Research ArticleBIOPHYSICS

Three-dimensional positioning and structure of chromosomes in a human prophase nucleus

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Science Advances  21 Jul 2017:
Vol. 3, no. 7, e1602231
DOI: 10.1126/sciadv.1602231
  • Fig. 1 Two slices from the acquired SBFSEM stack of 300 BSE micrographs of the nucleus with zoomed-in insets of two selected chromosomes.

    (A) Slice no. 36 of 300. (B) Slice no. 127 of 300. The insets in (A) and (B) are a broken chromosome and an identified intact chromosome A1 (assigned chromosome 1; see explanation in the text), respectively. The pixel size of the BSE micrographs is 11 nm × 11 nm, and the sectioning thickness is 20 nm.

  • Fig. 2 3D characterization and analysis of the imaged chromosomes within the human prophase nucleus.

    (A) 3D rendering of all the observed chromosomes in the nucleus, with its envelope in transparent blue, the intact chromosomes in yellow or green, and the broken ones in red. (B) 3D rendering of the intact chromosomes only in the nucleus. (C) Measured chromatid volumes versus the number of base pairs of the accordingly assigned chromosomes from the human genome sequence, with a linear fit. (D) Correlation between the measured chromatid volumes and the radii of the assigned chromosomes from the center of the nucleus. (E) Correlation between the gene density of measured chromosomes and their radii from the center of the nucleus. The chromosomes have been labeled by their assigned chromosome numbers in (E).

  • Fig. 3 Rendering of the identified intact individual chromosomes.

    (A) An SBFSEM slice of the chromosome A1, assigned as chromosome 1. This is a different slice from the inset in Fig. 1B. (B) 3D rendering of the chromatid separated chromosome A1 showing cavities in solid light green, green, and blue. (C) 3D rendering of the cavity network in chromosome A1 viewed through a transparent chromosome surface. Bottom: 3D rendering of the surface views of the chromatid-separated chromosomes A2 (D), A4 (E), and BC1 (F), which are assigned as chromosomes 2, 3, and 6, respectively. The insets at the bottom-right corners of (D), (E), and (F) are the SBFSEM slices of the chromosomes A2, A4, and BC1, respectively. Scale bars, 500 nm [(B to F), in each direction] and 1 μm (insets in D to F).

  • Fig. 4 Characterization and analysis of chromatid pairs of selected chromosomes.

    Left, middle, and right columns show chromosomes A3, BC3, and D2, respectively, which are assigned as chromosomes 3, 7, or X and 16 accordingly to the scheme of Table 1. (A to C) 3D rendering of the chromatid-separated chromosomes. (D to F) 3D superposition of the chromatid pairs through volume registration using Avizo. Scale bars, 500 nm (in each direction).

  • Table 1 Volume statistics of all the intact chromosomes in the nucleus.
    Chromosome
    (number)
    Chromatid volume (V1)
    (μm3)*
    Cavity volume (V2)
    (μm3)
    Ratio of V1/(V1 + V2)
    (%)
    Assigned chromosome
    (number)
    Calculated chromatid volume
    (μm3)
    A11.6550.08795.011.446
    A21.6500.04897.221.411
    A31.3280.04796.631.148
    A41.2630.03797.131.148
    BC11.1240.08393.151.049
    BC21.0920.07693.560.992
    BC31.0560.03696.77 (or X)0.923
    C10.9930.10290.780.849
    C20.9720.05394.890.819
    C30.9630.07492.9100.786
    C40.9550.05494.7110.783
    C50.8950.04994.8120.777
    D10.7760.05493.5130.668
    D20.6250.06690.4160.524
    D30.6030.03794.2170.471
    D40.5720.04592.7180.453
    D50.5090.02994.6190.343
    D60.4350.03592.6220.298
    D70.2130.01493.8210.279

    *Chromatid volume is half of the measured volume of the chromosomes without cavities, that is, half of the volume of the black region only in the imaged chromosomes, with chromatid pairs measured by Avizo.

    †Cavity volume is half of the volume of all the cavities in the chromosomes with chromatid pairs.

    ‡Calculated chromatid volumes were obtained by multiplying the sequence length of the assigned human chromosomes (in mega–base pairs) from the database by 5.80 nm3. The database is Archive Ensembl Release 68, July 2012 (28). The volume per base pair, 5.80 nm3, was obtained by theoretical calculation, which is explained in the main text.

    • Table 2 Statistical analysis of the chromatid pairs of selected intact chromosomes.

      SSC1, single sister chromatid 1; SSC2, single sister chromatid 2; Ave., average value of SSC1 and SSC2.

      Chromosome
      (number)
      Chromatid
      length (μm)*
      Ave. of cross-sectional
      area (μm2)
      Ave. diameter of cross
      sections (μm)
      Difference of the chromatid
      pairs in volume (%)
      A1SSC13.2410.5240.8191.84
      SSC22.8010.530
      Ave.3.0210.527
      A2SSC12.8690.5120.7996.54
      SSC23.0070.492
      Ave.2.9380.502
      A3SSC12.8280.4380.7603.50
      SSC22.7440.470
      Ave.2.7860.454
      A4SSC12.7660.4210.7420.31
      SSC22.7680.443
      Ave.2.7670.432
      BC1SSC12.2360.4920.7769.00
      SSC22.5310.453
      Ave.2.3840.473
      BC3SSC12.0850.4390.7604.61
      SSC21.9140.468
      Ave.2.0000.454
      D2SSC11.7320.3750.7019.13
      SSC21.6470.396
      Ave.1.6900.386

      *Chromatid lengths were obtained by measuring the separated two chromatids of the chromosomes using Avizo.

      †“Ave. of cross-sectional area” represents the average of the areas of a few cross sections along the central axis of the chromatids, which were carried out using Avizo. “Ave. diameter of cross sections” represents the diameter calculated from the values of the average of the “Ave. of cross-sectional area” when we assume the cross sections are circles.

      ‡Difference was obtained by superposition calculation on the two chromatids by 3D volume registration using Avizo. The difference (in percentage) equals 2*abs(VC1VC2)/(VC1 + VC2)*100%, where VC1 is the volume of the SSC1 and VC2 is the volume of the SSC2.

      Supplementary Materials

      • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/3/7/e1602231/DC1

        movie S1. 3D rendering of the measured prophase nucleus.

        movie S2. 3D rendering of the measured prophase nucleus from another orientation.

        table S1. Volume statistics of all the broken chromosomes in the nucleus.

        table S2. Reassignment of all the intact chromosomes as a comparison with the assignment in the main text.

        fig. S1. SBFSEM slices of the measured (smaller) chromosomes.

        fig. S2. Second linear fitting of the chromatid volumes against their base pair numbers.

      • Supplementary Materials

        This PDF file includes:

        • Legends for movies S1 and S2
        • table S1. Volume statistics of all the broken chromosomes in the nucleus.
        • table S2. Reassignment of all the intact chromosomes as a comparison with the assignment in the main text.
        • fig. S1. SBFSEM slices of the measured (smaller) chromosomes.
        • fig. S2. Second linear fitting of the chromatid volumes against their base pair numbers.

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        Other Supplementary Material for this manuscript includes the following:

        • movie S1 (.mpg format). 3D rendering of the measured prophase nucleus.
        • movie S2 (.mpg format). 3D rendering of the measured prophase nucleus from another orientation.

        Files in this Data Supplement:

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