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Lateral and End-On Kinetochore Attachments Are Coordinated to Achieve Bi-orientation in Drosophila Oocytes.

Radford SJ, Hoang TL, Głuszek AA, Ohkura H, McKim KS - PLoS Genet. (2015)

Bottom Line: We found that the initiation of spindle assembly results from chromosome-microtubule interactions that are kinetochore-independent.Stabilization of the spindle, however, depends on both central spindle and kinetochore components.We propose that the bi-orientation process begins with the kinetochores moving laterally along central spindle microtubules towards their minus ends.

View Article: PubMed Central - PubMed

Affiliation: Waksman Institute of Microbiology, Rutgers, The State University of New Jersey, Piscataway, New Jersey, United States of America.

ABSTRACT
In oocytes, where centrosomes are absent, the chromosomes direct the assembly of a bipolar spindle. Interactions between chromosomes and microtubules are essential for both spindle formation and chromosome segregation, but the nature and function of these interactions is not clear. We have examined oocytes lacking two kinetochore proteins, NDC80 and SPC105R, and a centromere-associated motor protein, CENP-E, to characterize the impact of kinetochore-microtubule attachments on spindle assembly and chromosome segregation in Drosophila oocytes. We found that the initiation of spindle assembly results from chromosome-microtubule interactions that are kinetochore-independent. Stabilization of the spindle, however, depends on both central spindle and kinetochore components. This stabilization coincides with changes in kinetochore-microtubule attachments and bi-orientation of homologs. We propose that the bi-orientation process begins with the kinetochores moving laterally along central spindle microtubules towards their minus ends. This movement depends on SPC105R, can occur in the absence of NDC80, and is antagonized by plus-end directed forces from the CENP-E motor. End-on kinetochore-microtubule attachments that depend on NDC80 are required to stabilize bi-orientation of homologs. A surprising finding was that SPC105R but not NDC80 is required for co-orientation of sister centromeres at meiosis I. Together, these results demonstrate that, in oocytes, kinetochore-dependent and -independent chromosome-microtubule attachments work together to promote the accurate segregation of chromosomes.

No MeSH data available.


Related in: MedlinePlus

Loss of CENP-E disrupts chromosome alignment in oocytes.Confocal images of karyosome and spindle organization in oocytes from wild type, cana/Df, after knockdown of cmet, Cenp-E germline clones, and after knockdown of Ndc80, Spc105R (HMS01548), Ndc80 and cmet, or Spc105R (HMS01548) and cmet. DNA is in blue and tubulin is in green in merged images. Single channel images show DNA in white. Scale bars represent 10 μm.
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pgen.1005605.g005: Loss of CENP-E disrupts chromosome alignment in oocytes.Confocal images of karyosome and spindle organization in oocytes from wild type, cana/Df, after knockdown of cmet, Cenp-E germline clones, and after knockdown of Ndc80, Spc105R (HMS01548), Ndc80 and cmet, or Spc105R (HMS01548) and cmet. DNA is in blue and tubulin is in green in merged images. Single channel images show DNA in white. Scale bars represent 10 μm.

Mentions: In cana mutant, cmet-depleted, or Cenp-E mutant oocytes, bipolar spindles formed (Fig 5). However, in cmet-depleted or Cenp-E mutant oocytes, the karyosome frequently split into multiple masses (Fig 5 and Table 1 and S3 Table). This karyosome defect was more frequent in Cenp-E mutant oocytes than in cmet-depleted oocytes (43% vs 15%, P = 0.001), demonstrating that both CENP-E homologs are required for proper karyosome organization. CANA and CMET are partially redundant because CMET is necessary for karyosome organization even when there is a functional copy of CANA. These results are the first evidence that the second Drosophila CENP-E homolog CANA is functional. Additionally, these results suggest that, although traditional chromosome congression does not occur in Drosophila oocytes, CENP-E is required to prevent chromosomes from becoming un-aligned and separated from the main karyosome mass.


Lateral and End-On Kinetochore Attachments Are Coordinated to Achieve Bi-orientation in Drosophila Oocytes.

Radford SJ, Hoang TL, Głuszek AA, Ohkura H, McKim KS - PLoS Genet. (2015)

Loss of CENP-E disrupts chromosome alignment in oocytes.Confocal images of karyosome and spindle organization in oocytes from wild type, cana/Df, after knockdown of cmet, Cenp-E germline clones, and after knockdown of Ndc80, Spc105R (HMS01548), Ndc80 and cmet, or Spc105R (HMS01548) and cmet. DNA is in blue and tubulin is in green in merged images. Single channel images show DNA in white. Scale bars represent 10 μm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4608789&req=5

pgen.1005605.g005: Loss of CENP-E disrupts chromosome alignment in oocytes.Confocal images of karyosome and spindle organization in oocytes from wild type, cana/Df, after knockdown of cmet, Cenp-E germline clones, and after knockdown of Ndc80, Spc105R (HMS01548), Ndc80 and cmet, or Spc105R (HMS01548) and cmet. DNA is in blue and tubulin is in green in merged images. Single channel images show DNA in white. Scale bars represent 10 μm.
Mentions: In cana mutant, cmet-depleted, or Cenp-E mutant oocytes, bipolar spindles formed (Fig 5). However, in cmet-depleted or Cenp-E mutant oocytes, the karyosome frequently split into multiple masses (Fig 5 and Table 1 and S3 Table). This karyosome defect was more frequent in Cenp-E mutant oocytes than in cmet-depleted oocytes (43% vs 15%, P = 0.001), demonstrating that both CENP-E homologs are required for proper karyosome organization. CANA and CMET are partially redundant because CMET is necessary for karyosome organization even when there is a functional copy of CANA. These results are the first evidence that the second Drosophila CENP-E homolog CANA is functional. Additionally, these results suggest that, although traditional chromosome congression does not occur in Drosophila oocytes, CENP-E is required to prevent chromosomes from becoming un-aligned and separated from the main karyosome mass.

Bottom Line: We found that the initiation of spindle assembly results from chromosome-microtubule interactions that are kinetochore-independent.Stabilization of the spindle, however, depends on both central spindle and kinetochore components.We propose that the bi-orientation process begins with the kinetochores moving laterally along central spindle microtubules towards their minus ends.

View Article: PubMed Central - PubMed

Affiliation: Waksman Institute of Microbiology, Rutgers, The State University of New Jersey, Piscataway, New Jersey, United States of America.

ABSTRACT
In oocytes, where centrosomes are absent, the chromosomes direct the assembly of a bipolar spindle. Interactions between chromosomes and microtubules are essential for both spindle formation and chromosome segregation, but the nature and function of these interactions is not clear. We have examined oocytes lacking two kinetochore proteins, NDC80 and SPC105R, and a centromere-associated motor protein, CENP-E, to characterize the impact of kinetochore-microtubule attachments on spindle assembly and chromosome segregation in Drosophila oocytes. We found that the initiation of spindle assembly results from chromosome-microtubule interactions that are kinetochore-independent. Stabilization of the spindle, however, depends on both central spindle and kinetochore components. This stabilization coincides with changes in kinetochore-microtubule attachments and bi-orientation of homologs. We propose that the bi-orientation process begins with the kinetochores moving laterally along central spindle microtubules towards their minus ends. This movement depends on SPC105R, can occur in the absence of NDC80, and is antagonized by plus-end directed forces from the CENP-E motor. End-on kinetochore-microtubule attachments that depend on NDC80 are required to stabilize bi-orientation of homologs. A surprising finding was that SPC105R but not NDC80 is required for co-orientation of sister centromeres at meiosis I. Together, these results demonstrate that, in oocytes, kinetochore-dependent and -independent chromosome-microtubule attachments work together to promote the accurate segregation of chromosomes.

No MeSH data available.


Related in: MedlinePlus