Skip to Content
Merck
  • ATRX proximal protein associations boast roles beyond histone deposition.

ATRX proximal protein associations boast roles beyond histone deposition.

PLoS genetics (2021-11-16)
William A Scott, Erum Z Dhanji, Boris J A Dyakov, Ema S Dreseris, Jonathon S Asa, Laura J Grange, Mila Mirceta, Christopher E Pearson, Grant S Stewart, Anne-Claude Gingras, Eric I Campos
ABSTRACT

The ATRX ATP-dependent chromatin remodelling/helicase protein associates with the DAXX histone chaperone to deposit histone H3.3 over repetitive DNA regions. Because ATRX-protein interactions impart functions, such as histone deposition, we used proximity-dependent biotinylation (BioID) to identify proximal associations for ATRX. The proteomic screen captured known interactors, such as DAXX, NBS1, and PML, but also identified a range of new associating proteins. To gauge the scope of their roles, we examined three novel ATRX-associating proteins that likely differed in function, and for which little data were available. We found CCDC71 to associate with ATRX, but also HP1 and NAP1, suggesting a role in chromatin maintenance. Contrastingly, FAM207A associated with proteins involved in ribosome biosynthesis and localized to the nucleolus. ATRX proximal associations with the SLF2 DNA damage response factor help inhibit telomere exchanges. We further screened for the proteomic changes at telomeres when ATRX, SLF2, or both proteins were deleted. The loss caused important changes in the abundance of chromatin remodelling, DNA replication, and DNA repair factors at telomeres. Interestingly, several of these have previously been implicated in alternative lengthening of telomeres. Altogether, this study expands the repertoire of ATRX-associating proteins and functions.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
TERGITOL solution, Type NP-40, 70% in H2O
Sigma-Aldrich
Duolink® In Situ Detection Reagents FarRed
Sigma-Aldrich
Monoclonal ANTI-FLAG® M2 antibody produced in mouse, 1 mg/mL, clone M2, affinity isolated antibody, buffered aqueous solution (50% glycerol, 10 mM sodium phosphate, and 150 mM NaCl, pH 7.4)
Sigma-Aldrich
Poly-L-lysine solution, 0.1 % (w/v) in H2O
Sigma-Aldrich
Protease Inhibitor Cocktail, for use with mammalian cell and tissue extracts, DMSO solution
Sigma-Aldrich
Anti-ZFHX4 antibody produced in rabbit, Prestige Antibodies® Powered by Atlas Antibodies, affinity isolated antibody, buffered aqueous glycerol solution
Sigma-Aldrich
bisBenzimide H 33258, for fluorescence, ≥98.0% (HPLC)
Roche
Blocking Reagent, For nucleic acid hybridization and detection
Sigma-Aldrich
Anti-EZH2 Antibody, clone AE25, clone AE25, from mouse
Millipore
Benzonase® Nuclease, Purity > 90%
Sigma-Aldrich
DAPI, for nucleic acid staining