A CRISPR-Cas9 based shuffle system for endogenous histone H3 and H4 combinatorial mutagenesis

Fu, Yu; Zhu, Zhenglin; Meng, Geng; Zhang, Rijun; Zhang, Yueping

doi:10.1038/s41598-021-82774-4

Cited by 10 publications

(7 citation statements)

References 25 publications

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“…An alternative approach is to test for the functional consequences of modifying the histones themselves. To achieve this, CRISPR-Cas9 has been used in S. cerevisiae to generate yeast strains with different combinations of mutations at histone tail lysines for histone H3 and H4, allowing the investigators to assess the effects of loss of different combinations of histone marks (Fu et al, 2021). In Trypanosoma brucei, precise editing of genes in multicopy arrays was performed with CRISPR-Cas9, allowing for the replacement of histone H4K4 with H4R4 to mimic the constitutively non-acetylated state.…”

Section: Gaps In the Field And Future Studiesmentioning

confidence: 99%

Is There a Histone Code for Cellular Quiescence?

Bonitto

Sarathy

Atai

et al. 2021

Front. Cell Dev. Biol.

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Many of the cells in our bodies are quiescent, that is, temporarily not dividing. Under certain physiological conditions such as during tissue repair and maintenance, quiescent cells receive the appropriate stimulus and are induced to enter the cell cycle. The ability of cells to successfully transition into and out of a quiescent state is crucial for many biological processes including wound healing, stem cell maintenance, and immunological responses. Across species and tissues, transcriptional, epigenetic, and chromosomal changes associated with the transition between proliferation and quiescence have been analyzed, and some consistent changes associated with quiescence have been identified. Histone modifications have been shown to play a role in chromatin packing and accessibility, nucleosome mobility, gene expression, and chromosome arrangement. In this review, we critically evaluate the role of different histone marks in these processes during quiescence entry and exit. We consider different model systems for quiescence, each of the most frequently monitored candidate histone marks, and the role of their writers, erasers and readers. We highlight data that support these marks contributing to the changes observed with quiescence. We specifically ask whether there is a quiescence histone “code,” a mechanism whereby the language encoded by specific combinations of histone marks is read and relayed downstream to modulate cell state and function. We conclude by highlighting emerging technologies that can be applied to gain greater insight into the role of a histone code for quiescence.

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Section: Gaps In the Field And Future Studiesmentioning

confidence: 99%

Is There a Histone Code for Cellular Quiescence?

Bonitto

Sarathy

Atai

et al. 2021

Front. Cell Dev. Biol.

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“…By coupling dCAS9 with TET proteins, 5-methylcytosine can be oxidized to 5-hydroxymethylcytosine, leading to DNA demethylation marker alterations. 39…”

Section: Discussionmentioning

confidence: 99%

CRISPR–Cas9 Gene Editing: Curing Genetic Diseases by Inherited Epigenetic Modifications

Kolanu

2024

Glob Med Genet

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Introduction CRISPR–Cas9 gene editing, leveraging bacterial defense mechanisms, offers precise DNA modifications, holding promise in curing genetic diseases. This review critically assesses its potential, analyzing evidence on therapeutic applications, challenges, and future prospects. Examining diverse genetic disorders, it evaluates efficacy, safety, and limitations, emphasizing the need for a thorough understanding among medical professionals and researchers. Acknowledging its transformative impact, a systematic review is crucial for informed decision-making, responsible utilization, and guiding future research to unlock CRISPR–Cas9's full potential in realizing the cure for genetic diseases. Methods A comprehensive literature search across PubMed, Scopus, and the Web of Science identified studies applying CRISPR–Cas9 gene editing for genetic diseases, following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. Inclusion criteria covered in vitro and in vivo models targeting various genetic diseases with reported outcomes on disease modification or potential cure. Quality assessment revealed a generally moderate to high risk of bias. Heterogeneity prevented quantitative meta-analysis, prompting a narrative synthesis of findings. Discussion CRISPR–Cas9 enables precise gene editing, correcting disease-causing mutations and offering hope for previously incurable genetic conditions. Leveraging inherited epigenetic modifications, it not only fixes mutations but also restores normal gene function and controls gene expression. The transformative potential of CRISPR–Cas9 holds promise for personalized treatments, improving therapeutic outcomes, but ethical considerations and safety concerns must be rigorously addressed to ensure responsible and safe application, especially in germline editing with potential long-term implications.

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“…A major advantage of removing endogenous histones in this strategy is that it increases the likelihood of detecting phenotypes associated with expression of histone mutants, which may otherwise be masked if competing wild-type histones are also present. Systematic mutagenesis experiments with core histones were initially conducted in Saccharomyces cerevisiae to reveal many new insights into histone function (Dai et al, 2008;Fu et al, 2021;Govin et al, 2010;Nakanishi et al, 2008). These experiments utilized histone shuffle systems to either provide an episomal plasmid expressing histone mutants in a background with the endogenous histone genes deleted, or to directly mutate the endogenous histone copies using homologous recombination.…”

Section: Introductionmentioning

confidence: 99%

“…These experiments utilized histone shuffle systems to either provide an episomal plasmid expressing histone mutants in a background with the endogenous histone genes deleted, or to directly mutate the endogenous histone copies using homologous recombination. The most recent system developed in S. cerevisiae utilized an efficient CRISPR/Cas9-based histone shuffle strategy that allows for the rapid development of multiplex histone mutations (Fu et al, 2021). In multicellular eukaryotes, Drosophila melanogaster is the first organism in which systematic histone mutagenesis was performed, and these systems used site-specific transgenesis to replace the endogenous histone coding region with that of a modified histone gene or histone array (Gunesdogan et al, 2010;Basler, 2009, 2012;McKay et al, 2015).…”

Section: Introductionmentioning

confidence: 99%

“…In contrast to the aforementioned biological model systems, plant systems present additional challenges to implementing complete histone gene replacement. While all replication-dependent histone genes are clustered at a single genomic locus in D. melanogaster (Lifton et al, 1978), and S. cerevisiae contains only two copies of each core histone gene at the haploid cell stage (Fu et al, 2021), there are 47 genes encoding H2A, H2B, H3, and H4 in Arabidopsis thaliana found dispersed throughout the genome (Tenea et al, 2009). Because the histone genes are not clustered together in plants, the establishment of complex histone deletion mutants necessary for partially or completely replacing endogenous histone genes with modified histone genes is more challenging.…”

Section: Introductionmentioning

confidence: 99%

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Systematic histone H4 replacement in Arabidopsis thaliana reveals a role for H4R17 in regulating flowering time

et al. 2022

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Despite the broad array of roles for epigenetic mechanisms on regulating diverse processes in eukaryotes, no experimental system is currently available in plants for the direct assessment of histone function. In this work, we present the development of a genetic strategy in Arabidopsis (Arabidopsis thaliana) whereby modified histone H4 transgenes can completely replace the expression of endogenous histone H4 genes. Accordingly, we established a collection of plants expressing different H4 point mutants targeting residues that may be post-translationally modified in vivo. To demonstrate its utility, we screened this new H4 mutant collection to uncover substitutions in H4 that alter flowering time. We identified different mutations in the H4 tail (H4R17A) and the H4 globular domain (H4R36A, H4R39K, H4R39A, and H4K44A) that strongly accelerate the floral transition. Furthermore, we identified a conserved regulatory relationship between H4R17 and the ISWI chromatin remodeling complex in plants: As with other biological systems, H4R17 regulates nucleosome spacing via ISWI. Overall, this work provides a large set of H4 mutants to the plant epigenetics community that can be used to systematically assess histone H4 function in Arabidopsis and a roadmap to replicate this strategy for studying other histone proteins in plants.

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A CRISPR-Cas9 based shuffle system for endogenous histone H3 and H4 combinatorial mutagenesis

Cited by 10 publications

References 25 publications

Is There a Histone Code for Cellular Quiescence?

Is There a Histone Code for Cellular Quiescence?

CRISPR–Cas9 Gene Editing: Curing Genetic Diseases by Inherited Epigenetic Modifications

Systematic histone H4 replacement in Arabidopsis thaliana reveals a role for H4R17 in regulating flowering time

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