rna-associated-chromatin-dna-dna-interaction-method

RNA-Associated Chromatin DNA-DNA Interaction Method

RNA-Associated Chromatin DNA-DNA Interaction Method (RDD) is a targeted strategy for mapping chromatin interactions anchored by a specific RNA. It ...

Southern University of Science and Technology

The 2020 China report of the Lancet Countdown on health and climate change.

The 2021 China report of the Lancet Countdown on health and climate change: seizing the window of opportunity.

The 2022 China report of the Lancet Countdown on health and climate change: leveraging climate actions for healthy ageing.

We investigated the residual rate and mass loss rate of litter, as well as the carbon release dynamics of litter and soil across seasons, to better understand the effects of seasonal fluctuations on carbon dynamics in mixed coniferous forests. The study was carried out in natural mixed coniferous forests in the Xiaoxinganling region of Heilongjiang Province, China, and the number of temperature cycles in the unfrozen season, freeze-thaw season, frozen season, and thaw season was controlled. The goal of the study was to examine how the carbon release dynamics of litter and soil respond to the freeze-thaw process and whether there are differences in carbon release dynamics under different seasons. Repeated-measures analysis of variance was used to analyze the residual mass rate and mass loss rate of litter, litter organic carbon and soil organic carbon during the unfrozen season, freeze-thaw season, frozen season, and thaw season. Litter decomposition was highest in the unfrozen season (15.9%~20.3%), and litter and soil carbon were sequestered throughout this process. Temperature swings above and below 0°C during the freeze-thaw season cause the litter to physically fragment and hasten its decomposition. Decomposition of litter was still feasible during the frozen season, and it was at its lowest during the thaw season (7.2%~7.8%), when its organic carbon was transported to the soil. Carbon migrates from undecomposed litter to semi-decomposed litter and then to soil. The carbon in the environment is fixed in the litter (11.3%~18.2%) and soil (34.4%~36.7%) in the unfrozen season, the carbon-fixing ability of the undecomposed litter in the freeze-thaw season is better, and the carbon in the semi-decomposed litter is mostly transferred to the soil; the carbon-fixing ability of the litter in the frozen season is worse (-3.9%~ -4.3%), and the organic carbon in the litter is gradually transferred to the soil. The carbon-fixing ability of the undecomposed litter in the thaw season is stronger, and the organic carbon in the semi-decomposed litter is mostly transferred to the soil. Both litter and soil can store carbon; however, from the unfrozen season until the thaw season, carbon is transported from undecomposed litter to semi-decomposed litter and to the soil over time.

Effects of seasonal changes on the carbon dynamics in mixed coniferous forests.

The three-dimensional (3D) organization of chromatin within the nucleus is crucial for gene regulation. However, the 3D architectural features that coordinate the activation of an entire chromosome remain largely unknown. We introduce an omics method, RNA-associated chromatin DNA-DNA interactions, that integrates RNA polymerase II (RNAPII)-mediated regulome with stochastic optical reconstruction microscopy to investigate the landscape of noncoding RNA -associated chromatin topology for gene equalization to achieve dosage compensation. Our findings reveal that  anchors to the target gene transcription end sites (TESs) and spreads in a distinctive boot-shaped configuration, promoting a more open chromatin state for hyperactivation. Furthermore,  arches TES to transcription start sites to enhance transcriptional loops, potentially facilitating RNAPII convoying and connecting proximal promoter-promoter transcriptional hubs for synergistic gene regulation. These TESs cluster as  compartments, surrounded by inactive domains for coactivation of multiple genes within the  territory. In addition,  structures gradually form and scaffold for stepwise coactivation in dosage compensation.

3D chromatin structures associated with ncRNA  for hyperactivation and coactivation across the entire X chromosome.

CRISPR/Cas-based transcriptional activators can be enhanced by intrinsically disordered regions (IDRs). However, the underlying mechanisms are still debatable. Here, we examine 12 well-known IDRs by fusing them to the dCas9-VP64 activator, of which only seven can augment activation, albeit independently of their phase separation capabilities. Moreover, modular domains (MDs), another class of multivalent molecules, though ineffective in enhancing dCas9-VP64 activity on their own, show substantial enhancement in transcriptional activation when combined with dCas9-VP64-IDR. By varying the number of gRNA binding sites and fusing dCas9-VP64 with different IDRs/MDs, we uncover that optimal, rather than maximal, cis-trans cooperativity enables the most robust activation. Finally, targeting promoter-enhancer pairs yields synergistic effects, which can be further amplified via enhancing chromatin interactions. Overall, our study develops a versatile platform for efficient gene activation and sheds important insights into CRIPSR-based transcriptional activators enhanced with multivalent molecules.

Specific multivalent molecules boost CRISPR-mediated transcriptional activation.

Three-dimensional genome organization reveals that gene regulatory elements, which are linearly distant on the genome, can spatially interact with target genes to regulate their expression. DNA fluorescence in situ hybridization (DNA-FISH) is an efficient method for studying the spatial proximity of genomic loci. In this study, we developed an optimized Tn5 transposome-based DNA-FISH method, termed Tn5-labeled DNA-FISH. This approach amplifies the target region and uses a self-assembled Tn5 transposome to simultaneously fragment the DNA into ~100 bp segments and label it with fluorescent oligonucleotides in a single step. This method enables the preparation of probes for regions as small as 4 kb and visualizes both endogenous and exogenous genomic loci at kb resolution. Tn5-labeled DNA-FISH provides a streamlined and cost-effective tool for probe generation, facilitating the investigation of chromatin spatial conformations, gene interactions, and genome architecture.

Tn5-Labeled DNA-FISH: An Optimized Probe Preparation Method for Probing Genome Architecture.

Transcription factors (TFs) play a crucial role in the regulation of gene expression and the structural organization of chromatin. They interact with proteins, RNA, and chromatin DNA to exert their functions. Therefore, an efficient and straightforward experimental approach that simultaneously captures the interactions of transcription factors with DNA, RNA, and proteins is essential for studying these regulatory proteins. In this study, we developed a novel method, TF-chRDP (Transcription Factor binding Chromatin-associated RNA, DNA, and Protein), which allows for the concurrent capture of these biomolecules in a single experiment. We enriched chromatin complexes using specific antibodies and divided the chromatin into three fractions: one for DNA library preparation to analyze the genomic binding sites of transcription factors, another for RNA library preparation to investigate the RNA associated with transcription factor binding, and the third for proteomic analysis to identify protein cofactors interacting with transcription factors. We applied this method to study the transcription factor p53 and its associated chromatin complexes. The results demonstrated high specificity in the enrichment of DNA, RNA and proteins. This method provides an efficient tool for simultaneously capturing chromatin-associated RNA, DNA and protein bound to specific TF, making it particularly useful for analyzing the role of protein-DNA-RNA complexes in transcriptional regulation.

TF-chRDP: a method for simultaneously capturing transcription factor binding chromatin-associated RNA, DNA and protein.

The three-dimensional (3D) chromatin structure of Epstein-Barr virus (EBV) within host cells and the underlying mechanisms of chromatin interaction and gene regulation, particularly those involving EBV's noncoding RNAs (ncRNAs), have remained incompletely characterized. In this study, we employed state-of-the-art techniques of 3D genome mapping, including protein-associated chromatin interaction analysis with paired-end tag sequencing (ChIA-PET), RNA-associated chromatin interaction technique (RDD), and super-resolution microscopy, to delineate the spatial architecture of EBV in human lymphoblastoid cells. We systematically analyzed EBV-to-EBV (E-E), EBV-to-host (E-H), and host-to-host (H-H) interactions linked to host proteins and EBV RNAs. Our findings reveal that EBV utilizes host CCCTC-binding factor (CTCF) and RNA polymerase II (RNAPII) to form distinct chromatin contact domains (CCDs) and RNAPII-associated interaction domains (RAIDs). The anchors of these chromatin domains serve as platforms for extensive interactions with host chromatin, thus modulating host gene expression. Notably, EBV ncRNAs, especially Epstein-Barr-encoded RNAs (EBERs), target and interact with less accessible regions of host chromatin to repress a subset of genes via the inhibition of RNAPII-associated chromatin loops. This process involves the cofactor nucleolin (NCL) and its RNA recognition motifs, and depletion of either NCL or EBERs alters expression of genes crucial for host infection control, immune response, and cell cycle regulation. These findings unveil a sophisticated interplay between EBV and host chromatin.

Article	Year
The 2020 China report of the Lancet Countdown on health and climate change. The Lancet. Public health\| PubMed ID: 33278345	2020
The 2021 China report of the Lancet Countdown on health and climate change: seizing the window of opportunity. The Lancet. Public health\| PubMed ID: 34758286	2021
The 2022 China report of the Lancet Countdown on health and climate change: leveraging climate actions for healthy ageing. The Lancet. Public health\| PubMed ID: 36354045	2022
Effects of seasonal changes on the carbon dynamics in mixed coniferous forests. PloS one\| PubMed ID: 37079645	2023
3D chromatin structures associated with ncRNA for hyperactivation and coactivation across the entire X chromosome. Science advances\| PubMed ID: 39058769	2024
Specific multivalent molecules boost CRISPR-mediated transcriptional activation. Nature communications\| PubMed ID: 39174527	2024
Tn5-Labeled DNA-FISH: An Optimized Probe Preparation Method for Probing Genome Architecture. International journal of molecular sciences\| PubMed ID: 40076846	2025
TF-chRDP: a method for simultaneously capturing transcription factor binding chromatin-associated RNA, DNA and protein. Frontiers in cell and developmental biology\| PubMed ID: 40123855	2025
Landscape of the Epstein-Barr virus-host chromatin interactome and gene regulation. The EMBO journal\| PubMed ID: 40425856	2025

Tong Gao

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