Chromatin regulators play important roles in the safeguarding of cell identities by opposing the induction of ectopic cell fates and, thereby, preventing forced conversion of cell identities by reprogramming approaches. Our knowledge of chromatin regulators acting as reprogramming barriers in living organisms needs improvement as most studies use tissue culture. We used Caenorhabditis elegans as an in vivo gene discovery model and automated solid-phase RNA interference screening, by which we identified 10 chromatin-regulating factors that protect cells against ectopic fate induction. Specifically, the chromodomain protein MRG-1 safeguards germ cells against conversion into neurons. MRG-1 is the ortholog of mammalian MRG15 (MORF-related gene on chromosome 15) and is required during germline development in C. elegans. However, MRG-1’s function as a barrier for germ cell reprogramming has not been revealed previously. Here, we further provide protein-protein and genome interactions of MRG-1 to characterize its molecular functions. Conserved chromatin regulators may have similar functions in higher organisms, and therefore, understanding cell fate protection in C. elegans may also help to facilitate reprogramming of human cells.

T+49 (0)30 450540501 (Translational design, muscle stem cells). One sentence summary: Patient primary muscle stems cells gene repaired with >90% efficiency by base editing maintain their regenerative properties for autologous cell replacement therapies of muscular dystrophy.

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Whether extension of lifespan provides an extended time without health deteriorations is an important issue for human aging. However, to which degree lifespan and aspects of healthspan regulation might be linked is not well understood. Chromatin factors could be involved in linking both aging aspects, as epigenetic mechanisms bridge regulation of different biological processes. The epigenetic factor LIN-53 (RBBP4/7) associates with different chromatin-regulating complexes to safeguard cell identities in Caenorhabditis elegans as well as mammals, and has a role in preventing memory loss and premature aging in humans. We show that LIN-53 interacts with the nucleosome remodeling and deacetylase (NuRD) complex in C. elegans muscles to ensure functional muscles during postembryonic development and in adults. While mutants for other NuRD members show a normal lifespan, animals lacking LIN-53 die early because LIN-53 depletion affects also the histone deacetylase complex Sin3, which is required for a normal lifespan. To determine why lin-53 and sin-3 mutants die early, we performed transcriptome and metabolomic analysis revealing that levels of the disaccharide trehalose are significantly decreased in both mutants. As trehalose is required for normal lifespan in C. elegans, lin-53 and sin-3 mutants could be rescued by either feeding with trehalose or increasing trehalose levels via the insulin/IGF1 signaling pathway. Overall, our findings suggest that LIN-53 is required for maintaining lifespan and muscle integrity through discrete chromatin regulatory mechanisms.Since both LIN-53 and its mammalian homologs safeguard cell identities, it is conceivable that its implication in lifespan regulation is also evolutionarily conserved. K E Y W O R D Saging, Caenorhabditis elegans, chromatin, epigenetics, healthspan, metabolome S U PP O RTI N G I N FO R M ATI O NAdditional supporting information may be found online in the Supporting Information section at the end of the article. How to cite this article: Müthel S, Uyar B, He M, et al. The conserved histone chaperone LIN-53 is required for normal lifespan and maintenance of muscle integrity in Caenorhabditis elegans. Aging Cell. 2019;18:e13012. https ://doi.

59Whether extension of lifespan provides an extended time without health deteriorations is an 60 important issue for human aging. However, to which degree lifespan and healthspan 61 regulation might be linked is not well understood. Chromatin factors could be involved in 62 linking both aging aspects, as epigenetic mechanisms bridge regulation of different biological 63 processes. The epigenetic factor LIN-53 (RBBP4/7) is required for safeguarding cell 64 identities in Caenorhabditis elegans as well as mammals and for preventing memory loss 65 and premature aging in humans. LIN-53 is a histone chaperone that associates with different 66 chromatin-regulating complexes. We show that LIN-53 interacts with the Nucleosome 67 remodeling and deacteylase (NuRD)-complex in C. elegans muscles to promote healthy 68 locomotion during aging. While mutants for other NuRD members show a normal lifespan, 69 animals lacking LIN-53 die early because LIN-53 depletion affects also the Histone 70 deacetylase complex Sin3, which is required for a normal lifespan. To determine why lin-53 71 and sin-3 mutants die early, we performed transcriptome and metabolome analysis and 72 found that levels of the disaccharide Trehalose are significantly decreased in both mutants. 73As Trehalose is required for normal lifespan in C. elegans, lin-53 and sin-3 mutants could be 74 rescued by either feeding with Trehalose or increasing Trehalose levels via the Insulin/IGF1 75 signaling pathway. Overall, our findings suggest that LIN-53 is required for maintaining 76 lifespan and promoting healthspan through discrete chromatin regulatory mechanisms. Since 77 both LIN-53 and its mammalian homologs safeguard cell identities, it is conceivable that its 78 implication in lifespan and healthspan regulation is also evolutionarily conserved. 79 80 extend the healthspan, meaning the time of life without unfavorable health conditions. 87

Who wants to live forever? Lifespan extension is a long-standing human desire. And sure enough, a longer lifespan is expected to also extend the time without diseases and frailty. But should we take for granted that lifespan and healthspan are inherently linked? Meaning, does increased lifespan also extend the time without unfavorable health conditions? It is not well understood to which extend lifespan and healthspan are indeed linked. Recent studies provide increasing evidence that epigenetic factors may play a pivotal role in connecting aging and healthspan regulation. Epigenetic factors control gene expression by regulating modifications and structure of chromatin. Loss of specific chromatin regulators can cause defects and diseases such as cancer and neurodegeneration. Also, histone demethylases maintain muscle stem cells and Sirtuins with histone deacetylation activity control organismal aging [1]. Another type of chromatinregulating proteins are histone chaperones. They are required for nuclear import of histone proteins, their assembly into nucleosomes and genomic localization, as well as for the post-translational modifications of histones. Histone chaperones interact with different chromatin-regulating factors to provide gene regulatory functions with a broad spectrum of physiological functions such as cell fate safeguarding and blocking reprogramming of cell identities [2]. Recently, it was found that the histone chaperone LIN-53 in the nematode Caenorhabditis elegans (C. elegans), known as RBBP4 and RBBP7 in humans, is important for lifespan as well as healthspan regulation [3]. Interestingly, a previous study in humans revealed that RBBP4/7 are implicated in aging and age-related memory loss [4,5]. Therefore, it is conceivable that LIN-53 represents and evolutionarily conserved link of lifespan and healthspan regulation. The histone chaperone LIN-53 and its mammalian homologs RBBP4/7 (older names: CAF-1p48, RbAp46/48) are found in different chromatin-regulating complexes including Polycomb Repressive Complex 2 (PRC2, histone methyltransferase activity), CAF1 histone chaperone complex, Sin3 histone deacetylase complex (HDAC), nucleosome remodeling complex (NuRD), and DRM (Dp/Rb/Muv) complex [6]. Recently, LIN-53 and the RBBP4/7-containing CAF1 complex have been identified as reprogramming barriers in the nematode C. elegans and in mouse fibro-www.aging-us.com

Critical illness myopathy (CIM) is an acquired, devastating, multifactorial muscle-wasting disease with incomplete recovery. The impact on hospital costs and permanent loss of quality of life is enormous. Incomplete recovery might imply that the function of muscle stem cells (MuSC) is impaired. We tested whether epigenetic alterations could be in part responsible. We characterized human muscle stem cells (MuSC) isolated from early CIM and analyzed epigenetic alterations (CIM n = 15, controls n = 21) by RNA-Seq, immunofluorescence, analysis of DNA repair, and ATAC-Seq. CIM-MuSC were transplanted into immunodeficient NOG mice to assess their regenerative potential. CIM-MuSC exhibited significant growth deficits, reduced ability to differentiate into myotubes, and impaired DNA repair. The chromatin structure was damaged, as characterized by alterations in mRNA of histone 1, depletion or dislocation of core proteins of nucleosome remodeling and deacetylase complex, and loosening of multiple nucleosome-spanning sites. Functionally, CIM-MuSC had a defect in building new muscle fibers. Further, MuSC obtained from the electrically stimulated muscle of CIM patients was very similar to control MuSC, indicating the impact of muscle contraction in the onset of CIM. CIM not only affects working skeletal muscle but has a lasting and severe epigenetic impact on MuSC.

LMNA-related muscular dystrophy is an autosomal-dominant progressive disorder caused by mutations in LMNA. LMNA missense mutations are becoming correctable with CRISPR/Cas9-derived tools. Evaluating the functional recovery of LMNA after gene editing bears challenges as there is no reported direct loss of function of lamin A/C proteins in patient-derived cells. The proteins encoded by LMNA are lamins A/C, important ubiquitous nuclear envelope proteins but absent in pluripotent stem cells. We induced lamin A/C expression in induced pluripotent stem cells (iPSCs) of two patients with LMNA-related muscular dystrophy, NM_170707.4 (LMNA): c.1366A > G, p.(Asn456Asp) and c.1494G > T, p.(Trp498Cys), using a short three-day, serum-induced differentiation protocol and analyzed expression profiles of co-regulated genes, examples being COL1A2 and S100A6. We then performed precise gene editing of LMNA c.1366A > G using the near-PAMless (PAM: protospacer-adjacent motif) cytosine base editor. We show that the mutation can be repaired to 100% efficiency in individual iPSC clones. The fast differentiation protocol provided a functional readout and demonstrated increased lamin A/C expression as well as normalized expression of co-regulated genes. Collectively, our findings demonstrate the power of CRISPR/Cas9-mediated gene correction and effective outcome measures in a disease with, so far, little perspective on therapies.