Open AccessProteomic Analysis of Histones H2A/H2B and Variant Hv1 in Tetrahymena thermophila Reveals an Ancient Network of Chaperones
Author(s) -
Kanwal Ashraf,
Syed NabeelShah,
Jyoti Garg,
Alejandro Saettone,
Joanna Derynck,
AnneClaude Gingras,
Jean-Philippe Lambert,
Ronald E. Pearlman,
Jeffrey Fillingham
Publication year - 2019
Publication title -
molecular biology and evolution
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 6.637
H-Index - 218
eISSN - 1537-1719
pISSN - 0737-4038
DOI - 10.1093/molbev/msz039
Subject(s) - tetrahymena , biology , histone h2b , histone , genetics , evolutionary biology , microbiology and biotechnology , gene
Epigenetic information, which can be passed on independently of the DNA sequence, is stored in part in the form of histone posttranslational modifications and specific histone variants. Although complexes necessary for deposition have been identified for canonical and variant histones, information regarding the chromatin assembly pathways outside of the Opisthokonts remains limited. Tetrahymena thermophila, a ciliated protozoan, is particularly suitable to study and unravel the chromatin regulatory layers due to its unique physical separation of chromatin states in the form of two distinct nuclei present within the same cell. Using a functional proteomics pipeline, we carried out affinity purification followed by mass spectrometry of endogenously tagged T. thermophila histones H2A, H2B and variant Hv1.We identified a set of interacting proteins shared among the three analyzed histones that includes the FACT-complex, as well as H2A- or Hv1-specific chaperones. We find that putative subunits of T. thermophila versions of SWR- and INO80-complexes, as well as transcription-related histone chaperone Spt6Tt specifically copurify with Hv1. We also identified importin β6 and the T. thermophila ortholog of nucleoplasmin 1 (cNpl1Tt) as H2A-H2B interacting partners. Our results further implicate Poly [ADP-ribose] polymerases in histone metabolism. Molecular evolutionary analysis, reciprocal affinity purification coupled to mass spectrometry experiments, and indirect immunofluorescence studies using endogenously tagged Spt16Tt (FACT-complex subunit), cNpl1Tt, and PARP6Tt underscore the validity of our approach and offer mechanistic insights. Our results reveal a highly conserved regulatory network for H2A (Hv1)-H2B concerning their nuclear import and assembly into chromatin.