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Three-dimensional localization of CENP-A suggests a complex higher order structure of centromeric chromatin.

Marshall OJ, Marshall AT, Choo KH - J. Cell Biol. (2008)

Bottom Line: Combined with previous chromatin immunoprecipitation results (Saffery, R., H.Choo. 2005.Genet. 14:85-93), our data suggest that centromeric chromatin is arranged in a coiled 30-nm fiber that is itself coiled or folded to form a higher order structure.

View Article: PubMed Central - PubMed

Affiliation: Chromosome and Chromatin Research, Murdoch Childrens Research Institute, Royal Children's Hospital, Parkville, Victoria 3052, Australia.

ABSTRACT
The histone H3 variant centromere protein A (CENP-A) is central to centromere formation throughout eukaryotes. A long-standing question in centromere biology has been the organization of CENP-A at the centromere and its implications for the structure of centromeric chromatin. In this study, we describe the three-dimensional localization of CENP-A at the inner kinetochore plate through serial-section transmission electron microscopy of human mitotic chromosomes. At the kinetochores of normal centromeres and at a neocentromere, CENP-A occupies a compact domain at the inner kinetochore plate, stretching across two thirds of the length of the constriction but encompassing only one third of the constriction width and height. Within this domain, evidence of substructure is apparent. Combined with previous chromatin immunoprecipitation results (Saffery, R., H. Sumer, S. Hassan, L.H. Wong, J.M. Craig, K. Todokoro, M. Anderson, A. Stafford, and K.H.A. Choo. 2003. Mol. Cell. 12:509-516; Chueh, A.C., L.H. Wong, N. Wong, and K.H.A. Choo. 2005. Hum. Mol. Genet. 14:85-93), our data suggest that centromeric chromatin is arranged in a coiled 30-nm fiber that is itself coiled or folded to form a higher order structure.

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Examples of labeled putative chromatin fibers. The distance between parallel lines represents the measured width of the fiber, which is listed below each example. Five of the fibers depicted are cropped and rotated from centromeres in Figs. 2 and 3. Bar, 200 nm.
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fig4: Examples of labeled putative chromatin fibers. The distance between parallel lines represents the measured width of the fiber, which is listed below each example. Five of the fibers depicted are cropped and rotated from centromeres in Figs. 2 and 3. Bar, 200 nm.

Mentions: Localization of CENP-A to the inner kinetochore of human chromosomes. Each panel represents a complete set of serial sections through the centromere, and a schematic of the orientation of each chromosome precedes each set. (A) 60-nm serial sections through an acetone-fixed, colcemid-blocked mitotic chromosome. (C and E) 70-nm serial sections through PFA-fixed, colcemid-blocked mitotic chromosomes in different orientations. (G) 50-nm serial sections through an acetone-fixed unblocked mitotic chromosome. Details of the centromere regions (asterisks) from A, C, and G are shown in B, D, and H, respectively. Black arrowheads in B and D point to the kinetochore outer plate; white arrowheads demarcate putative chromatin fibers illustrated in Fig. 4. Arrowheads in H point to microtubules attached to the kinetochore. (F) A stylized representation of the CENP-A binding domain (red) on human chromosomes drawn to the proportions observed in this study (Table I). Bars, 200 nm.


Three-dimensional localization of CENP-A suggests a complex higher order structure of centromeric chromatin.

Marshall OJ, Marshall AT, Choo KH - J. Cell Biol. (2008)

Examples of labeled putative chromatin fibers. The distance between parallel lines represents the measured width of the fiber, which is listed below each example. Five of the fibers depicted are cropped and rotated from centromeres in Figs. 2 and 3. Bar, 200 nm.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2606971&req=5

fig4: Examples of labeled putative chromatin fibers. The distance between parallel lines represents the measured width of the fiber, which is listed below each example. Five of the fibers depicted are cropped and rotated from centromeres in Figs. 2 and 3. Bar, 200 nm.
Mentions: Localization of CENP-A to the inner kinetochore of human chromosomes. Each panel represents a complete set of serial sections through the centromere, and a schematic of the orientation of each chromosome precedes each set. (A) 60-nm serial sections through an acetone-fixed, colcemid-blocked mitotic chromosome. (C and E) 70-nm serial sections through PFA-fixed, colcemid-blocked mitotic chromosomes in different orientations. (G) 50-nm serial sections through an acetone-fixed unblocked mitotic chromosome. Details of the centromere regions (asterisks) from A, C, and G are shown in B, D, and H, respectively. Black arrowheads in B and D point to the kinetochore outer plate; white arrowheads demarcate putative chromatin fibers illustrated in Fig. 4. Arrowheads in H point to microtubules attached to the kinetochore. (F) A stylized representation of the CENP-A binding domain (red) on human chromosomes drawn to the proportions observed in this study (Table I). Bars, 200 nm.

Bottom Line: Combined with previous chromatin immunoprecipitation results (Saffery, R., H.Choo. 2005.Genet. 14:85-93), our data suggest that centromeric chromatin is arranged in a coiled 30-nm fiber that is itself coiled or folded to form a higher order structure.

View Article: PubMed Central - PubMed

Affiliation: Chromosome and Chromatin Research, Murdoch Childrens Research Institute, Royal Children's Hospital, Parkville, Victoria 3052, Australia.

ABSTRACT
The histone H3 variant centromere protein A (CENP-A) is central to centromere formation throughout eukaryotes. A long-standing question in centromere biology has been the organization of CENP-A at the centromere and its implications for the structure of centromeric chromatin. In this study, we describe the three-dimensional localization of CENP-A at the inner kinetochore plate through serial-section transmission electron microscopy of human mitotic chromosomes. At the kinetochores of normal centromeres and at a neocentromere, CENP-A occupies a compact domain at the inner kinetochore plate, stretching across two thirds of the length of the constriction but encompassing only one third of the constriction width and height. Within this domain, evidence of substructure is apparent. Combined with previous chromatin immunoprecipitation results (Saffery, R., H. Sumer, S. Hassan, L.H. Wong, J.M. Craig, K. Todokoro, M. Anderson, A. Stafford, and K.H.A. Choo. 2003. Mol. Cell. 12:509-516; Chueh, A.C., L.H. Wong, N. Wong, and K.H.A. Choo. 2005. Hum. Mol. Genet. 14:85-93), our data suggest that centromeric chromatin is arranged in a coiled 30-nm fiber that is itself coiled or folded to form a higher order structure.

Show MeSH
Related in: MedlinePlus