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The microtubule-dependent motor centromere-associated protein E (CENP-E) is an integral component of kinetochore corona fibers that link centromeres to spindle microtubules.

Yao X, Anderson KL, Cleveland DW - J. Cell Biol. (1997)

Bottom Line: Centromere-associated protein E (CENP-E) is a kinesin-related microtubule motor protein that is essential for chromosome congression during mitosis.In congressing chromosomes, CENP-E is preferentially associated with (or accessible at) the stretched, leading kinetochore known to provide the primary power for chromosome movement.Taken together, this evidence strongly supports a model in which CENP-E functions in congression to tether kinetochores to the disassembling microtubule plus ends.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Cell Biology, Ludwig Institute for Cancer Research, School of Medicine, University of California, La Jolla, CA 92093-0660, USA.

ABSTRACT
Centromere-associated protein E (CENP-E) is a kinesin-related microtubule motor protein that is essential for chromosome congression during mitosis. Using immunoelectron microscopy, CENP-E is shown to be an integral component of the kinetochore corona fibers that tether centromeres to the spindle. Immediately upon nuclear envelope fragmentation, an associated plus end motor trafficks cytoplasmic CENP-E toward chromosomes along astral microtubules that enter the nuclear volume. Before or concurrently with initial lateral attachment of spindle microtubules, CENP-E targets to the outermost region of the developing kinetochores. After stable attachment, throughout chromosome congression, at metaphase, and throughout anaphase A, CENP-E is a constituent of the corona fibers, extending at least 50 nm away from the kinetochore outer plate and intertwining with spindle microtubules. In congressing chromosomes, CENP-E is preferentially associated with (or accessible at) the stretched, leading kinetochore known to provide the primary power for chromosome movement. Taken together, this evidence strongly supports a model in which CENP-E functions in congression to tether kinetochores to the disassembling microtubule plus ends.

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The leading kinetochore of a congressing chromosomes pair has increased  level or accessibility of  CENP-E. HeLa cells were  processed as described in Fig.  2. (A and D) Low magnification views of a prometaphase  HeLa cell (poles of the bipolar spindle are labeled with  asterisks). (B and E) Intermediate magnification of two  examples of a bioriented chromosome. (B) Boxed area of  A showing chromosomes pair  partially congressed from spindle poles toward the equator  of spindle poles, but not yet  aligned at the equator. (C  and F) High magnification  of the two bioriented chromosomes in B and E. 10-nm  gold particles representing  CENP-E decorate the outer  kinetochore surface. A trilaminar structure of the kinetochore is not yet apparent,  indicating that these kinetochores are not fully mature.  (G–I) Double immunofluorescence demonstrating preferential CENP-E staining on  the kinetochore closest to the  midzone on a chromosome  not yet congressed to the metaphase plate. (G) CENP-B,  (H) CENP-E, and (I) DAPI  to display chromosome positioning. Bars: (A and D) 2 μm;  (B and E) 230 nm; (C) 90 nm;  (F) 110 nm; (H–I) 10 μm.
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Figure 4: The leading kinetochore of a congressing chromosomes pair has increased level or accessibility of CENP-E. HeLa cells were processed as described in Fig. 2. (A and D) Low magnification views of a prometaphase HeLa cell (poles of the bipolar spindle are labeled with asterisks). (B and E) Intermediate magnification of two examples of a bioriented chromosome. (B) Boxed area of A showing chromosomes pair partially congressed from spindle poles toward the equator of spindle poles, but not yet aligned at the equator. (C and F) High magnification of the two bioriented chromosomes in B and E. 10-nm gold particles representing CENP-E decorate the outer kinetochore surface. A trilaminar structure of the kinetochore is not yet apparent, indicating that these kinetochores are not fully mature. (G–I) Double immunofluorescence demonstrating preferential CENP-E staining on the kinetochore closest to the midzone on a chromosome not yet congressed to the metaphase plate. (G) CENP-B, (H) CENP-E, and (I) DAPI to display chromosome positioning. Bars: (A and D) 2 μm; (B and E) 230 nm; (C) 90 nm; (F) 110 nm; (H–I) 10 μm.

Mentions: To probe for the localization of CENP-E in chromosomes as they congress toward the spindle equator, we examined bioriented chromosomes in prometaphase cells. Two different examples from a single cell are highlighted in Fig. 4, A–C and D–F. At higher magnifications, nine gold particles representing specific labeling of CENP-E can be clearly seen on one leading kinetochore (i.e., defined here to be the one closer to the midzone; Fig. 4 C, arrow), while five particles are found on the other (trailing kinetochore; Fig. 4 C, arrowhead). Again, there was virtually no gold staining on other microtubules or at other surface regions of the chomosome (Fig. 4, B and E). In a second example (Fig. 4, D–F), seven gold particles were associated with the trailing sister kinetochore (Fig. 4 E), while the leading one reveals 14 particles, plus two adjacent clusters of CENP-E associated with a kinetochore microtubule. By examining 23 serial sections from five bioriented chromosomes, we determined that the leading kinetochore always displayed more intense CENP-E reactivity (12 ± 4 gold particles for the leading vs 7 ± 3 for the trailing), demonstrating a difference in abundance, conformation, or accessibility of kinetochore-bound CENP-E during chromosome congression. The increased immunoreactivity on the apparently leading kinetochore was confirmed at the light microscopic level. While CENP-B showed comparable staining on leading and trailing kinetochores of a lagging chromosome pair (Fig. 4 G, arrowhead and arrow, respectively), CENP-E staining was more intense on the kinetochore closest to the midzone (Fig. 4 H, arrowhead). Furthermore, in serial sections, none displayed the clear trilaminar structure seen at metaphase (e.g., see Fig. 5 B), demonstrating that kinetochore assembly is both multistep and incomplete even as late as bipolar microtubule attachment.


The microtubule-dependent motor centromere-associated protein E (CENP-E) is an integral component of kinetochore corona fibers that link centromeres to spindle microtubules.

Yao X, Anderson KL, Cleveland DW - J. Cell Biol. (1997)

The leading kinetochore of a congressing chromosomes pair has increased  level or accessibility of  CENP-E. HeLa cells were  processed as described in Fig.  2. (A and D) Low magnification views of a prometaphase  HeLa cell (poles of the bipolar spindle are labeled with  asterisks). (B and E) Intermediate magnification of two  examples of a bioriented chromosome. (B) Boxed area of  A showing chromosomes pair  partially congressed from spindle poles toward the equator  of spindle poles, but not yet  aligned at the equator. (C  and F) High magnification  of the two bioriented chromosomes in B and E. 10-nm  gold particles representing  CENP-E decorate the outer  kinetochore surface. A trilaminar structure of the kinetochore is not yet apparent,  indicating that these kinetochores are not fully mature.  (G–I) Double immunofluorescence demonstrating preferential CENP-E staining on  the kinetochore closest to the  midzone on a chromosome  not yet congressed to the metaphase plate. (G) CENP-B,  (H) CENP-E, and (I) DAPI  to display chromosome positioning. Bars: (A and D) 2 μm;  (B and E) 230 nm; (C) 90 nm;  (F) 110 nm; (H–I) 10 μm.
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Figure 4: The leading kinetochore of a congressing chromosomes pair has increased level or accessibility of CENP-E. HeLa cells were processed as described in Fig. 2. (A and D) Low magnification views of a prometaphase HeLa cell (poles of the bipolar spindle are labeled with asterisks). (B and E) Intermediate magnification of two examples of a bioriented chromosome. (B) Boxed area of A showing chromosomes pair partially congressed from spindle poles toward the equator of spindle poles, but not yet aligned at the equator. (C and F) High magnification of the two bioriented chromosomes in B and E. 10-nm gold particles representing CENP-E decorate the outer kinetochore surface. A trilaminar structure of the kinetochore is not yet apparent, indicating that these kinetochores are not fully mature. (G–I) Double immunofluorescence demonstrating preferential CENP-E staining on the kinetochore closest to the midzone on a chromosome not yet congressed to the metaphase plate. (G) CENP-B, (H) CENP-E, and (I) DAPI to display chromosome positioning. Bars: (A and D) 2 μm; (B and E) 230 nm; (C) 90 nm; (F) 110 nm; (H–I) 10 μm.
Mentions: To probe for the localization of CENP-E in chromosomes as they congress toward the spindle equator, we examined bioriented chromosomes in prometaphase cells. Two different examples from a single cell are highlighted in Fig. 4, A–C and D–F. At higher magnifications, nine gold particles representing specific labeling of CENP-E can be clearly seen on one leading kinetochore (i.e., defined here to be the one closer to the midzone; Fig. 4 C, arrow), while five particles are found on the other (trailing kinetochore; Fig. 4 C, arrowhead). Again, there was virtually no gold staining on other microtubules or at other surface regions of the chomosome (Fig. 4, B and E). In a second example (Fig. 4, D–F), seven gold particles were associated with the trailing sister kinetochore (Fig. 4 E), while the leading one reveals 14 particles, plus two adjacent clusters of CENP-E associated with a kinetochore microtubule. By examining 23 serial sections from five bioriented chromosomes, we determined that the leading kinetochore always displayed more intense CENP-E reactivity (12 ± 4 gold particles for the leading vs 7 ± 3 for the trailing), demonstrating a difference in abundance, conformation, or accessibility of kinetochore-bound CENP-E during chromosome congression. The increased immunoreactivity on the apparently leading kinetochore was confirmed at the light microscopic level. While CENP-B showed comparable staining on leading and trailing kinetochores of a lagging chromosome pair (Fig. 4 G, arrowhead and arrow, respectively), CENP-E staining was more intense on the kinetochore closest to the midzone (Fig. 4 H, arrowhead). Furthermore, in serial sections, none displayed the clear trilaminar structure seen at metaphase (e.g., see Fig. 5 B), demonstrating that kinetochore assembly is both multistep and incomplete even as late as bipolar microtubule attachment.

Bottom Line: Centromere-associated protein E (CENP-E) is a kinesin-related microtubule motor protein that is essential for chromosome congression during mitosis.In congressing chromosomes, CENP-E is preferentially associated with (or accessible at) the stretched, leading kinetochore known to provide the primary power for chromosome movement.Taken together, this evidence strongly supports a model in which CENP-E functions in congression to tether kinetochores to the disassembling microtubule plus ends.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Cell Biology, Ludwig Institute for Cancer Research, School of Medicine, University of California, La Jolla, CA 92093-0660, USA.

ABSTRACT
Centromere-associated protein E (CENP-E) is a kinesin-related microtubule motor protein that is essential for chromosome congression during mitosis. Using immunoelectron microscopy, CENP-E is shown to be an integral component of the kinetochore corona fibers that tether centromeres to the spindle. Immediately upon nuclear envelope fragmentation, an associated plus end motor trafficks cytoplasmic CENP-E toward chromosomes along astral microtubules that enter the nuclear volume. Before or concurrently with initial lateral attachment of spindle microtubules, CENP-E targets to the outermost region of the developing kinetochores. After stable attachment, throughout chromosome congression, at metaphase, and throughout anaphase A, CENP-E is a constituent of the corona fibers, extending at least 50 nm away from the kinetochore outer plate and intertwining with spindle microtubules. In congressing chromosomes, CENP-E is preferentially associated with (or accessible at) the stretched, leading kinetochore known to provide the primary power for chromosome movement. Taken together, this evidence strongly supports a model in which CENP-E functions in congression to tether kinetochores to the disassembling microtubule plus ends.

Show MeSH
Related in: MedlinePlus