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International Journal of Molecular... Feb 2024Fibrosis represents a process characterized by excessive deposition of extracellular matrix (ECM) proteins. It often represents the evolution of pathological conditions,... (Review)
Review
Fibrosis represents a process characterized by excessive deposition of extracellular matrix (ECM) proteins. It often represents the evolution of pathological conditions, causes organ failure, and can, in extreme cases, compromise the functionality of organs to the point of causing death. In recent years, considerable efforts have been made to understand the molecular mechanisms underlying fibrotic evolution and to identify possible therapeutic strategies. Great interest has been aroused by the discovery of a molecular association between epithelial to mesenchymal plasticity (EMP), in particular epithelial to mesenchymal transition (EMT), and fibrogenesis, which has led to the identification of complex molecular mechanisms closely interconnected with each other, which could explain EMT-dependent fibrosis. However, the result remains unsatisfactory from a therapeutic point of view. In recent years, advances in epigenetics, based on chromatin remodeling through various histone modifications or through the intervention of non-coding RNAs (ncRNAs), have provided more information on the fibrotic process, and this could represent a promising path forward for the identification of innovative therapeutic strategies for organ fibrosis. In this review, we summarize current research on epigenetic mechanisms involved in organ fibrosis, with a focus on epigenetic regulation of EMP/EMT-dependent fibrosis.
Topics: Humans; Epigenesis, Genetic; DNA Methylation; Epithelial-Mesenchymal Transition; Fibrosis; Chromatin Assembly and Disassembly
PubMed: 38474021
DOI: 10.3390/ijms25052775 -
MBio Apr 2024The STimulator of INterferon Genes (STING) constitutes a major DNA-sensing pathway that restricts HSV-1 infection in different models by activating type I interferon and...
The STimulator of INterferon Genes (STING) constitutes a major DNA-sensing pathway that restricts HSV-1 infection in different models by activating type I interferon and pro-inflammatory responses. To counteract STING, HSV-1 has evolved numerous strategies including mechanisms to interfere with its oligomerization, post-translational modifications, and downstream signaling. Previously, we demonstrated that STING is packaged in extracellular vesicles (EVs) produced from HSV-1-infected cells. These EVs activated antiviral responses in uninfected recipient cells and suppressed a subsequent HSV-1 infection in a STING-dependent manner. Here, we provide information on the packaging of STING in EVs and its exocytosis. We found that STING exocytosis did not occur in CD63 knockdown cells supporting that STING follows the CD63 exocytosis pathway. Consistently, we found that STING co-localized with CD63 in cytoplasmic globular structures and exosomal STING and CD63 co-fractionated. Both golgicide A and brefeldin A prevented STING exocytosis during HSV-1 infection suggesting that STING trafficking through the Golgi is required. A STING ligand was insufficient for STING exocytosis, and downstream signaling through TBK1 was not required. However, STING palmitoylation and tethering to the ER by STIM1 were required for STING exocytosis. Finally, we found that HSV-1 replication/late gene expression triggered CD63 exocytosis that was required for STING exocytosis. Surprisingly, HSV-2 strain G did not trigger CD63 or STING exocytosis as opposed to VZV and HCMV. Also, EVs from HSV-1(F)- and HSV-2(G)-infected cells displayed differences in their ability to restrict these viruses. Overall, STING exocytosis is induced by certain viruses and shapes the microenvironment of infection.IMPORTANCEExtracellular vesicles (EVs) are released by all types of cells as they constitute a major mechanism of intercellular communication. The packaging of specific cargo in EVs and the pathway of exocytosis are not fully understood. STING is a sensor of a broad spectrum of pathogens and a key component of innate immunity. STING exocytosis during HSV-1 infection has been an intriguing observation, raising questions of whether this is a virus-induced process, the purpose it serves, and whether it is observed after infection with other viruses. Here, we have provided insights into the pathway of STING exocytosis and determined factors involved. STING exocytosis is a virus-induced process and not a response of the host to the infection. Besides HSV-1, other herpes viruses triggered STING exocytosis, but HSV-2(G) did not. HSV-1 EVs displayed different restriction capabilities compared with HSV-2(G) EVs. Overall, STING exocytosis is triggered by viruses to shape the microenvironment of infection.
Topics: Humans; Exocytosis; Herpes Simplex; Herpesvirus 1, Human; Immunity, Innate; Membrane Proteins
PubMed: 38470056
DOI: 10.1128/mbio.00373-24 -
Nucleus (Austin, Tex.) Dec 2024Cell migration involves the actin cytoskeleton, and recently recognized nuclear involvement. In this study, we explore the impact of chromatin remodeling on cell...
Cell migration involves the actin cytoskeleton, and recently recognized nuclear involvement. In this study, we explore the impact of chromatin remodeling on cell migration using NIH 3T3 cells and a scratch wound assay subjected to pharmacological interventions. We inhibit histone deacetylases (HDACs) with Trichostatin A (TSA) and methyltransferase EZH2 with GSK126 to modulate chromatin compaction. Our results indicate that chromatin modifications impair wound closure efficiency, reduce individual cell migration speed, and disrupt migration persistence. Live-cell imaging reveals dynamic intranuclear chromatin remodeling and nuclear shape parameters during migration, influenced by both small- and large-scale chromatin remodeling. The altered nuclear shape is associated with disrupted cell and nuclear mechanics, suggesting a crucial interplay between chromatin remodeling, nuclear mechanics and migration. These findings shed light on the intricate connection between intranuclear chromatin dynamics, nuclear mechanics, and cell migration, providing a basis for further investigations into the molecular mechanisms governing these processes.
Topics: Mice; Animals; Chromatin; Chromatin Assembly and Disassembly; Cell Movement
PubMed: 38465796
DOI: 10.1080/19491034.2024.2325961 -
Molecular Therapy. Methods & Clinical... Mar 2024The unique palindromic inverted terminal repeats (ITRs) and single-stranded nature of adeno-associated virus (AAV) DNA are major hurdles to current sequencing...
The unique palindromic inverted terminal repeats (ITRs) and single-stranded nature of adeno-associated virus (AAV) DNA are major hurdles to current sequencing technologies. Due to these characteristics, sequencing noncanonical AAV genomes present in AAV vector preparations remains challenging. To address this limitation, we developed thorough molecule configuration analysis of noncanonical AAV genomes (TMCA-AAV-seq). TMCA-AAV-seq takes advantage of the documented AAV packaging mechanism in which encapsidation initiates from its 3' ITR, for AAV-seq library construction. Any AAV genome with a 3' ITR is converted to a template suitable to adapter addition by a Bst DNA polymerase-mediated extension reaction. This extension reaction helps fix ITR heterogeneity in the AAV population and allows efficient adapter addition to even noncanonical AAV genomes. The resulting library maintains the original AAV genome configurations without introducing undesired changes. Subsequently, long-read sequencing can be performed by the Pacific Biosciences (PacBio) single-molecule, real-time (SMRT) sequencing technology platform. Finally, through comprehensive data analysis, we can recover canonical, noncanonical AAV DNA, and non-AAV vector DNA sequences, along with their molecular configurations. Our method is a robust tool for profiling thorough AAV-population genomes. TMCA-AAVseq can be further extended to all parvoviruses and their derivative vectors.
PubMed: 38463141
DOI: 10.1016/j.omtm.2024.101215 -
Biochemical Society Transactions Apr 2024Eukaryotic genomes are compacted and organized into distinct three-dimensional (3D) structures, which range from small-scale nucleosome arrays to large-scale chromatin... (Review)
Review
Eukaryotic genomes are compacted and organized into distinct three-dimensional (3D) structures, which range from small-scale nucleosome arrays to large-scale chromatin domains. These chromatin structures play an important role in the regulation of transcription and other nuclear processes. The molecular mechanisms that drive the formation of chromatin structures across scales and the relationship between chromatin structure and function remain incompletely understood. Because the processes involved are complex and interconnected, it is often challenging to dissect the underlying principles in the nuclear environment. Therefore, in vitro reconstitution systems provide a valuable approach to gain insight into the molecular mechanisms by which chromatin structures are formed and to determine the cause-consequence relationships between the processes involved. In this review, we give an overview of in vitro approaches that have been used to study chromatin structures across scales and how they have increased our understanding of the formation and function of these structures. We start by discussing in vitro studies that have given insight into the mechanisms of nucleosome positioning. Next, we discuss recent efforts to reconstitute larger-scale chromatin domains and loops and the resulting insights into the principles of genome organization. We conclude with an outlook on potential future applications of chromatin reconstitution systems and how they may contribute to answering open questions concerning chromatin architecture.
Topics: Nucleosomes; Chromatin; Genome; Humans; Chromatin Assembly and Disassembly; Animals
PubMed: 38451192
DOI: 10.1042/BST20230883 -
Nature Mar 2024DNA and histone modifications combine into characteristic patterns that demarcate functional regions of the genome. While many 'readers' of individual modifications have...
DNA and histone modifications combine into characteristic patterns that demarcate functional regions of the genome. While many 'readers' of individual modifications have been described, how chromatin states comprising composite modification signatures, histone variants and internucleosomal linker DNA are interpreted is a major open question. Here we use a multidimensional proteomics strategy to systematically examine the interaction of around 2,000 nuclear proteins with over 80 modified dinucleosomes representing promoter, enhancer and heterochromatin states. By deconvoluting complex nucleosome-binding profiles into networks of co-regulated proteins and distinct nucleosomal features driving protein recruitment or exclusion, we show comprehensively how chromatin states are decoded by chromatin readers. We find highly distinctive binding responses to different features, many factors that recognize multiple features, and that nucleosomal modifications and linker DNA operate largely independently in regulating protein binding to chromatin. Our online resource, the Modification Atlas of Regulation by Chromatin States (MARCS), provides in-depth analysis tools to engage with our results and advance the discovery of fundamental principles of genome regulation by chromatin states.
Topics: Humans; Binding Sites; Chromatin; DNA; Enhancer Elements, Genetic; Heterochromatin; Histones; Nuclear Proteins; Nucleosomes; Promoter Regions, Genetic; Protein Binding; Proteomics; Chromatin Assembly and Disassembly
PubMed: 38448585
DOI: 10.1038/s41586-024-07141-5 -
Nature Communications Mar 2024Artificial biomolecular condensates are emerging as a versatile approach to organize molecular targets and reactions without the need for lipid membranes. Here we ask...
Artificial biomolecular condensates are emerging as a versatile approach to organize molecular targets and reactions without the need for lipid membranes. Here we ask whether the temporal response of artificial condensates can be controlled via designed chemical reactions. We address this general question by considering a model problem in which a phase separating component participates in reactions that dynamically activate or deactivate its ability to self-attract. Through a theoretical model we illustrate the transient and equilibrium effects of reactions, linking condensate response and reaction parameters. We experimentally realize our model problem using star-shaped DNA motifs known as nanostars to generate condensates, and we take advantage of strand invasion and displacement reactions to kinetically control the capacity of nanostars to interact. We demonstrate reversible dissolution and growth of DNA condensates in the presence of specific DNA inputs, and we characterize the role of toehold domains, nanostar size, and nanostar valency. Our results will support the development of artificial biomolecular condensates that can adapt to environmental changes with prescribed temporal dynamics.
Topics: DNA Packaging; Biomolecular Condensates; DNA Replication; Gene Conversion; Nucleotide Motifs
PubMed: 38429336
DOI: 10.1038/s41467-024-46266-z -
Molecular Genetics and Metabolism May 2024The Mendelian disorders of chromatin machinery (MDCMs) represent a distinct subgroup of disorders that present with neurodevelopmental disability. The chromatin... (Review)
Review
The Mendelian disorders of chromatin machinery (MDCMs) represent a distinct subgroup of disorders that present with neurodevelopmental disability. The chromatin machinery regulates gene expression by a range of mechanisms, including by post-translational modification of histones, responding to histone marks, and remodelling nucleosomes. Some of the MDCMs that impact on histone modification may have potential therapeutic interventions. Two potential treatment strategies are to enhance the intracellular pool of metabolites that can act as substrates for histone modifiers and the use of medications that may inhibit or promote the modification of histone residues to influence gene expression. In this article we discuss the influence and potential treatments of histone modifications involving histone acetylation and histone methylation. Genomic technologies are facilitating earlier diagnosis of many Mendelian disorders, providing potential opportunities for early treatment from infancy. This has parallels with how inborn errors of metabolism have been afforded early treatment with newborn screening. Before this promise can be fulfilled, we require greater understanding of the biochemical fingerprint of these conditions, which may provide opportunities to supplement metabolites that can act as substrates for chromatin modifying enzymes. Importantly, understanding the metabolomic profile of affected individuals may also provide disorder-specific biomarkers that will be critical for demonstrating efficacy of treatment, as treatment response may not be able to be accurately assessed by clinical measures.
Topics: Humans; Chromatin; Metabolic Networks and Pathways; Histones; Protein Processing, Post-Translational; Acetylation; Metabolism, Inborn Errors; Chromatin Assembly and Disassembly; Genetic Diseases, Inborn; Infant, Newborn; Methylation
PubMed: 38428378
DOI: 10.1016/j.ymgme.2024.108360 -
Cell Reports Mar 2024SWI/SNF complexes are evolutionarily conserved, ATP-dependent chromatin remodeling machines. Here, we characterize the features of SWI/SNF-dependent genes using BRM014,...
SWI/SNF complexes are evolutionarily conserved, ATP-dependent chromatin remodeling machines. Here, we characterize the features of SWI/SNF-dependent genes using BRM014, an inhibitor of the ATPase activity of the complexes. We find that SWI/SNF activity is required to maintain chromatin accessibility and nucleosome occupancy for most enhancers but not for most promoters. SWI/SNF activity is needed for expression of genes with low to medium levels of expression that have promoters with (1) low chromatin accessibility, (2) low levels of active histone marks, (3) high H3K4me1/H3K4me3 ratio, (4) low nucleosomal phasing, and (5) enrichment in TATA-box motifs. These promoters are mostly occupied by the canonical Brahma-related gene 1/Brahma-associated factor (BAF) complex. These genes are surrounded by SWI/SNF-dependent enhancers and mainly encode signal transduction, developmental, and cell identity genes (with almost no housekeeping genes). Machine-learning models trained with different chromatin characteristics of promoters and their surrounding regulatory regions indicate that the chromatin landscape is a determinant for establishing SWI/SNF dependency.
Topics: Chromatin; Transcription Factors; Nucleosomes; Chromatin Assembly and Disassembly
PubMed: 38427563
DOI: 10.1016/j.celrep.2024.113855 -
The Journal of Clinical Investigation Mar 2024Early gestational loss occurs in approximately 20% of all clinically recognized human pregnancies and is an important cause of morbidity. Either embryonic or maternal...
Early gestational loss occurs in approximately 20% of all clinically recognized human pregnancies and is an important cause of morbidity. Either embryonic or maternal defects can cause loss, but a functioning and receptive uterine endometrium is crucial for embryo implantation. We report that the switch/sucrose nonfermentable (SWI/SNF) remodeling complex containing polybromo-1 (PBRM1) and Brahma-related gene 1 (BRG1) is essential for implantation of the embryonic blastocyst on the wall of the uterus in mice. Although preimplantation development is unaffected, conditional ablation of Pbrm1 in uterine stromal cells disrupts progesterone pathways and uterine receptivity. Heart and neural crest derivatives expressed 2 (Hand2) encodes a basic helix-loop-helix (bHLH) transcription factor required for embryo implantation. We identify an enhancer of the Hand2 gene in stromal cells that requires PBRM1 for epigenetic histone modifications/coactivator recruitment and looping with the promoter. In Pbrm1cKO mice, perturbation of chromatin assembly at the promoter and enhancer sites compromises Hand2 transcription, adversely affects fibroblast growth factor signaling pathways, prevents normal stromal-epithelial crosstalk, and disrupts embryo implantation. The mutant female mice are infertile and provide insight into potential causes of early pregnancy loss in humans.
Topics: Animals; Female; Humans; Mice; Pregnancy; Basic Helix-Loop-Helix Transcription Factors; Chromatin; Chromatin Assembly and Disassembly; Embryo Implantation; Transcription Factors; Uterus
PubMed: 38426493
DOI: 10.1172/JCI174194