The histone H3/H4 chaperone CHAF1B prevents the mislocalization of CENP-A for chromosomal stability

Roshan L. Shrestha; Vinutha Balachandra; Jee Hun Kim; Austin Rossi; Pranathi Vadlamani; Subhash Chandra Sethi; Laurent Ozbun; Shinjen Lin; Ken Chin Chien Cheng; Raj Chari; Tatiana S. Karpova; Gianluca Pegoraro; Daniel R. Foltz; Natasha J. Caplen; Munira A. Basrai

doi:10.1242/JCS.260944

The histone H3/H4 chaperone CHAF1B prevents the mislocalization of CENP-A for chromosomal stability

Roshan L. Shrestha, Vinutha Balachandra, Jee Hun Kim, Austin Rossi, Pranathi Vadlamani, Subhash Chandra Sethi, Laurent Ozbun, Shinjen Lin, Ken Chin Chien Cheng, Raj Chari, Tatiana S. Karpova, Gianluca Pegoraro, Daniel R. Foltz, Natasha J. Caplen, Munira A. Basrai^*

^*Corresponding author for this work

Biochemistry and Molecular Genetics

Research output: Contribution to journal › Article › peer-review

9 Scopus citations

Abstract

Restricting the localization of the evolutionarily conserved centromeric histone H3 variant CENP-A to centromeres prevents chromosomal instability (CIN). The mislocalization of CENP-A to noncentromeric regions contributes to CIN in yeasts, flies and human cells. Even though overexpression and mislocalization of CENP-A have been reported in cancers, the mechanisms responsible for its mislocalization remain poorly understood. Here, we used an imagingbased high-throughput RNAi screen to identify factors that prevent mislocalization of overexpressed YFP-tagged CENP-A (YFP-CENPA) in HeLa cells. Among the top five candidates in the screen - the depletion of which showed increased nuclear YFP-CENP-A fluorescence - were the histone chaperones CHAF1B (or p60) and CHAF1A (or p150). Follow-up validation and characterization experiments showed that CHAF1B-depleted cells exhibited CENPA mislocalization, CIN phenotypes and increased enrichment of CENP-A in chromatin fractions. The depletion of DAXX, a histone H3.3 chaperone, suppressed CENP-A mislocalization and CIN in CHAF1B-depleted cells. We propose that in CHAF1B-depleted cells, DAXX promotes mislocalization of the overexpressed CENP-A to non-centromeric regions, resulting in CIN. In summary, we identified regulators of CENP-A localization and defined a role for CHAF1B in preventing DAXX-dependent CENP-A mislocalization and CIN.

Original language	English (US)
Article number	jcs260944
Journal	Journal of cell science
Volume	136
Issue number	10
DOIs	https://doi.org/10.1242/JCS.260944
State	Published - May 2023

Funding

This work was supported by the Intramural Research Program of the National Cancer Institute (NCI), Center for Cancer Research (CCR): project numbers ZIA BC 010822 (M.A.B.), ZIA BC 011704 (N.J.C.) and 1-ZIC-BC011567-01 (L.O. and G.P.). R.C. was supported by the National Cancer Institute/National Institutes of Health under the contract no. HHSN261201500003I. S.L. and K.C. were supported by the Intramural Research Program of the National Institutes of Health. D.R.F. and P.V. were supported by the National Institutes of Health grants R01GM111907 and UC01CA260699. Open access funding provided by the National Institutes of Health. Deposited in PMC for immediate release.

Keywords

CENP-A
CHAF1B
Chromosomal instability
DAXX

ASJC Scopus subject areas

Cell Biology

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1242/JCS.260944

Cite this

Shrestha, R. L., Balachandra, V., Kim, J. H., Rossi, A., Vadlamani, P., Sethi, S. C., Ozbun, L., Lin, S., Cheng, K. C. C., Chari, R., Karpova, T. S., Pegoraro, G., Foltz, D. R., Caplen, N. J., & Basrai, M. A. (2023). The histone H3/H4 chaperone CHAF1B prevents the mislocalization of CENP-A for chromosomal stability. Journal of cell science, 136(10), Article jcs260944. https://doi.org/10.1242/JCS.260944

@article{33334b35291841bd88ce1414949b67ef,

title = "The histone H3/H4 chaperone CHAF1B prevents the mislocalization of CENP-A for chromosomal stability",

abstract = "Restricting the localization of the evolutionarily conserved centromeric histone H3 variant CENP-A to centromeres prevents chromosomal instability (CIN). The mislocalization of CENP-A to noncentromeric regions contributes to CIN in yeasts, flies and human cells. Even though overexpression and mislocalization of CENP-A have been reported in cancers, the mechanisms responsible for its mislocalization remain poorly understood. Here, we used an imagingbased high-throughput RNAi screen to identify factors that prevent mislocalization of overexpressed YFP-tagged CENP-A (YFP-CENPA) in HeLa cells. Among the top five candidates in the screen - the depletion of which showed increased nuclear YFP-CENP-A fluorescence - were the histone chaperones CHAF1B (or p60) and CHAF1A (or p150). Follow-up validation and characterization experiments showed that CHAF1B-depleted cells exhibited CENPA mislocalization, CIN phenotypes and increased enrichment of CENP-A in chromatin fractions. The depletion of DAXX, a histone H3.3 chaperone, suppressed CENP-A mislocalization and CIN in CHAF1B-depleted cells. We propose that in CHAF1B-depleted cells, DAXX promotes mislocalization of the overexpressed CENP-A to non-centromeric regions, resulting in CIN. In summary, we identified regulators of CENP-A localization and defined a role for CHAF1B in preventing DAXX-dependent CENP-A mislocalization and CIN.",

keywords = "CENP-A, CHAF1B, Chromosomal instability, DAXX",

author = "Shrestha, {Roshan L.} and Vinutha Balachandra and Kim, {Jee Hun} and Austin Rossi and Pranathi Vadlamani and Sethi, {Subhash Chandra} and Laurent Ozbun and Shinjen Lin and Cheng, {Ken Chin Chien} and Raj Chari and Karpova, {Tatiana S.} and Gianluca Pegoraro and Foltz, {Daniel R.} and Caplen, {Natasha J.} and Basrai, {Munira A.}",

note = "Publisher Copyright: {\textcopyright} 2023. Published by The Company of Biologists Ltd.",

year = "2023",

month = may,

doi = "10.1242/JCS.260944",

language = "English (US)",

volume = "136",

journal = "Journal of cell science",

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Shrestha, RL, Balachandra, V, Kim, JH, Rossi, A, Vadlamani, P, Sethi, SC, Ozbun, L, Lin, S, Cheng, KCC, Chari, R, Karpova, TS, Pegoraro, G, Foltz, DR, Caplen, NJ & Basrai, MA 2023, 'The histone H3/H4 chaperone CHAF1B prevents the mislocalization of CENP-A for chromosomal stability', Journal of cell science, vol. 136, no. 10, jcs260944. https://doi.org/10.1242/JCS.260944

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T1 - The histone H3/H4 chaperone CHAF1B prevents the mislocalization of CENP-A for chromosomal stability

AU - Shrestha, Roshan L.

AU - Balachandra, Vinutha

AU - Kim, Jee Hun

AU - Rossi, Austin

AU - Vadlamani, Pranathi

AU - Sethi, Subhash Chandra

AU - Ozbun, Laurent

AU - Lin, Shinjen

AU - Cheng, Ken Chin Chien

AU - Chari, Raj

AU - Karpova, Tatiana S.

AU - Pegoraro, Gianluca

AU - Foltz, Daniel R.

AU - Caplen, Natasha J.

AU - Basrai, Munira A.

PY - 2023/5

Y1 - 2023/5

N2 - Restricting the localization of the evolutionarily conserved centromeric histone H3 variant CENP-A to centromeres prevents chromosomal instability (CIN). The mislocalization of CENP-A to noncentromeric regions contributes to CIN in yeasts, flies and human cells. Even though overexpression and mislocalization of CENP-A have been reported in cancers, the mechanisms responsible for its mislocalization remain poorly understood. Here, we used an imagingbased high-throughput RNAi screen to identify factors that prevent mislocalization of overexpressed YFP-tagged CENP-A (YFP-CENPA) in HeLa cells. Among the top five candidates in the screen - the depletion of which showed increased nuclear YFP-CENP-A fluorescence - were the histone chaperones CHAF1B (or p60) and CHAF1A (or p150). Follow-up validation and characterization experiments showed that CHAF1B-depleted cells exhibited CENPA mislocalization, CIN phenotypes and increased enrichment of CENP-A in chromatin fractions. The depletion of DAXX, a histone H3.3 chaperone, suppressed CENP-A mislocalization and CIN in CHAF1B-depleted cells. We propose that in CHAF1B-depleted cells, DAXX promotes mislocalization of the overexpressed CENP-A to non-centromeric regions, resulting in CIN. In summary, we identified regulators of CENP-A localization and defined a role for CHAF1B in preventing DAXX-dependent CENP-A mislocalization and CIN.

AB - Restricting the localization of the evolutionarily conserved centromeric histone H3 variant CENP-A to centromeres prevents chromosomal instability (CIN). The mislocalization of CENP-A to noncentromeric regions contributes to CIN in yeasts, flies and human cells. Even though overexpression and mislocalization of CENP-A have been reported in cancers, the mechanisms responsible for its mislocalization remain poorly understood. Here, we used an imagingbased high-throughput RNAi screen to identify factors that prevent mislocalization of overexpressed YFP-tagged CENP-A (YFP-CENPA) in HeLa cells. Among the top five candidates in the screen - the depletion of which showed increased nuclear YFP-CENP-A fluorescence - were the histone chaperones CHAF1B (or p60) and CHAF1A (or p150). Follow-up validation and characterization experiments showed that CHAF1B-depleted cells exhibited CENPA mislocalization, CIN phenotypes and increased enrichment of CENP-A in chromatin fractions. The depletion of DAXX, a histone H3.3 chaperone, suppressed CENP-A mislocalization and CIN in CHAF1B-depleted cells. We propose that in CHAF1B-depleted cells, DAXX promotes mislocalization of the overexpressed CENP-A to non-centromeric regions, resulting in CIN. In summary, we identified regulators of CENP-A localization and defined a role for CHAF1B in preventing DAXX-dependent CENP-A mislocalization and CIN.

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KW - CHAF1B

KW - Chromosomal instability

KW - DAXX

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DO - 10.1242/JCS.260944

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AN - SCOPUS:85160877726

SN - 0021-9533

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JO - Journal of cell science

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The histone H3/H4 chaperone CHAF1B prevents the mislocalization of CENP-A for chromosomal stability

Abstract

Funding

Keywords

ASJC Scopus subject areas

UN SDGs

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