Mitotic Activation of a Novel Histone Deacetylase 3-Linker Histone H1.3 Protein Complex by Protein Kinase CK2

Hemangi Patil,Carrie Wilks,Rhiannon W Gonzalez,Sudheer Dhanireddy,Heather Conrad-Webb,Michael Bergel

doi:10.1074/jbc.m115.643874

Hemangi Patil, Carrie Wilks + Show 4 more

Open Access

https://doi.org/10.1074/jbc.m115.643874

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Abstract

Histone deacetylase 3 (HDAC3) and linker histone H1 are involved in both chromatin compaction and the regulation of mitotic progression. However, the mechanisms by which HDAC3 and H1 regulate mitosis and the factors controlling HDAC3 and H1 activity during mitosis are unclear. Furthermore, as of now, no association between class I, II, or IV (non-sirtuin) HDACs and linker histones has been reported. Here we describe a novel HDAC3-H1.3 complex containing silencing mediator of retinoic acid and thyroid hormone receptor (SMRT) and nuclear receptor corepressor 1 (N-CoR) that accumulated in synchronized HeLa cells in late G2 phase and mitosis. Nonetheless, the deacetylation activity by HDAC3 in the complex was evident only in mitotic complexes. HDAC3 associated with H1.3 was highly phosphorylated on Ser-424 only during mitosis. Isolation of inactive HDAC3-H1.3 complexes from late G2 phase cells, and phosphorylation of HDAC3 in the complexes at serine 424 by protein kinase CK2 (also known as casein kinase 2) activated the HDAC3 in vitro. In vivo, CK2α and CK2α' double knockdown cells demonstrated a significant decrease in HDAC3 Ser-424 phosphorylation during mitosis. HDAC3 and H1.3 co-localized in between the chromosomes, with polar microtubules and spindle poles during metaphase through telophase, and partially co-localized with chromatin during prophase and interphase. H1 has been reported previously to associate with microtubules and, therefore, could potentially function in targeting HDAC3 to the microtubules. We suggest that phosphorylation of HDAC3 in the complex by CK2 during mitosis activates the complex for a dual role: compaction of the mitotic chromatin and regulation of polar microtubules dynamic instability.

Highlights

Mitotic Activation of a Novel Histone Deacetylase 3-Linker Histone H1.3 Protein Complex by Protein Kinase CK2*
Histone Deacetylase 3 Associates with Histone H1.3 in Vivo and in Vitro—Histone deacetylases (HDACs) and linker histone H1 are known to play a role in the progression of mitosis and in chromatin compaction (4 – 6, 8)
When anti-H1 antibody was used for the reciprocal immunoprecipitations and anti-HDACs antibodies for the Western blotting analysis, an association could only be confirmed between H1 and Histone deacetylase 3 (HDAC3) (Fig. 1D)

Summary

Introduction

Mitotic Activation of a Novel Histone Deacetylase 3-Linker Histone H1.3 Protein Complex by Protein Kinase CK2*. Co-immunoprecipitation assays were carried out on protein extracts from each of these cell cycle phases using antibodies against either histone H1 (Fig. 2H) or HDAC3 (Fig. 2I), followed by Western blotting analysis for detection of HDAC3 (Fig. 2H) or H1.3 (Fig. 2I).

Results

Conclusion