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Telomeres cluster de novo before the initiation of synapsis: a three-dimensional spatial analysis of telomere positions before and during meiotic prophase.

Bass HW, Marshall WF, Sedat JW, Agard DA, Cande WZ - J. Cell Biol. (1997)

Bottom Line: We found that nuclei at the last somatic prophase before meiosis exhibit a nonrandom, polarized chromosome organization resulting in a loose grouping of telomeres.The stage-dependent changes in telomere arrangements are suggestive of specific, active telomere-associated motility processes with meiotic functions.Thus, the formation of the cluster itself is an early event in the nuclear reorganizations associated with meiosis and may reflect a control point in the initiation of synapsis or crossing over.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, University of California at Berkeley 94720, USA.

ABSTRACT
We have analyzed the progressive changes in the spatial distribution of telomeres during meiosis using three-dimensional, high resolution fluorescence microscopy. Fixed meiotic cells of maize (Zea mays L.) were subjected to in situ hybridization under conditions that preserved chromosome structure, allowing identification of stage-dependent changes in telomere arrangements. We found that nuclei at the last somatic prophase before meiosis exhibit a nonrandom, polarized chromosome organization resulting in a loose grouping of telomeres. Quantitative measurements on the spatial arrangements of telomeres revealed that, as cells passed through premeiotic interphase and into leptotene, there was an increase in the frequency of large telomere-to-telomere distances and a decrease in the bias toward peripheral localization of telomeres. By leptotene, there was no obvious evidence of telomere grouping, and the large, singular nucleolus was internally located, nearly concentric with the nucleus. At the end of leptotene, telomeres clustered de novo at the nuclear periphery, coincident with a displacement of the nucleolus to one side. The telomere cluster persisted throughout zygotene and into early pachytene. The nucleolus was adjacent to the cluster at zygotene. At the pachytene stage, telomeres rearranged again by dispersing throughout the nuclear periphery. The stage-dependent changes in telomere arrangements are suggestive of specific, active telomere-associated motility processes with meiotic functions. Thus, the formation of the cluster itself is an early event in the nuclear reorganizations associated with meiosis and may reflect a control point in the initiation of synapsis or crossing over.

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Polarized chromosomes and telomere grouping at the last prophase before meiosis. 3-dimensional telomere FISH was carried  out on acrylamide-embedded cells (see Materials and Methods) from 0.2-mm anthers. Two nuclei at the last prophase before meiosis are  shown (A–D, and E–F). To convey the 3-dimensional data in 2-dimensional images, the original data sets (60–80 optical sections each)  were converted to a series of four sequential projections each with an effective focal planes of 1/4 the nuclear depth (∼5 microns). The  four 1/4 nucleus projections are displayed in sequence to give visual access to the full complement of data in the original data set (A).  Pseudocolor overlays showing the DNA (DAPI image, red) and the telomeres (bright dots in the FITC images, green/yellow) for each  projection show a loose grouping telomere signals (green dots). (B) A 3-dimensional model of the nucleus in A is shown. The edges of  the nucleus (purple wire) and the nucleolus (red wire) are indicated along with the positions of the telomeres (yellow spheres). (C) Volume-rendered projections of the entire nucleus is also shown to convey the overall paths of the chromosomes. The telomere hemisphere  (T*) and the presumed centromere hemisphere (C*) are indicated. (D) The axial paths of continuous chromosome were determined  and they are individually displayed side by side. (E) A volume-rendered projection of a different nucleus is shown along with the paths  of the 12 longest chromosome tracings (F). The tracings in F are rotated clockwise about 10° relative to the projection in E. Each nucleus has a large, singular internal nucleolus (not shown for nucleus in E). Bar, 5 μm.
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Figure 3: Polarized chromosomes and telomere grouping at the last prophase before meiosis. 3-dimensional telomere FISH was carried out on acrylamide-embedded cells (see Materials and Methods) from 0.2-mm anthers. Two nuclei at the last prophase before meiosis are shown (A–D, and E–F). To convey the 3-dimensional data in 2-dimensional images, the original data sets (60–80 optical sections each) were converted to a series of four sequential projections each with an effective focal planes of 1/4 the nuclear depth (∼5 microns). The four 1/4 nucleus projections are displayed in sequence to give visual access to the full complement of data in the original data set (A). Pseudocolor overlays showing the DNA (DAPI image, red) and the telomeres (bright dots in the FITC images, green/yellow) for each projection show a loose grouping telomere signals (green dots). (B) A 3-dimensional model of the nucleus in A is shown. The edges of the nucleus (purple wire) and the nucleolus (red wire) are indicated along with the positions of the telomeres (yellow spheres). (C) Volume-rendered projections of the entire nucleus is also shown to convey the overall paths of the chromosomes. The telomere hemisphere (T*) and the presumed centromere hemisphere (C*) are indicated. (D) The axial paths of continuous chromosome were determined and they are individually displayed side by side. (E) A volume-rendered projection of a different nucleus is shown along with the paths of the 12 longest chromosome tracings (F). The tracings in F are rotated clockwise about 10° relative to the projection in E. Each nucleus has a large, singular internal nucleolus (not shown for nucleus in E). Bar, 5 μm.

Mentions: All images were recorded using an IMT-2 wide-field microscope (Olympus Corp., Lake Success, NY), making use of one of two oil immersion lenses, ×60 NA 1.4 PlanApo (Olympus Corp.) or ×100 NA 1.4 PlanApo (Nikon Inc., Garden City, NY) (30). In both cases, the data were oversampled in the X, Y, and Z dimensions with typical XYZ voxel dimensions of 0.07 × 0.07 × 0.2 μm3. The computerized light microscope workstation has been described elsewhere (14, 15). Original data collection was made on an area extending at least 2 μm beyond the nuclear border in Z, and extending at least 5 μm in X and Y from the widest edges of the nuclei. After three-dimensional iterative deconvolution (7) of the original fullsized data sets, individual nuclei were computationally cropped in all dimensions. The resulting data subsets were used for all subsequent model building and image display, resulting in the exclusion of the surrounding cytoplasm and neighboring cells from the figures presented. The images presented were adjusted for brightness and contrast using linear scaling of the minimum and maximum intensities, but no additional image processing such as edge or local contrast enhancements were used, except for the volume-rendered projections in Fig. 3, C and E.


Telomeres cluster de novo before the initiation of synapsis: a three-dimensional spatial analysis of telomere positions before and during meiotic prophase.

Bass HW, Marshall WF, Sedat JW, Agard DA, Cande WZ - J. Cell Biol. (1997)

Polarized chromosomes and telomere grouping at the last prophase before meiosis. 3-dimensional telomere FISH was carried  out on acrylamide-embedded cells (see Materials and Methods) from 0.2-mm anthers. Two nuclei at the last prophase before meiosis are  shown (A–D, and E–F). To convey the 3-dimensional data in 2-dimensional images, the original data sets (60–80 optical sections each)  were converted to a series of four sequential projections each with an effective focal planes of 1/4 the nuclear depth (∼5 microns). The  four 1/4 nucleus projections are displayed in sequence to give visual access to the full complement of data in the original data set (A).  Pseudocolor overlays showing the DNA (DAPI image, red) and the telomeres (bright dots in the FITC images, green/yellow) for each  projection show a loose grouping telomere signals (green dots). (B) A 3-dimensional model of the nucleus in A is shown. The edges of  the nucleus (purple wire) and the nucleolus (red wire) are indicated along with the positions of the telomeres (yellow spheres). (C) Volume-rendered projections of the entire nucleus is also shown to convey the overall paths of the chromosomes. The telomere hemisphere  (T*) and the presumed centromere hemisphere (C*) are indicated. (D) The axial paths of continuous chromosome were determined  and they are individually displayed side by side. (E) A volume-rendered projection of a different nucleus is shown along with the paths  of the 12 longest chromosome tracings (F). The tracings in F are rotated clockwise about 10° relative to the projection in E. Each nucleus has a large, singular internal nucleolus (not shown for nucleus in E). Bar, 5 μm.
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Related In: Results  -  Collection

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Figure 3: Polarized chromosomes and telomere grouping at the last prophase before meiosis. 3-dimensional telomere FISH was carried out on acrylamide-embedded cells (see Materials and Methods) from 0.2-mm anthers. Two nuclei at the last prophase before meiosis are shown (A–D, and E–F). To convey the 3-dimensional data in 2-dimensional images, the original data sets (60–80 optical sections each) were converted to a series of four sequential projections each with an effective focal planes of 1/4 the nuclear depth (∼5 microns). The four 1/4 nucleus projections are displayed in sequence to give visual access to the full complement of data in the original data set (A). Pseudocolor overlays showing the DNA (DAPI image, red) and the telomeres (bright dots in the FITC images, green/yellow) for each projection show a loose grouping telomere signals (green dots). (B) A 3-dimensional model of the nucleus in A is shown. The edges of the nucleus (purple wire) and the nucleolus (red wire) are indicated along with the positions of the telomeres (yellow spheres). (C) Volume-rendered projections of the entire nucleus is also shown to convey the overall paths of the chromosomes. The telomere hemisphere (T*) and the presumed centromere hemisphere (C*) are indicated. (D) The axial paths of continuous chromosome were determined and they are individually displayed side by side. (E) A volume-rendered projection of a different nucleus is shown along with the paths of the 12 longest chromosome tracings (F). The tracings in F are rotated clockwise about 10° relative to the projection in E. Each nucleus has a large, singular internal nucleolus (not shown for nucleus in E). Bar, 5 μm.
Mentions: All images were recorded using an IMT-2 wide-field microscope (Olympus Corp., Lake Success, NY), making use of one of two oil immersion lenses, ×60 NA 1.4 PlanApo (Olympus Corp.) or ×100 NA 1.4 PlanApo (Nikon Inc., Garden City, NY) (30). In both cases, the data were oversampled in the X, Y, and Z dimensions with typical XYZ voxel dimensions of 0.07 × 0.07 × 0.2 μm3. The computerized light microscope workstation has been described elsewhere (14, 15). Original data collection was made on an area extending at least 2 μm beyond the nuclear border in Z, and extending at least 5 μm in X and Y from the widest edges of the nuclei. After three-dimensional iterative deconvolution (7) of the original fullsized data sets, individual nuclei were computationally cropped in all dimensions. The resulting data subsets were used for all subsequent model building and image display, resulting in the exclusion of the surrounding cytoplasm and neighboring cells from the figures presented. The images presented were adjusted for brightness and contrast using linear scaling of the minimum and maximum intensities, but no additional image processing such as edge or local contrast enhancements were used, except for the volume-rendered projections in Fig. 3, C and E.

Bottom Line: We found that nuclei at the last somatic prophase before meiosis exhibit a nonrandom, polarized chromosome organization resulting in a loose grouping of telomeres.The stage-dependent changes in telomere arrangements are suggestive of specific, active telomere-associated motility processes with meiotic functions.Thus, the formation of the cluster itself is an early event in the nuclear reorganizations associated with meiosis and may reflect a control point in the initiation of synapsis or crossing over.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, University of California at Berkeley 94720, USA.

ABSTRACT
We have analyzed the progressive changes in the spatial distribution of telomeres during meiosis using three-dimensional, high resolution fluorescence microscopy. Fixed meiotic cells of maize (Zea mays L.) were subjected to in situ hybridization under conditions that preserved chromosome structure, allowing identification of stage-dependent changes in telomere arrangements. We found that nuclei at the last somatic prophase before meiosis exhibit a nonrandom, polarized chromosome organization resulting in a loose grouping of telomeres. Quantitative measurements on the spatial arrangements of telomeres revealed that, as cells passed through premeiotic interphase and into leptotene, there was an increase in the frequency of large telomere-to-telomere distances and a decrease in the bias toward peripheral localization of telomeres. By leptotene, there was no obvious evidence of telomere grouping, and the large, singular nucleolus was internally located, nearly concentric with the nucleus. At the end of leptotene, telomeres clustered de novo at the nuclear periphery, coincident with a displacement of the nucleolus to one side. The telomere cluster persisted throughout zygotene and into early pachytene. The nucleolus was adjacent to the cluster at zygotene. At the pachytene stage, telomeres rearranged again by dispersing throughout the nuclear periphery. The stage-dependent changes in telomere arrangements are suggestive of specific, active telomere-associated motility processes with meiotic functions. Thus, the formation of the cluster itself is an early event in the nuclear reorganizations associated with meiosis and may reflect a control point in the initiation of synapsis or crossing over.

Show MeSH