An ATPase-Deficient Variant of the SNF2 Family Member HELLS Shows Altered Dynamics at Pericentromeric Heterochromatin. Issue 10 (22nd May 2015)

• Record Type: Journal Article
• Authors: Lungu, Cristiana; Muegge, Kathrin; Jeltsch, Albert; Jurkowska, Renata Z.
• Is Part Of: Journal of molecular biology. Volume 427:Issue 10(2015:May 15)
• Page Start: 1903
• Page End: 1915
• Publication Date: 2015-05-22
• Subjects: DAPI 4′, 6-diamidino-2-phenylindole -- FRAP fluorescence recovery after photobleaching -- KO knockout -- MEF mouse embryonic fibroblast; chromatin remodeling -- ATP hydrolysis -- protein dynamics -- fluorescence recovery after photobleaching -- heterochromatin; Molecular biology -- Periodicals; Biology -- Periodicals; Biochemistry -- Periodicals; Bacteriology -- Periodicals; Molecular Biology -- Periodicals; Biochemistry -- Periodicals; Biologie moléculaire -- Périodiques; Biologie -- Périodiques; Biochimie -- Périodiques; Moleculaire biologie; Biochemistry; Biology; Molecular biology; Periodicals; 572.805
• DOI: 10.1016/j.jmb.2015.03.014
• Languages: English
• ISSNs: 0022-2836

Abstract

Abstract: The HELLS ( hel icase, l ymphoid s pecific, also known as lymphoid-specific helicase) protein is related to the SNF2 ( s ucrose n on- f ermentable 2 ) family of chromatin remodeling ATPases. It is required for efficient DNA methylation in mammals, particularly at heterochromatin-located repetitive sequences. In this study, we investigated the interaction of HELLS with chromatin and used an ATPase-deficient HELLS variant to address the role of ATP hydrolysis in this process. Chromatin fractionation experiments demonstrated that, in the absence of the ATPase activity, HELLS is retained at the nuclear matrix compartment, defined in part by lamin B1. Microscopy studies revealed a stronger association of the ATPase-deficient mutant with heterochromatin. These results were further supported by fluorescence recovery after photobleaching measurements, which showed that, at heterochromatic sites, wild-type HELLS is very dynamic, with a recovery half-time of 0.8 s and a mobile protein fraction of 61%. In contrast, the ATPase-deficient mutant displayed 4.5-s recovery half-time and a reduction in the mobile fraction to 30%. We also present evidence suggesting that, in addition to the ATPase activity, a functional H3K9me3 signaling pathway contributes to an efficient release of HELLS from pericentromeric chromatin. Overall, our results show that a functional ATPase activity is not required for the recruitment of HELLS to heterochromatin, but it is important for the release of the enzyme from these sites. Graphical Abstract: Highlights: We studied the chromatin interaction of an ATPase-deficient HELLS mutant. The mutant was enriched in the nuclear matrix fraction of chromatin. The mutant showed higher accumulation at pericentromeric heterochromatin. Fluorescence recovery after photobleaching measurements showed a 5-fold decreased mobility of the mutant. ATPase activity is essential for the release of HELLS from compacted chromatin.