression levels of the housekeeping gene HPRT1. (GIF 26 kb)

High resolution image (TIFF 35038 kb)

Fig. S3

Positioning of differentially expressed genes within their corresponding CTs in LoVo cells. Boxplots showing radial distances (i.e., eccentricity values) of MYC vs DLC1 and CCND1 vs SCN3B in a G1-phase cells, and b S-phase cells. Histograms and distributions of the distances between the center of the gene and the closest CT border depicted in bins of 0.2 μm are plotted for c G1- and d S-phase cells. Dashed-line at 0 indicates the border of the CT, thus positive values indicate that co-localization of the gene signal and the CT painting probe was lower than 50%. In contrast, negative values indicate that co-localization of the gene signal and the CT painting probe was higher than 50%. Adjusted distributions computed using Kernel density estimator method have been added to the plots. Indicated by arrowheads in the representative examples are loci looping out of the corresponding CT in e G1- and f S-phase. Scale bars correspond to 5 μm. The Wilcoxon test was used to compare median radial measurements between over- and under-expressed genes (n > 50). * corresponds to p < 0.05; ** correspond to p < 0.01, **** correspond to p < 0.0001 and n.s., non significant. (GIF 156 kb)

High resolution image (TIFF 70752 kb)

Fig. S4

Co-localization profiles between gene probes and CTs. Representative maximum projections of pseudoconfocal images stacks of S-phase DLD-1 cells and fluorescence intensity profiles corresponding to the line drawn over the CTs from the previous images of a CT8, MYC and DLC1 FISH panel and b CT11, CCND1 and SCN3B FISH panel. Scale bar corresponds to 5 μm. (GIF 93 kb)

High resolution image (TIFF 30193 kb)

Fig S5

Radial positioning of MYC in CT8 after TCF7L2 silencing. a Boxplot showing MYC radial distances in siTCF7L2 treated cells compared to siNEG and untreated cultures. b Histogram showing the distribution of the distance between MYC and the closest border of CT8 depicted in bins of 0.2 μm of siTCF7L2, siNEG and untreated cells. Adjusted distributions computed using Kernel density estimator method have been added to the plots. Mann-Whitney sum-rank test was used to compare MYC median radial distances between different experiments (untreated, siNEG and siTCF7L2). n.s., non significant. (GIF 29 kb)

High resolution image (TIFF 16500 kb)

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Torabi, K., Wangsa, D., Ponsa, I. et al. Transcription-dependent radial distribution of TCF7L2 regulated genes in chromosome territories. Chromosoma 126, 655–667 (2017). https://doi.org/10.1007/s00412-017-0629-6

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