TY - JOUR
T1 - Cse4 Is Part of an Octameric Nucleosome in Budding Yeast
AU - Camahort, Raymond
AU - Shivaraju, Manjunatha
AU - Mattingly, Mark
AU - Li, Bing
AU - Nakanishi, Shima
AU - Zhu, Dongxiao
AU - Shilatifard, Ali
AU - Workman, Jerry L.
AU - Gerton, Jennifer L.
N1 - Funding Information:
We would like to thank Sue Biggins for strains and plasmids, Chris Seidel for assistance with design and printing of DNA microarrays, Carl Wu for strains and antibodies, and the Stowers Institute Molecular Biology Facility for help with mutagenesis. We would like to thank Laurence Florens for her help with proteomic analysis. We would like to thank Swami Venkatesh and Jeong-Hoon Kim for reagents and helpful discussion. We would also like to thank Joan Conaway for thoughtful discussion and feedback, as well as all of the members of the Gerton lab for their help and support. Some of the experiments described herein were done to fulfill, in part, requirements for M.S.'s PhD thesis research as a student registered with the Open University. This research was supported by National Institutes of Health (NIH) R01GM080477 and the Stowers Institute for Medical Research. B.L. is supported by a grant from the Welch Foundation (I-1713) and is a W.A. “Tex” Moncrief, Jr. Scholar in Medical Research.
PY - 2009/9/24
Y1 - 2009/9/24
N2 - The budding yeast CenH3 histone variant Cse4 localizes to centromeric nucleosomes and is required for kinetochore assembly and chromosome segregation. The exact composition of centromeric Cse4-containing nucleosomes is a subject of debate. Using unbiased biochemical, cell-biological, and genetic approaches, we have tested the composition of Cse4-containing nucleosomes. Using micrococcal nuclease-treated chromatin, we find that Cse4 is associated with the histones H2A, H2B, and H4, but not H3 or the nonhistone protein Scm3. Overexpression of Cse4 rescues the lethality of a scm3 deletion, indicating that Scm3 is not essential for the formation of functional centromeric chromatin. We also find that octameric Cse4 nucleosomes can be reconstituted in vitro. Furthermore, Cse4-Cse4 dimerization occurs in vivo at the centromeric nucleosome, and this requires the predicted Cse4-Cse4 dimerization interface. Taken together, our experimental evidence supports the model that the Cse4 nucleosome is an octamer, containing two copies each of Cse4, H2A, H2B, and H4.
AB - The budding yeast CenH3 histone variant Cse4 localizes to centromeric nucleosomes and is required for kinetochore assembly and chromosome segregation. The exact composition of centromeric Cse4-containing nucleosomes is a subject of debate. Using unbiased biochemical, cell-biological, and genetic approaches, we have tested the composition of Cse4-containing nucleosomes. Using micrococcal nuclease-treated chromatin, we find that Cse4 is associated with the histones H2A, H2B, and H4, but not H3 or the nonhistone protein Scm3. Overexpression of Cse4 rescues the lethality of a scm3 deletion, indicating that Scm3 is not essential for the formation of functional centromeric chromatin. We also find that octameric Cse4 nucleosomes can be reconstituted in vitro. Furthermore, Cse4-Cse4 dimerization occurs in vivo at the centromeric nucleosome, and this requires the predicted Cse4-Cse4 dimerization interface. Taken together, our experimental evidence supports the model that the Cse4 nucleosome is an octamer, containing two copies each of Cse4, H2A, H2B, and H4.
KW - CELLCYCLE
KW - DNA
KW - PROTEINS
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U2 - 10.1016/j.molcel.2009.07.022
DO - 10.1016/j.molcel.2009.07.022
M3 - Article
C2 - 19782029
AN - SCOPUS:70349168454
SN - 1097-2765
VL - 35
SP - 794
EP - 805
JO - Molecular cell
JF - Molecular cell
IS - 6
ER -