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Histone hypercitrullination mediates chromatin decondensation and neutrophil extracellular trap formation.

Wang Y, Li M, Stadler S, Correll S, Li P, Wang D, Hayama R, Leonelli L, Han H, Grigoryev SA, Allis CD, Coonrod SA - J. Cell Biol. (2009)

Bottom Line: The inhibition of PAD4 decreases histone hypercitrullination and the formation of NET-like structures, whereas PAD4 treatment of HL-60 cells facilitates these processes.The loss of heterochromatin and multilobular nuclear structures is detected in HL-60 granulocytes after PAD4 activation.Importantly, citrullination of biochemically defined avian nucleosome arrays inhibits their compaction by the linker histone H5 to form higher order chromatin structures.

View Article: PubMed Central - PubMed

Affiliation: Center for Gene Regulation, Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park, PA 16802, USA. yuw12@psu.edu

ABSTRACT
Peripheral blood neutrophils form highly decondensed chromatin structures, termed neutrophil extracellular traps (NETs), that have been implicated in innate immune response to bacterial infection. Neutrophils express high levels of peptidylarginine deiminase 4 (PAD4), which catalyzes histone citrullination. However, whether PAD4 or histone citrullination plays a role in chromatin structure in neutrophils is unclear. In this study, we show that the hypercitrullination of histones by PAD4 mediates chromatin decondensation. Histone hypercitrullination is detected on highly decondensed chromatin in HL-60 granulocytes and blood neutrophils. The inhibition of PAD4 decreases histone hypercitrullination and the formation of NET-like structures, whereas PAD4 treatment of HL-60 cells facilitates these processes. The loss of heterochromatin and multilobular nuclear structures is detected in HL-60 granulocytes after PAD4 activation. Importantly, citrullination of biochemically defined avian nucleosome arrays inhibits their compaction by the linker histone H5 to form higher order chromatin structures. Together, these results suggest that histone hypercitrullination has important functions in chromatin decondensation in granulocytes/neutrophils.

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Histone citrullination inhibits nucleosome array compaction by linker histone H5. (A) A 35-S structure was detected after treatment of the 207 × 12 array (containing 12 nucleosome core particles in an array) with the GST-PAD4C645S mutant or GST-PAD4. (B) After adding the linker histone H5, the 207 × 12 nucleosome core particle array pretreated with the PAD4C645S mutant was detected as a 50-S structure, whereas the 207 × 12 nucleosome core particle array pretreated with PAD4 was detected as a 40-S structure. (C) The H5-bound 207 × 12 array treated with the PAD4C645S mutant was detected as a 53-S structure, whereas the H5-bound 207 × 12 array treated with PAD4 was detected as a 45-S structure.
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fig5: Histone citrullination inhibits nucleosome array compaction by linker histone H5. (A) A 35-S structure was detected after treatment of the 207 × 12 array (containing 12 nucleosome core particles in an array) with the GST-PAD4C645S mutant or GST-PAD4. (B) After adding the linker histone H5, the 207 × 12 nucleosome core particle array pretreated with the PAD4C645S mutant was detected as a 50-S structure, whereas the 207 × 12 nucleosome core particle array pretreated with PAD4 was detected as a 40-S structure. (C) The H5-bound 207 × 12 array treated with the PAD4C645S mutant was detected as a 53-S structure, whereas the H5-bound 207 × 12 array treated with PAD4 was detected as a 45-S structure.

Mentions: To analyze whether histone citrullination affects the nucleosome compaction, we analyzed the density of the 207 × 12 nucleosome core particle array (Springhetti et al., 2003), which contains 12 nucleosome core particles assembled with core histones H3, H2B, H2A, and H4 on a defined DNA template, treated with GST-PAD4 or the GST-PAD4C645S mutant. The ability of PAD4 to citrullinate nucleosomal histones was confirmed by Western blotting (unpublished data). Consistent with the maintenance of the primary nucleosome structure (Fig. 3 I), there was no difference between the density of the 207 × 12 nucleosome core particle array treated with PAD4 or the PAD4C645S mutant (Fig. 5 A). This result suggests that under current experimental conditions, citrullination of histones by PAD4 did not generate detectable conformation changes in the nucleosomal core particle arrays.


Histone hypercitrullination mediates chromatin decondensation and neutrophil extracellular trap formation.

Wang Y, Li M, Stadler S, Correll S, Li P, Wang D, Hayama R, Leonelli L, Han H, Grigoryev SA, Allis CD, Coonrod SA - J. Cell Biol. (2009)

Histone citrullination inhibits nucleosome array compaction by linker histone H5. (A) A 35-S structure was detected after treatment of the 207 × 12 array (containing 12 nucleosome core particles in an array) with the GST-PAD4C645S mutant or GST-PAD4. (B) After adding the linker histone H5, the 207 × 12 nucleosome core particle array pretreated with the PAD4C645S mutant was detected as a 50-S structure, whereas the 207 × 12 nucleosome core particle array pretreated with PAD4 was detected as a 40-S structure. (C) The H5-bound 207 × 12 array treated with the PAD4C645S mutant was detected as a 53-S structure, whereas the H5-bound 207 × 12 array treated with PAD4 was detected as a 45-S structure.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2654299&req=5

fig5: Histone citrullination inhibits nucleosome array compaction by linker histone H5. (A) A 35-S structure was detected after treatment of the 207 × 12 array (containing 12 nucleosome core particles in an array) with the GST-PAD4C645S mutant or GST-PAD4. (B) After adding the linker histone H5, the 207 × 12 nucleosome core particle array pretreated with the PAD4C645S mutant was detected as a 50-S structure, whereas the 207 × 12 nucleosome core particle array pretreated with PAD4 was detected as a 40-S structure. (C) The H5-bound 207 × 12 array treated with the PAD4C645S mutant was detected as a 53-S structure, whereas the H5-bound 207 × 12 array treated with PAD4 was detected as a 45-S structure.
Mentions: To analyze whether histone citrullination affects the nucleosome compaction, we analyzed the density of the 207 × 12 nucleosome core particle array (Springhetti et al., 2003), which contains 12 nucleosome core particles assembled with core histones H3, H2B, H2A, and H4 on a defined DNA template, treated with GST-PAD4 or the GST-PAD4C645S mutant. The ability of PAD4 to citrullinate nucleosomal histones was confirmed by Western blotting (unpublished data). Consistent with the maintenance of the primary nucleosome structure (Fig. 3 I), there was no difference between the density of the 207 × 12 nucleosome core particle array treated with PAD4 or the PAD4C645S mutant (Fig. 5 A). This result suggests that under current experimental conditions, citrullination of histones by PAD4 did not generate detectable conformation changes in the nucleosomal core particle arrays.

Bottom Line: The inhibition of PAD4 decreases histone hypercitrullination and the formation of NET-like structures, whereas PAD4 treatment of HL-60 cells facilitates these processes.The loss of heterochromatin and multilobular nuclear structures is detected in HL-60 granulocytes after PAD4 activation.Importantly, citrullination of biochemically defined avian nucleosome arrays inhibits their compaction by the linker histone H5 to form higher order chromatin structures.

View Article: PubMed Central - PubMed

Affiliation: Center for Gene Regulation, Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park, PA 16802, USA. yuw12@psu.edu

ABSTRACT
Peripheral blood neutrophils form highly decondensed chromatin structures, termed neutrophil extracellular traps (NETs), that have been implicated in innate immune response to bacterial infection. Neutrophils express high levels of peptidylarginine deiminase 4 (PAD4), which catalyzes histone citrullination. However, whether PAD4 or histone citrullination plays a role in chromatin structure in neutrophils is unclear. In this study, we show that the hypercitrullination of histones by PAD4 mediates chromatin decondensation. Histone hypercitrullination is detected on highly decondensed chromatin in HL-60 granulocytes and blood neutrophils. The inhibition of PAD4 decreases histone hypercitrullination and the formation of NET-like structures, whereas PAD4 treatment of HL-60 cells facilitates these processes. The loss of heterochromatin and multilobular nuclear structures is detected in HL-60 granulocytes after PAD4 activation. Importantly, citrullination of biochemically defined avian nucleosome arrays inhibits their compaction by the linker histone H5 to form higher order chromatin structures. Together, these results suggest that histone hypercitrullination has important functions in chromatin decondensation in granulocytes/neutrophils.

Show MeSH
Related in: MedlinePlus