Control Of Gene Activity Research Articles

Current Updates on the Understanding of the Role of DNA Methylation on Obesity.

Obesity is a condition in which there is an accumulation of excess body fat leading to a weight far above the normal range that poses significant health risks. According to WHO, 8 billion people in the world were obese in 2022. Consequently, obesity has become a pandemic with negative impacts on both global health and economies. Obesity is influenced by various factors including environmental influences, lifestyle choices, gut microbiota, genetic factors, and epigenetic mechanisms such as DNA methylation. DNA methylation can affect an individual's phenotype and condition without altering their DNA sequence. It is the most extensively studied epigenetic alteration and it plays an important part in controlling gene activity associated with obesity. Numerous studies have indicated that DNA methylation is implicated in obesity, thus this review aims to elaborate the roles of DNA methylation to inform the development of preventive measures for obesity.

KIAA1429 regulates lung adenocarcinoma proliferation and metastasis through the PI3K/AKT pathway by modulating ARHGAP30 expression.

Alterations in epigenetic factors are recognized as key contributors to the emergence of human cancer. The active and reversible alteration of N6-methyladenosine (m6A) RNA is crucial for controlling gene activity and determining cellular destiny. Even with these insights, the triggering of KIAA1429 (also called VIRMA) and its role in lung adenocarcinoma (LUAD) is mostly unclear. As a result, the objective of this study was to elucidate how KIAA1429 contributes to cancer development in LUAD. This study utilized multiple methods for investigation, encompassing the in vitro functional examination of KIAA1429 in lung adenocarcinoma cells, transcriptome sequencing, methylation RNA immunoprecipitation sequencing (MeRIP-seq), as well as RNA stability tests to ascertain the half-life and stability of the target genes. The results indicated that modifying the expression of KIAA1429 regulated the proliferation and metastasis of LUAD. By employing transcriptome sequencing alongside MeRIP-seq analysis, the research pinpointed genes affected by m6A alterations triggered by KIAA1429. In a more detailed manner, it was discovered that KIAA1429 plays a regulatory role in the expression of ARHGAP30. Suppressing KIAA1429 results in reduced m6A levels in the mRNA of the target gene ARHGAP30, boosting its stability and expression, thus inhibiting tumor proliferation and metastasis. This study revealed the activation mechanism and pivotal function of KIAA1429 in LUAD tumor development, paving the way for molecular-based interventions for LUAD.

P073 Functional investigation of Osteoarthritis-associated non-coding variants using CRISPR/Cas9-mediated perturbation of active enhancer regions

Abstract Background/Aims OA is a common, complex disease, affecting millions of people worldwide with no effective treatments. Recent studies have implicated over 100 regions of the genome that carry DNA changes that increase the risk of developing OA. Although these findings have advanced our understanding of the disease, implicating biological pathways and cell types as being important in disease, there is still much work required to understand the mechanism by which risk variants cause OA and fully translate the findings. This is primarily due to the nature of these DNA risk variants, often located outside gene coding regions, in regions that control gene activity. Here we use CRISPR technology to investigate two DNA regions implicated in OA risk to determine the target genes that are regulated and the likely cell types in which they operate. Methods We annotated OA associated SNPs utilising in-house ATAC-seq for open chromatin regions and CUT&Tag for H3K27ac marks generated for the first time here in two chondrocyte cell lines (TC28a2 and SW1353), along with H3K27ac marks from primary chondrocytes derived from the ENCODE database. Additionally we focused on regions which had genes expressed in chondrocytes within 1Mb of the lead SNP. Together this information guided prioritisation of OA regions and residing SNPs for functional validation. The two chondrocyte cell lines were engineered to constitutively express either CRISPR activation (dCas9-VPR) or CRISPR inhibition (dCas9-MeCP2-KRAB) machinery and subsequently designed guide RNAs targeted the prioritised SNPs. This enabled the activation or repression of regulatory regions, with the resultant effect on gene expression of potential target genes measured through qPCR. Results Through functional annotation of OA-associated regions, we focused on two SNPs prioritised across two distinct regions: rs1149611 in the TSKU region and rs3795159 in the UQCC1 region. These SNPs coincided with active chromatin marks in the chondrocyte cell lines and were targeted by CRISPR activation or inhibition systems in both chondrocyte cell lines. Both activation and inhibition of rs3795159 in the SW1353 chondrocyte cell line resulted in significant differential expression of the UQCC1 gene. Similarly, activation of rs1149611 upregulated TSKU in this cell line, whilst deletion of the SNP region downregulated TSKU expression in the Tc28a cell line. UQCC1 is a fibroblast growth factor, and TSKU is an inhibitor of TGFB1, and has been implicated in bone growth. Conclusion Although these results should be replicated in primary chondrocytes, here we determine firm evidence that DNA regions containing OA risk SNPs can regulate UQCC1 and TSKU in cell lines. This implicates these genes, and biological pathways, being perturbed prior to disease onset, and as such constitute novel targets for OA intervention. Disclosure A. Frantzeskos: None. J. Srinivasan: None. S. Morris: None. C. Shi: None. A. McGovern: None. E. Richards: None. S. Rossi: None. S. Eyre: None. G. Orozco: None.

New molecular aspects of the pathogenesis of osteoporosis – perspectives for early diagnosis and treatment

Osteoporosis (OP) is a common disease leading to low-trauma fractures and is a serious medical and social problem. Often a fracture is the first clinical manifestation of OP that has been asymptomatic for a long time, necessitating the development of methods for early detection and risk assessment of this disease. OP is a multifactorial disease with a strong hereditary component. However, as the data from the study of genetic factors show, only 15% of the heritability of this trait can be explained. In this context, the focus of research is shifting to the area of epigenetic regulation, which controls gene activity without altering the primary structure of the DNA. One of the most promising mechanisms of epigenetic control is methylation, which affects DNA as well as RNA and histones. The characteristics of these mechanisms and the possibilities of their use for the diagnosis and treatment of OP are presented in this review.

Optogenetic control of YAP reveals a dynamic communication code for stem cell fate and proliferation

YAP is a transcriptional regulator that controls pluripotency, cell fate, and proliferation. How cells ensure the selective activation of YAP effector genes is unknown. This knowledge is essential to rationally control cellular decision-making. Here we leverage optogenetics, live-imaging of transcription, and cell fate analysis to understand and control gene activation and cell behavior. We reveal that cells decode the steady-state concentrations and timing of YAP activation to control proliferation, cell fate, and expression of the pluripotency regulators Oct4 and Nanog. While oscillatory YAP inputs induce Oct4 expression and proliferation optimally at frequencies that mimic native dynamics, cellular differentiation requires persistently low YAP levels. We identify the molecular logic of the Oct4 dynamic decoder, which acts through an adaptive change sensor. Our work reveals how YAP levels and dynamics enable multiplexing of information transmission for the regulation of developmental decision-making and establishes a platform for the rational control of these behaviors.

Advances in epigenetic modifications of autophagic process in pulmonary hypertension.

Pulmonary hypertension is characterized by pulmonary arterial remodeling that results in increased pulmonary vascular resistance, right ventricular failure, and premature death. It is a threat to public health globally. Autophagy, as a highly conserved self-digestion process, plays crucial roles with autophagy-related (ATG) proteins in various diseases. The components of autophagy in the cytoplasm have been studied for decades and multiple studies have provided evidence of the importance of autophagic dysfunction in pulmonary hypertension. The status of autophagy plays a dynamic suppressive or promotive role in different contexts and stages of pulmonary hypertension development. Although the components of autophagy have been well studied, the molecular basis for the epigenetic regulation of autophagy is less understood and has drawn increasing attention in recent years. Epigenetic mechanisms include histone modifications, chromatin modifications, DNA methylation, RNA alternative splicing, and non-coding RNAs, which control gene activity and the development of an organism. In this review, we summarize the current research progress on epigenetic modifications in the autophagic process, which have the potential to be crucial and powerful therapeutic targets against the autophagic process in pulmonary hypertension development.

Estimating gene action and combining ability for yield and fiber quality in cotton (Gossypium hirsutum L.)

Genetic evaluation of segregating populations is one of the primary activities of cotton breeders. Seven cotton genotypes were crossed in a line ? tester mating design. The population of 12 F1, 12 F2 and their parents were tested in Randomized Complete Block Design with three replications at the Nazilli Cotton Research Institute of T?rkiye during the 2014/2015 cotton-growing season. The genetic variability was significant for all characters studied. ?2GCA/?2SCA indicated non-additive gene action in controlling seed cotton yield (F2), ginning out-turn (F1) and fiber quality traits for both generations. The highly favourable GCA effects for seed cotton yield and fiber strength were recorded in the Carmen cultivar. Carmen ? Carisma hybrid exhibited high mean performance and SCA effects for seed cotton yield and fiber strength, while the performance of Gloria ? ST-468 was superior in terms of ginning out-turn. We concluded that individual plants with desired traits should be selected in later generations due to the preponderance of non-additive gene actions.

Developmental cell fate choice in neural tube progenitors employs two distinct cis-regulatory strategies.

In many developing tissues, the patterns of gene expression that assign cell fate are organized by graded secreted signals. Cis-regulatory elements (CREs) interpret these signals to control gene expression, but how this is accomplished remains poorly understood. In the neural tube, a gradient of the morphogen sonic hedgehog (Shh) patterns neural progenitors. We identify two distinct ways in which CREs translate graded Shh into differential gene expression in mouse neural progenitors. In most progenitors, a common set of CREs control gene activity by integrating cell-type-specific inputs. By contrast, the most ventral progenitors use a unique set of CREs, established by the pioneer factor FOXA2. This parallels the role of FOXA2 in endoderm, where FOXA2 binds some of the same sites. Together, the data identify distinct cis-regulatory strategies for the interpretation of morphogen signaling and raise the possibility of an evolutionarily conserved role for FOXA2 across tissues.

Shaping Hox gene activity to generate morphological diversity across vertebrate phylogeny.

The importance of Hox genes for the development and evolution of the vertebrate axial skeleton and paired appendages has been recognized for already several decades. The steady growth of genomic sequence data from an increasing number of vertebrate species, together with the improvement of methods to analyze genomic structure and interactions, as well as to control gene activity in various species has refined our understanding of Hox gene activity in development and evolution. Here, I will review recent data addressing the influence of Hox regulatory processes in the evolution of the fins and the emergence of the tetrapod limb. In addition, I will discuss the involvement of posterior Hox genes in the control of vertebrate axial extension, focusing on an apparently divergent activity that Hox13 paralog group genes have on the regulation of tail bud development in mouse and zebrafish embryos.

Transcriptional and post-transcriptional mechanisms that regulate the genetic program in Zika virus-infected macrophages.

Besides our understanding of the effects of ZIKA virus (ZIKV) infection on neural progenitors' cells the pathogenesis of this RNA virus also involves antigen-presenting cells, including macrophages. However, the molecular mechanisms that control gene activation and repression associated with the macrophage response to acute ZIKV infection are not fully understood. We approached the issue by RNA-seq and miRNA-seq datasets to understand the genetic program of ZIKV-infected macrophages. Results indicate that macrophage activates a regulatory program, involving 1067 differentially expressed genes. These genetic programs induced an inflammatory response mediated by chemokines as well as an interferon-independent anti-viral response, presumptively activated by IL-27. Additionally, the pathogenetic process involves changes in other signaling pathways such as cellular stress, cell signaling, metabolism, and cell differentiation. Furthermore, transcriptional control analysis revealed regulatory functions of key transcription factors principally, NFκB and STAT1, as well as HIF1A, ETV7, and PRMD1 that are associated with metabolic reprogramming during viral infection. We also noted six long-noncoding RNAs (lncRNAs) that may act in the regulation of gene expression, including MROCKI and ZC2HC1A-2, that are involved in the inflammatory response and expression of the cytokines, respectively. On the other hand, post-transcriptional control by miRNAs, including miR-155-5p and miR-146a-5p, are associated with modulation of genes related to inflammatory and antiviral responses. Relevant to the post-transcriptional control, our data unveiled the role of RNA binding proteins that have diverse functions such as ribonucleases (PNPT1, ZC3H12A, and ZC3HAV1), splicing factors (SSB, RBM11, and RAVER2), and RNA modifiers (PARP10 and PARP14). Overall, the results establish an unbiased approach to discerning the wiring of a regulatory mechanism controlling the genetic program in ZIKV-infected macrophages.

Evaluating Histone Acetylation in Mouse Hematopoietic Stem and Progenitor Cells Using Chromatin Immunoprecipitation.

Epigenetics is the study of how cells control gene activity without changing the DNA sequence. Various epigenetic processes have been identified, including methylation, acetylation, phosphorylation, and ubiquitylation. Epigenetic processes are natural and essential to cell functions; however, when they occur improperly or at the wrong time, adverse effects can occur. A significant epigenetic process is chromatin modification. Chromatin-DNA complexes can be modified by acetylation, altering chromatin structure to influence gene expression. Stresses to hematopoietic stem and progenitor cells, such as ionizing radiation and aging, have significant effects on genomic function. Understanding epigenetic regulation in hematopoietic cells, particularly under stress, offers the potential for therapeutic intervention. We have utilized Chromatin immunoprecipitation (ChIP) in HSPCs to understand epigenetic regulation in response to ionizing radiation. This technique can be applied reliably to rare hematopoietic cells and offers a powerful tool to explore epigenetic regulation in HSPCs.

Characterization of demethylating DNA glycosylase ROS1 from Nicotiana tabacum L.

One of the main mechanisms of epigenetic regulation in higher eukaryotes is based on the methylation of cytosine at the C5 position with the formation of 5-methylcytosine (mC), which is further recognized by regulatory proteins. In mammals, methylation mainly occurs in CG dinucleotides, while in plants it targets CG, CHG, and CHH sequences (H is any base but G). Correct maintenance of the DNA methylation status is based on the balance of methylation, passive demethylation, and active demethylation. While in mammals active demethylation is based on targeted regulated damage to mC in DNA followed by the action of repair enzymes, demethylation in plants is performed by specialized DNA glycosylases that hydrolyze the N-glycosidic bond of mC nucleotides. The genome of the model plant Arabidopsis thaliana encodes four paralogous proteins, two of which, DEMETER (DME) and REPRESSOR OF SILENCING 1 (ROS1), possess 5-methylcytosine-DNA glycosylase activity and are necessary for the regulation of development, response to infections and abiotic stress and silencing of transgenes and mobile elements. Homologues of DME and ROS1 are present in all plant groups; however, outside A. thaliana, they are poorly studied. Here we report the properties of a recombinant fragment of the ROS1 protein from Nicotiana tabacum (NtROS1), which contains all main structural domains required for catalytic activity. Using homologous modeling, we have constructed a structural model of NtROS1, which revealed folding characteristic of DNA glycosylases of the helix– hairpin–helix structural superfamily. The recombinant NtROS1 protein was able to remove mC bases from DNA, and the enzyme activity was barely affected by the methylation status of CG dinucleotides in the opposite strand. The enzyme removed 5-hydroxymethylcytosine (hmC) from DNA with a lower eff iciency, showing minimal activity in the presence of mC in the opposite strand. Expression of the NtROS1 gene in cultured human cells resulted in a global decrease in the level of genomic DNA methylation. In general, it can be said that the NtROS1 protein and other homologues of DME and ROS1 represent a promising scaffold for engineering enzymes to analyze the status of epigenetic methylation and to control gene activity.

The Physics of DNA Folding: Polymer Models and Phase-Separation.

Within cell nuclei, several biophysical processes occur in order to allow the correct activities of the genome such as transcription and gene regulation. To quantitatively investigate such processes, polymer physics models have been developed to unveil the molecular mechanisms underlying genome functions. Among these, phase-separation plays a key role since it controls gene activity and shapes chromatin spatial structure. In this paper, we review some recent experimental and theoretical progress in the field and show that polymer physics in synergy with numerical simulations can be helpful for several purposes, including the study of molecular condensates, gene-enhancer dynamics, and the three-dimensional reconstruction of real genomic regions.

RNA Design Principles for Riboswitches that Regulate RNase P-Mediated tRNA Processing.

Riboswitches are an attractive target for the directed design of RNA-based regulators by in silico prediction. These noncoding RNA elements consist of an aptamer platform for the highly selective ligand recognition and an expression platform which controls gene activity typically at the level of transcription or translation. In previous work, we could successfully apply RNA folding prediction to implement a new riboswitch mechanism regulating processing of a tRNA by RNase P. In this contribution, we present detailed information about our pipeline consisting of in silico design combined with the biochemical analysis for the verification of the implemented mechanism. Furthermore, we discuss the applicability of the presented biochemical in vivo and in vitro methods for the characterization of other artificial riboswitches.

Relationship between High Expression of Kaiso Protein and Poor Prognosis of Lung Cancer and the Regulation Mechanism of Malignant Phenotype of Lung Cancer Cells.

In this study, Kaiso was discovered to be a unique member of the POZ-zinc fingers family of transcription factors, which has been implicated in the genesis and progression of cancer. Although there is still some debate, Kaiso is believed to be implicated in the development of human cancer. It should be noted that there is minimal evidence available on the therapeutic relevance of nuclear Kaiso in lung cancer in humans. Histone or DNA modifications that control gene activity outside of the underlying sequence are examples of epigenetic alternations. Epigenetic alterations are heritable but reversible. Human illness, such as lung cancer, is often related to epigenetic dysregulation. In preclinical and clinical studies, epigenetic-targeted therapy has shown significant therapeutic promise for solid tumours and has been used in the treatment of haematological malignancies using different medicines targeting epigenetic regulators. It is important to note that the abnormal activities of Kaiso enzymes in tumour growth are summarised below and the development of inhibitors or medicines targeting epigenetic enzyme regulation is highlighted.

RH17 restricts reproductive fate and represses autonomous seed coat development in sexual Arabidopsis.

Plant sexual and asexual reproduction through seeds (apomixis) is tightly controlled by complex gene regulatory programs, which are not yet fully understood. Recent findings suggest that RNA helicases are required for plant germline development. This resembles their crucial roles in animals, where they are involved in controlling gene activity and the maintenance of genome integrity. Here, we identified previously unknown roles of Arabidopsis RH17 during reproductive development. Interestingly, RH17 is involved in repression of reproductive fate and of elements of seed development in the absence of fertilization. In lines carrying a mutant rh17 allele, development of supernumerary reproductive cell lineages in the female flower tissues (ovules) was observed, occasionally leading to formation of two embryos per seed. Furthermore, seed coat, and putatively also endosperm development, frequently initiated autonomously. Such induction of several features phenocopying distinct elements of apomixis by a single mutation is unusual and suggests that RH17 acts in regulatory control of plant reproductive development. Furthermore, an in-depth understanding of its action might be of use for agricultural applications.

Optical Control of Base Editing and Transcription through Light‐Activated Guide RNA

AbstractWe developed a method to control gene activation and base editing in mammalian cells and zebrafish embryos via photochemically activated, caged guide RNAs. Four evenly distributed, caged nucleobases are installed in the 5’‐protospacer region of the gRNA to block interaction between the dCas9:gRNA complex and the dsDNA target. Upon light activation, gRNA:dsDNA hybridization is restored and catalytically dead Cas9‐based CRISPR activation or base editing is achieved. Caged gRNAs are customizable in design, offer spatiotemporal control of transcription activation and base editing in both mammalian cells and zebrafish embryos, and preserve the ability to form dCas9:gRNA complexes for enhanced gRNA stability. We hereby offer a versatile tool for conditional control of interactions between dCas9‐based regulators with their programmed genomic loci, thereby facilitating parsing of spatiotemporally complex genetic events, such as transcription regulation and single‐base mutagenesis.

Circular L-RNA aptamer promotes target recognition and controls gene activity.

Rational design of aptamers to incorporate unnatural nucleotides and special chemical moieties can expand their functional complexity and diversity. Spiegelmer (L-RNA aptamer) is a unique class of aptamer that is composed of unnatural L-RNA nucleotides, and so far there are limited L-RNA aptamer candidates and applications being reported. Moreover, the target binding properties of current L-RNA aptamers require significant improvement. Here, using L-Apt.4-1c as an example, we develop a simple and robust strategy to generate the first circular L-RNA aptamer, cycL-Apt.4-1c, quantitatively, demonstrate substantial enhancement in binding affinity and selectivity toward its target, and notably report novel applications of circular L-RNA aptamer in controlling RNA–protein interaction, and gene activity including telomerase activity and gene expression. Our approach and findings will be applicable to any L-RNA aptamers and open up a new avenue for diverse applications.

Scalable control of developmental timetables by epigenetic switching networks.

During development, progenitor cells follow timetables for differentiation that span many cell generations. These developmental timetables are robustly encoded by the embryo, yet scalably adjustable by evolution, facilitating variation in organism size and form. Epigenetic switches, involving rate-limiting activation steps at regulatory gene loci, control gene activation timing in diverse contexts, and could profoundly impact the dynamics of gene regulatory networks controlling developmental lineage specification. Here, we develop a mathematical framework to model regulatory networks with genes controlled by epigenetic switches. Using this framework, we show that such epigenetic switching networks uphold developmental timetables that robustly span many cell generations, and enable the generation of differentiated cells in precisely defined numbers and fractions. Changes to epigenetic switching networks can readily alter the timing of developmental events within a timetable, or alter the overall speed at which timetables unfold, enabling scalable control over differentiated population sizes. With their robust, yet flexibly adjustable nature, epigenetic switching networks could represent central targets on which evolution acts to manufacture diversity in organism size and form.

DAF-16/FoxO and DAF-12/VDR control cellular plasticity both cell-autonomously and via interorgan signaling.

Many cell types display the remarkable ability to alter their cellular phenotype in response to specific external or internal signals. Such phenotypic plasticity is apparent in the nematode Caenorhabditis elegans when adverse environmental conditions trigger entry into the dauer diapause stage. This entry is accompanied by structural, molecular, and functional remodeling of a number of distinct tissue types of the animal, including its nervous system. The transcription factor (TF) effectors of 3 different hormonal signaling systems, the insulin-responsive DAF-16/FoxO TF, the TGFβ-responsive DAF-3/SMAD TF, and the steroid nuclear hormone receptor, DAF-12/VDR, a homolog of the vitamin D receptor (VDR), were previously shown to be required for entering the dauer arrest stage, but their cellular and temporal focus of action for the underlying cellular remodeling processes remained incompletely understood. Through the generation of conditional alleles that allowed us to spatially and temporally control gene activity, we show here that all 3 TFs are not only required to initiate tissue remodeling upon entry into the dauer stage, as shown before, but are also continuously required to maintain the remodeled state. We show that DAF-3/SMAD is required in sensory neurons to promote and then maintain animal-wide tissue remodeling events. In contrast, DAF-16/FoxO or DAF-12/VDR act cell-autonomously to control anatomical, molecular, and behavioral remodeling events in specific cell types. Intriguingly, we also uncover non-cell autonomous function of DAF-16/FoxO and DAF-12/VDR in nervous system remodeling, indicating the presence of several insulin-dependent interorgan signaling axes. Our findings provide novel perspectives into how hormonal systems control tissue remodeling.