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Histone H4 Lysine 16 Hypoacetylation Is Associated with Defective DNA Repair and Premature Senescence in Zmpste24-Deficient Mice

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Histone H4 Lysine 16 Hypoacetylation Is Associated with Defective DNA Repair and Premature Senescence in Zmpste24-Deficient Mice Histone H4 lysine 16 hypoacetylation is associated with defective DNA repair and premature senescence in Zmpste24-deficient mice Vaidehi Krishnana,b,1, Maggie Zi Ying Chowa,1, Zimei Wanga,c,1, Le Zhanga,1, Baohua Liua,d, Xinguang Liud, and Zhongjun Zhoua,2 aDepartment of Biochemistry, Li Ka Shing Faculty of Medicine, Center of Development, Reproduction and Growth, University of Hong Kong, Hong Kong; bCancer Science Institute of Singapore, National University of Singapore, Singapore 117456; cDepartment of Biochemistry and Molecular Medicine, School of Medicine, Shenzhen University, Shenzhen 518060, China; and dInstitute for Aging Research, Guang Dong Medical College, Dongguan, China Edited by Neal G. Copeland, Institute of Molecular and Cell Biology, Proteos, Singapore, and approved June 20, 2011 (received for review February 25, 2011) Specific point mutations in lamin A gene have been shown to Zinc metallopeptidase, STE24 homolog (Zmpste24) is a met- alloproteinase responsible for the posttranslational processing accelerate aging in humans and mice. Particularly, a de novo −/− mutation at G608G position impairs lamin A processing to produce and cleavage of prelamin A into mature lamin A. The Zmpste24 the mutant protein progerin, which causes the Hutchinson Gilford mice exhibit premature aging features characterized by hair loss, growth retardation, osteoporosis, and premature mortality (12). progeria syndrome. The premature aging phenotype of Hutchin- These progeroid features are also manifested in Hutchinson son Gilford progeria syndrome is largely recapitulated in mice de- Gilford progeria syndrome (HGPS) patients, a syndrome that ficient for the lamin A-processing enzyme, Zmpste24. We have arises due to a de novo lamin A gene (LMNA) point mutation previously reported that Zmpste24 deficiency results in genomic giving rise to a truncated protein, progerin (13). In our previous instability and early cellular senescence due to the delayed recruit- work, a relationship between genomic instability and premature− − ment of repair proteins to sites of DNA damage. Here, we further aging was established (14). We showed that Zmpste24 / mouse investigate the molecular mechanism underlying delayed DNA embryonic fibroblasts (MEFs) and HGPS cells in culture accu- −/− damage response and identify a histone acetylation defect in mulated DNA damage and chromosome aberrations. Zmpste24 −/− CELL BIOLOGY Zmpste24 mice. Specifically, histone H4 was hypoacetylated at MEFs were highly sensitive to a wide variety of DNA-damaging a lysine 16 residue (H4K16), and this defect was attributed to the agents and exhibited early cellular senescence in culture. In par- ticular, upon DSB induction, a significant delay in the recruitment reduced association of a histone acetyltransferase, Mof, to the of 53BP1 and Rad51 was noted in Zmpste24-null cells and HGPS nuclear matrix. Given the reversible nature of epigenetic changes, fibroblasts, a finding corroborated later by several other reports rescue experiments performed either by Mof overexpression or by (15–18). Together, these studies have led to the idea that prel- histone deacetylase inhibition promoted repair protein recruit- amin A/progerin accumulation interferes with the loading of ment to DNA damage sites and substantially ameliorated aging- DNA repair proteins to DSB sites and that, as a consequence, associated phenotypes, both in vitro and in vivo. The life span of nonrepairable DNA damage remains and early cellular senes- −/− Zmpste24 mice was also extended with the supplementation of cence occurs to cause premature aging. a histone deacetylase inhibitor, sodium butyrate, to drinking wa- The proper conformation of chromatin is essential for the re- ter. Consistent with recent data showing age-dependent buildup cruitment of DNA repair proteins to DSBs. In response to DNA damage, chromatin structure is altered by enzymes that remodel of unprocessable lamin A in physiological aging, aged wild-type chromatin or carry out the epigenetic modification of histones. mice also showed hypoacetylation of H4K16. The above results Consequently, chromatin surrounding DSBs undergoes re- shed light on how chromatin modifications regulate the DNA dam- laxation to generate a compartment capable of recruiting and age response and suggest that the reversal of epigenetic marks retaining repair factors (19–21). We hypothesized that mutant could make an attractive therapeutic target against laminopathy- lamin A might have a severe effect on chromatin organization due based progeroid pathologies. to aberrant epigenetic modification of histones. Recent reports have shown that histone H4 acetylation at lysine 16 (H4K16) plays an important role in DNA damage response and DSB repair (22, ukaryotic cells are equipped with a surveillance machinery to fi orchestrate the rapid detection and repair of DNA damage. 23). Speci cally, H4K16 acetylation neutralizes the positive E charge on the H4 tail, which then poses a structural constraint on When DNA damage occurs, chromatin surrounding the double- fi the formation of higher-order chromatin and forces the chro- strand breaks (DSBs) is altered and histones are modi ed to matin to stay in a more open configuration. facilitate access for repair proteins (1). As a rapid response to Here, we describe impaired histone H4 acetylation in the DSB induction, the histone H2A variant, H2AX, is phosphory- Zmpste24-deficient cells. Our data indicate that reduced H4K16 lated at Ser139 (γ-H2AX), which in turn interacts with MDC1, acetylation contributes to the− − delayed recruitment of DNA re- a DSB repair mediator, to facilitate the further recruitment of pair proteins in Zmpste24 / MEFs. Furthermore, pharmaco- DNA repair proteins, such as 53BP1 and BRCA1 (2–4). In- γ logical modulation of histone acetylation was found to delay terestingly, -H2AX accumulation has been documented both in premature cellular senescence and aging. Together with similar fi human senescent cells and in the broblasts of aged mice and epigenetic patterns observed in wild-type mice during normal primates (5–8). It has been proposed that these age-associated physiological aging, these findings not only provide a paradigm γ-H2AX foci contain nonrepairable DSBs and may have a role in initiating aging, especially because DSBs are very toxic and are one of the most lethal forms of DNA damage. Direct evidence Author contributions: V.K., M.Z.Y.C., and Z.Z. designed research; V.K., M.Z.Y.C., Z.W., L.Z., for nonrepairable DNA damage as an inducer of premature and B.L. performed research; X.L. contributed new reagents/analytic tools; V.K., M.Z.Y.C., aging has been obtained from mouse models that lack DNA Z.W., L.Z., B.L., X.L., and Z.Z. analyzed data; and V.K., M.Z.Y.C., and Z.Z. wrote the paper. repair proteins, such as ATM, Ku70, Ku80, DNA ligase IV, and The authors declare no conflict of interest. Ercc1, as well as from humans with premature aging syndromes (9, 10). Together, these studies support the idea that the inability This article is a PNAS Direct Submission. to recruit repair proteins to sites of DNA lesions leads to the 1V.K., M.Z.Y.C., Z.W., and L.Z. contributed equally to this work. accumulation of irreparable DNA damage, which predisposes 2To whom correspondence should be addressed. E-mail: [email protected]. cells to premature senescence, growth retardation, and acceler- This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. ated aging (11). 1073/pnas.1102789108/-/DCSupplemental. www.pnas.org/cgi/doi/10.1073/pnas.1102789108 PNAS Early Edition | 1of6 Downloaded by guest on September 28, 2021 for the role of epigenetic signaling in DNA damage repair, but point at which the mice had not yet developed any signs of pre- also extend the mechanistic link between histone modifications, mature aging (Fig. 1F). Then, to ensure that the down-regulation genomic instability, and aging. of H4 acetylation was not a reflection of global aberrations in epigenetic modifications, other modification sites on histone H3 Results and H4 were analyzed. No significant alteration could be detec- Reduced H4K16 Acetylation in Zmpste24-Deficient MEFs. To evaluate ted in the overall H3 acetylation, H3K79 di-methylation, or the association between epigenetic events and aging, we com- H4K20 di-methylation (Fig. S1A). The relationship between pared the histone modification patterns between wild-type and prelamin A and H4 acetylation was further investigated using − − Zmpste24 / mutant cells. Under identical culture conditions, HEK293 cells that ectopically expressed unprocessable prelamin +/+ −/− A (C-terminal Ser-Ile-Met replaced with Phe-Phe-Met). There Zmpste24 and Zmpste24 MEFs were negative for senes- β β was a marked reduction of H4 and H4K16 acetylation in cence-associated -galactosidase (SA -gal) staining at early HEK293 cells upon transfection with unprocessable prelamin A passage (passage 3) (Fig. 1A, Upper panels). However, upon ∼ −/− (Fig. S1B). Thus, these results suggest that prelamin A accumu- continuous passaging (passage 6), 60% of the Zmpste24 lation impairs H4K16 acetylation. MEFs showed SA β-gal positivity and flattened morphology (Fig. 1A, Lower panels). To study whether early senescence in − − Prelamin A Accumulation Decreased Mof Association on the Nuclear Zmpste24 / MEFs was caused by altered chromatin mod- fi fi fi Matrix. H4K16 acetylation in mammalian cells is catalyzed mainly i cations, histone modi cation at speci c sites was examined in by the histone acetyltransferase Mof (24, 25). Mof is important
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