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The dissection of meiotic chromosome movement in mice using an in vivo electroporation technique.

Shibuya H, Morimoto A, Watanabe Y - PLoS Genet. (2014)

Bottom Line: Further, during bouquet stage, telomeres are constrained near the MTOC, resulting in the transient suppression of telomere mobility and nuclear rotation.In contrast, actin regulates the oscillatory changes in nuclear shape.Our data provide the mechanical scheme for meiotic chromosome movement throughout prophase I in mammals.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Chromosome Dynamics, Institute of Molecular and Cellular Biosciences, University of Tokyo, Tokyo, Japan.

ABSTRACT
During meiosis, the rapid movement of telomeres along the nuclear envelope (NE) facilitates pairing/synapsis of homologous chromosomes. In mammals, the mechanical properties of chromosome movement and the cytoskeletal structures responsible for it remain poorly understood. Here, applying an in vivo electroporation (EP) technique in live mouse testis, we achieved the quick visualization of telomere, chromosome axis and microtubule organizing center (MTOC) movements. For the first time, we defined prophase sub-stages of live spermatocytes morphologically according to GFP-TRF1 and GFP-SCP3 signals. We show that rapid telomere movement and subsequent nuclear rotation persist from leptotene/zygotene to pachytene, and then decline in diplotene stage concomitant with the liberation of SUN1 from telomeres. Further, during bouquet stage, telomeres are constrained near the MTOC, resulting in the transient suppression of telomere mobility and nuclear rotation. MTs are responsible for these movements by forming cable-like structures on the NE, and, probably, by facilitating the rail-tacking movements of telomeres on the MT cables. In contrast, actin regulates the oscillatory changes in nuclear shape. Our data provide the mechanical scheme for meiotic chromosome movement throughout prophase I in mammals.

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Related in: MedlinePlus

Inter-centromere connection observed in live-diplotene cells.A, Diplotene spermatocytes (from 21 dpp male mouse testis) expressing GFP-TRF1 and GFP-SCP3 with the trajectories of pairs of telomeres shown by red and green (time 0:00–3:30 min), overlaid on a projection of the first (0:00) and last (3:30) time points. The arrows indicate the pericentromeric heterochromatin, on which the marked short-arm telomeres are co-localized. Magnified images show the original images of marked pairs of telomeres. Whole images are in S5D Figure and S5 Movie. B, Trajectories of pairs of telomeres shown in A at the indicated time points. The original and overlaid images are shown in S6 Figure. C, The interpretation of long-arm and short-arm telomere dynamics in diplotene stage (top). Quantification of the 3D-distances between homolog telomeres throughout the time-lapse analysis in the cell shown in A (bottom). 3 pairs of telomeres (shown as blue, red and green) were traced for 3.5 minutes (7 sec intervals) for both long-arm and short-arm telomeres. The inter-telomere distances were calculated from the original 3-dimensional data for each time-points. Whole images are in S5D Figure and S5 Movie. Bars, 5 µm (unless otherwise indicated).
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pgen-1004821-g004: Inter-centromere connection observed in live-diplotene cells.A, Diplotene spermatocytes (from 21 dpp male mouse testis) expressing GFP-TRF1 and GFP-SCP3 with the trajectories of pairs of telomeres shown by red and green (time 0:00–3:30 min), overlaid on a projection of the first (0:00) and last (3:30) time points. The arrows indicate the pericentromeric heterochromatin, on which the marked short-arm telomeres are co-localized. Magnified images show the original images of marked pairs of telomeres. Whole images are in S5D Figure and S5 Movie. B, Trajectories of pairs of telomeres shown in A at the indicated time points. The original and overlaid images are shown in S6 Figure. C, The interpretation of long-arm and short-arm telomere dynamics in diplotene stage (top). Quantification of the 3D-distances between homolog telomeres throughout the time-lapse analysis in the cell shown in A (bottom). 3 pairs of telomeres (shown as blue, red and green) were traced for 3.5 minutes (7 sec intervals) for both long-arm and short-arm telomeres. The inter-telomere distances were calculated from the original 3-dimensional data for each time-points. Whole images are in S5D Figure and S5 Movie. Bars, 5 µm (unless otherwise indicated).

Mentions: To dissect the different nature of short-arm and long-arm telomeres in diplotene stage, we traced a pair of GFP-TRF1 foci co-localizing with pericentromeric heterochromatin (at short-arm) and not co-localizing with pericentromeric heterochromatin (at long-arm) in the same diplotene cells (Fig. 4A). We found that these pairs of GFP-TRF1 foci move in coordination maintaining a distance within 4.5 µm, implying that these pairs of GFP-TRF1 foci correspond to telomeres of homologous chromosomes, which are physically connected by a chiasmata (Fig. 4A, B, C). The tracing of GFP-TRF1 foci revealed that a pair of heterochromatin-associated telomeres draws parallel trajectories while heterochromatin-unassociated telomeres draws rather disordered ones, suggesting that the heterochromatin-associated telomeres are more tightly connected and move in greater coordination than the heterochromatin-unassociated ones (Fig. 4A, B and original images in S5D and S6 Figures). The quantification of 3-dimensional distances between a pair of GFP-TRF1 foci further confirmed the stable and close association between heterochromatin-associated telomeres (<1 µm) and a rather loose association between heterochromatin-unassociated telomeres (<4.5 µm) (Fig. 4C). The tight association of telomeres on heterochromatin observed in our time-lapse imaging likely reflects the presence of the inter-centromere connection seen in fixed diplotene cells, which might facilitate the correct co-orientation of homolog centromeres and the subsequent disjunction of homologous chromosomes in anaphase I [34], [35].


The dissection of meiotic chromosome movement in mice using an in vivo electroporation technique.

Shibuya H, Morimoto A, Watanabe Y - PLoS Genet. (2014)

Inter-centromere connection observed in live-diplotene cells.A, Diplotene spermatocytes (from 21 dpp male mouse testis) expressing GFP-TRF1 and GFP-SCP3 with the trajectories of pairs of telomeres shown by red and green (time 0:00–3:30 min), overlaid on a projection of the first (0:00) and last (3:30) time points. The arrows indicate the pericentromeric heterochromatin, on which the marked short-arm telomeres are co-localized. Magnified images show the original images of marked pairs of telomeres. Whole images are in S5D Figure and S5 Movie. B, Trajectories of pairs of telomeres shown in A at the indicated time points. The original and overlaid images are shown in S6 Figure. C, The interpretation of long-arm and short-arm telomere dynamics in diplotene stage (top). Quantification of the 3D-distances between homolog telomeres throughout the time-lapse analysis in the cell shown in A (bottom). 3 pairs of telomeres (shown as blue, red and green) were traced for 3.5 minutes (7 sec intervals) for both long-arm and short-arm telomeres. The inter-telomere distances were calculated from the original 3-dimensional data for each time-points. Whole images are in S5D Figure and S5 Movie. Bars, 5 µm (unless otherwise indicated).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4263375&req=5

pgen-1004821-g004: Inter-centromere connection observed in live-diplotene cells.A, Diplotene spermatocytes (from 21 dpp male mouse testis) expressing GFP-TRF1 and GFP-SCP3 with the trajectories of pairs of telomeres shown by red and green (time 0:00–3:30 min), overlaid on a projection of the first (0:00) and last (3:30) time points. The arrows indicate the pericentromeric heterochromatin, on which the marked short-arm telomeres are co-localized. Magnified images show the original images of marked pairs of telomeres. Whole images are in S5D Figure and S5 Movie. B, Trajectories of pairs of telomeres shown in A at the indicated time points. The original and overlaid images are shown in S6 Figure. C, The interpretation of long-arm and short-arm telomere dynamics in diplotene stage (top). Quantification of the 3D-distances between homolog telomeres throughout the time-lapse analysis in the cell shown in A (bottom). 3 pairs of telomeres (shown as blue, red and green) were traced for 3.5 minutes (7 sec intervals) for both long-arm and short-arm telomeres. The inter-telomere distances were calculated from the original 3-dimensional data for each time-points. Whole images are in S5D Figure and S5 Movie. Bars, 5 µm (unless otherwise indicated).
Mentions: To dissect the different nature of short-arm and long-arm telomeres in diplotene stage, we traced a pair of GFP-TRF1 foci co-localizing with pericentromeric heterochromatin (at short-arm) and not co-localizing with pericentromeric heterochromatin (at long-arm) in the same diplotene cells (Fig. 4A). We found that these pairs of GFP-TRF1 foci move in coordination maintaining a distance within 4.5 µm, implying that these pairs of GFP-TRF1 foci correspond to telomeres of homologous chromosomes, which are physically connected by a chiasmata (Fig. 4A, B, C). The tracing of GFP-TRF1 foci revealed that a pair of heterochromatin-associated telomeres draws parallel trajectories while heterochromatin-unassociated telomeres draws rather disordered ones, suggesting that the heterochromatin-associated telomeres are more tightly connected and move in greater coordination than the heterochromatin-unassociated ones (Fig. 4A, B and original images in S5D and S6 Figures). The quantification of 3-dimensional distances between a pair of GFP-TRF1 foci further confirmed the stable and close association between heterochromatin-associated telomeres (<1 µm) and a rather loose association between heterochromatin-unassociated telomeres (<4.5 µm) (Fig. 4C). The tight association of telomeres on heterochromatin observed in our time-lapse imaging likely reflects the presence of the inter-centromere connection seen in fixed diplotene cells, which might facilitate the correct co-orientation of homolog centromeres and the subsequent disjunction of homologous chromosomes in anaphase I [34], [35].

Bottom Line: Further, during bouquet stage, telomeres are constrained near the MTOC, resulting in the transient suppression of telomere mobility and nuclear rotation.In contrast, actin regulates the oscillatory changes in nuclear shape.Our data provide the mechanical scheme for meiotic chromosome movement throughout prophase I in mammals.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Chromosome Dynamics, Institute of Molecular and Cellular Biosciences, University of Tokyo, Tokyo, Japan.

ABSTRACT
During meiosis, the rapid movement of telomeres along the nuclear envelope (NE) facilitates pairing/synapsis of homologous chromosomes. In mammals, the mechanical properties of chromosome movement and the cytoskeletal structures responsible for it remain poorly understood. Here, applying an in vivo electroporation (EP) technique in live mouse testis, we achieved the quick visualization of telomere, chromosome axis and microtubule organizing center (MTOC) movements. For the first time, we defined prophase sub-stages of live spermatocytes morphologically according to GFP-TRF1 and GFP-SCP3 signals. We show that rapid telomere movement and subsequent nuclear rotation persist from leptotene/zygotene to pachytene, and then decline in diplotene stage concomitant with the liberation of SUN1 from telomeres. Further, during bouquet stage, telomeres are constrained near the MTOC, resulting in the transient suppression of telomere mobility and nuclear rotation. MTs are responsible for these movements by forming cable-like structures on the NE, and, probably, by facilitating the rail-tacking movements of telomeres on the MT cables. In contrast, actin regulates the oscillatory changes in nuclear shape. Our data provide the mechanical scheme for meiotic chromosome movement throughout prophase I in mammals.

Show MeSH
Related in: MedlinePlus