Direct visualization of replication and R-loop collision using single-molecule imaging.

R-loops are three-stranded nucleic acid structures that can cause replication stress by blocking replication fork progression. However, the detailed mechanism underlying the collision of DNA replication forks and R-loops remains elusive. To investigate how R-loops induce replication stress, we use single-molecule fluorescence imaging to directly visualize the collision of replicating Phi29 DNA polymerase (Phi29 DNAp), the simplest replication system, and R-loops. We demonstrate that a single R-loop can block replication, and the blockage is more pronounced when an RNA-DNA hybrid is on the non-template strand. We show that this asymmetry results from secondary structure formation on the non-template strand, which impedes the progression of Phi29 DNAp. We also show that G-quadruplex formation on the displaced single-stranded DNA in an R-loop enhances the replication stalling. Moreover, we observe the collision between Phi29 DNAp and RNA transcripts synthesized by T7 RNA polymerase (T7 RNAp). RNA transcripts cause more stalling because of the presence of T7 RNAp. Our work provides insights into how R-loops impede DNA replication at single-molecule resolution.

Alteration of replication protein A binding mode on single-stranded DNA by NSMF potentiates RPA phosphorylation by ATR kinase.

Replication protein A (RPA), a eukaryotic single-stranded DNA (ssDNA) binding protein, dynamically interacts with ssDNA in different binding modes and plays essential roles in DNA metabolism such as replication, repair, and recombination. RPA accumulation on ssDNA due to replication stress triggers the DNA damage response (DDR) by activating the ataxia telangiectasia and RAD3-related (ATR) kinase, which phosphorylates itself and downstream DDR factors, including RPA. We recently reported that the N-methyl-D-aspartate receptor synaptonuclear signaling and neuronal migration factor (NSMF), a neuronal protein associated with Kallmann syndrome, promotes RPA32 phosphorylation via ATR upon replication stress. However, how NSMF enhances ATR-mediated RPA32 phosphorylation remains elusive. Here, we demonstrate that NSMF colocalizes and physically interacts with RPA at DNA damage sites in vivo and in vitro. Using purified RPA and NSMF in biochemical and single-molecule assays, we find that NSMF selectively displaces RPA in the more weakly bound 8- and 20-nucleotide binding modes from ssDNA, allowing the retention of more stable RPA molecules in the 30-nt binding mode. The 30-nt binding mode of RPA enhances RPA32 phosphorylation by ATR, and phosphorylated RPA becomes stabilized on ssDNA. Our findings provide new mechanistic insight into how NSMF facilitates the role of RPA in the ATR pathway.

MSH2-MSH3 promotes DNA end resection during homologous recombination and blocks polymerase theta-mediated end-joining through interaction with SMARCAD1 and EXO1.

DNA double-strand break (DSB) repair via homologous recombination is initiated by end resection. The extent of DNA end resection determines the choice of the DSB repair pathway. Nucleases for end resection have been extensively studied. However, it is still unclear how the potential DNA structures generated by the initial short resection by MRE11-RAD50-NBS1 are recognized and recruit proteins, such as EXO1, to DSB sites to facilitate long-range resection. We found that the MSH2-MSH3 mismatch repair complex is recruited to DSB sites through interaction with the chromatin remodeling protein SMARCAD1. MSH2-MSH3 facilitates the recruitment of EXO1 for long-range resection and enhances its enzymatic activity. MSH2-MSH3 also inhibits access of POLθ, which promotes polymerase theta-mediated end-joining (TMEJ). Collectively, we present a direct role of MSH2-MSH3 in the initial stages of DSB repair by promoting end resection and influencing the DSB repair pathway by favoring homologous recombination over TMEJ.

Advances in Biocarbon and Soft Material Assembly for Enthalpy Storage: Fundamentals, Mechanisms, and Multimodal Applications.

High-value-added biomass materials like biocarbon are being actively pursued integrating them with soft materials in a broad range of advanced renewable energy technologies owing to their advantages, such as lightweight, relatively low-cost, diverse structural engineering applications, and high energy storage potential. Consequently, the hybrid integration of soft and biomass-derived materials shall store energy to mitigate intermittency issues, primarily through enthalpy storage during phase change. This paper introduces the recent advances in the development of natural biomaterial-derived carbon materials in soft material assembly and its applications in multidirectional renewable energy storage. Various emerging biocarbon materials (biochar, carbon fiber, graphene, nanoporous carbon nanosheets (2D), and carbon aerogel) with intrinsic structures and engineered designs for enhanced enthalpy storage and multimodal applications are discussed. The fundamental design approaches, working mechanisms, and feature applications, such as including thermal management and electromagnetic interference shielding, sensors, flexible electronics and transparent nanopaper, and environmental applications of biocarbon-based soft material composites are highlighted. Furthermore, the challenges and potential opportunities of biocarbon-based composites are identified, and prospects in biomaterial-based soft materials composites are presented.

Factors determining contamination and time trends in cyclic and linear siloxanes in sediments from an industrialized lake in Korea.

Siloxanes, widely used in various consumer and industrial products, are emerging concerns of contaminants. Despite this, limited studies have been conducted on contamination and time trends on siloxanes in coastal environments. In the present study, four cyclic and 15 linear siloxanes were measured in sediments collected from an artificial saltwater lake in Korea during 2001-2016 to investigate contamination, time trends, and ecotoxicological concerns. Cyclic siloxanes were detected in all sediment samples, whereas linear siloxanes were not frequently detected. The highest siloxane concentrations were observed in creeks passing through various industrial complexes, indicating that industrial activities predominantly contributed to siloxane contamination in coastal environments. Decamethylcyclopentasiloxane (D5) and dodecylcyclohexasiloxane (D6) were predominant siloxanes in sediments over the last two decades. Siloxane concentrations significantly increased in creek sediments from 2008 to 2016, whereas those in inshore and offshore regions significantly decreased due to a strong dilution effect by the operation of tidal power plant. This suggests that consumption patterns and coastal development activities are crucial factors determining the contamination and time trends in the sedimentary siloxanes. The sedimentary concentrations of octamethylcyclotetrasiloxane (D4) and D5 exceeded several thresholds, raising the potentials for ecological risks to aquatic organisms.

Lactate oxidase/catalase-displaying nanoparticles efficiently consume lactate in the tumor microenvironment to effectively suppress tumor growth.

The aggressive proliferation of tumor cells often requires increased glucose uptake and excessive anaerobic glycolysis, leading to the massive production and secretion of lactate to form a unique tumor microenvironment (TME). Therefore, regulating appropriate lactate levels in the TME would be a promising approach to control tumor cell proliferation and immune suppression. To effectively consume lactate in the TME, lactate oxidase (LOX) and catalase (CAT) were displayed onto Aquifex aeolicus lumazine synthase protein nanoparticles (AaLS) to form either AaLS/LOX or AaLS/LOX/CAT. These complexes successfully consumed lactate produced by CT26 murine colon carcinoma cells under both normoxic and hypoxic conditions. Specifically, AaLS/LOX generated a large amount of H2O2 with complete lactate consumption to induce drastic necrotic cell death regardless of culture condition. However, AaLS/LOX/CAT generated residual H2O2, leading to necrotic cell death only under hypoxic condition similar to the TME. While the local administration of AaLS/LOX to the tumor site resulted in mice death, that of AaLS/LOX/CAT significantly suppressed tumor growth without any severe side effects. AaLS/LOX/CAT effectively consumed lactate to produce adequate amounts of H2O2 which sufficiently suppress tumor growth and adequately modulate the TME, transforming environments that are favorable to tumor suppressive neutrophils but adverse to tumor-supportive tumor-associated macrophages. Collectively, these findings showed that the modular functionalization of protein nanoparticles with multiple metabolic enzymes may offer the opportunity to develop new enzyme complex-based therapeutic tools that can modulate the TME by controlling cancer metabolism.

Multiple CTCF sites cooperate with each other to maintain a TAD for enhancer-promoter interaction in the ß-globin locus.

Insulators are cis-regulatory elements that block enhancer activity and prevent heterochromatin spreading. The binding of CCCTC-binding factor (CTCF) protein is essential for insulators to play the roles in a chromatin context. The ß-globin locus, consisting of multiple genes and enhancers, is flanked by two insulators 3'HS1 and HS5. However, it has been reported that the absence of these insulators did not affect the ß-globin transcription. To explain the unexpected finding, we have deleted a CTCF motif at 3'HS1 or HS5 in the human ß-globin locus and analyzed chromatin interactions around the locus. It was found that a topologically associating domain (TAD) containing the ß-globin locus is maintained by neighboring CTCF sites in the CTCF motif-deleted loci. The additional deletions of neighboring CTCF motifs disrupted the ß-globin TAD, resulting in decrease of the ß-globin transcription. Chromatin interactions of the ß-globin enhancers with gene promoter were weakened in the multiple CTCF motifs-deleted loci, even though the enhancers have still active chromatin features such as histone H3K27ac and histone H3 depletion. Genome-wide analysis using public CTCF ChIA-PET and ChIP-seq data showed that chromatin domains possessing multiple CTCF binding sites tend to contain super-enhancers like the ß-globin enhancers. Taken together, our results show that multiple CTCF sites surrounding the ß-globin locus cooperate with each other to maintain a TAD. The ß-globin TAD appears to provide a compact spatial environment that enables enhancers to interact with promoter.

Histone H3K4me1 and H3K27ac play roles in nucleosome eviction and eRNA transcription, respectively, at enhancers.

Histone H3K4me1 and H3K27ac are enhancer-specific modifications and are required for enhancers to activate transcription of target genes. However, the reciprocal effects of these histone modifications on each other and their roles in enhancers are not clear. Here to comparatively analyze the role of these modifications, we inhibited H3K4me1 and H3K27ac by deleting the SET domains of histone methyltransferases MLL3 and MLL4 and the HAT domain of histone acetyltransferase p300, respectively, in erythroid K562 cells. The loss of H3K4me1 reduced H3K27ac at the ß-globin enhancer LCR HSs, but H3K27ac reduction did not affect H3K4me1. This unequal relationship between two modifications was revealed in putative enhancers by genome-wide analysis using ChIP-seq. Histone H3 eviction at putative enhancers was weakened by the loss of H3K4me1 but not by the loss of H3K27ac. Chromatin remodeling complexes were recruited into the ß-globin LCR HSs in a H3K4me1-dependent manner. In contrast, H3K27ac was required for enhancer RNA (eRNA) transcription, and H3K4me1 was not enough for it. Forced H3K27ac-induced eRNA transcription without affecting H3K4me1 at the ß-globin LCR HSs. These results indicate that H3K4me1 and H3K27ac affect each other in different ways and play more direct roles in nucleosome eviction and eRNA transcription, respectively, at enhancers.

Use of biochar co-mediated chitosan mesopores to encapsulate alkane and improve thermal properties.

Phase-change materials (PCMs) plays a significant role in energy conservation and thermal management systems. However, excessive seepage and insufficient thermal conductivity of pristine PCMs are restricting its real-world applications. Herein, "anisotropic-like" biochar with favorable pore characteristics is designed by combining it with chitosan for dodecane encapsulation. The use of biochar could overcome high manufacturing costs and associated environmental issues of PCM supporting materials. Biochar co-mediated chitosan enrich the mesopore proportion (96.5%) and provide interactive synergistic architecture. The prepared composite PCM exhibited outstanding latent heat retention of 95.9% after repeated cycling, high loading ratio, enhanced thermal conductivity (0.373 W/(m·K)), leakage-free, and repeatable utilization properties above the melting point of pristine dodecane. A figure of merit of 33.94 × 106 W2 S/(m4oC) was achieved, far surpassing that measure among reported biochar-based composite PCMs. This study provides insights into next-generation sustainable energy storage development for a key global sustainability goal.

The miR-182-5p/NDRG1 Axis Controls Endometrial Receptivity through the NF-κB/ZEB1/E-Cadherin Pathway.

Endometrial receptivity is essential for successful pregnancy, and its impairment is a major cause of embryo-implantation failure. MicroRNAs (miRNAs) that regulate epigenetic modifications have been associated with endometrial receptivity. However, the molecular mechanisms whereby miRNAs regulate endometrial receptivity remain unclear. Therefore, we investigated whether miR-182 and its potential targets influence trophoblast cell attachment. miR-182 was expressed at lower levels in the secretory phase than in the proliferative phase of endometrium tissues from fertile donors. However, miR-182 expression was upregulated during the secretory phase in infertile women. Transfecting a synthetic miR-182-5p mimic decreased spheroid attachment of human JAr choriocarcinoma cells and E-cadherin expression (which is important for endometrial receptivity). miR-182-5p also downregulated N-Myc downstream regulated 1 (NDRG1), which was studied further. NDRG1 was upregulated in the secretory phase of the endometrium tissues and induced E-cadherin expression through the nuclear factor-κΒ (NF-κΒ)/zinc finger E-box binding homeobox 1 (ZEB1) signaling pathway. NDRG1-overexpressing or -depleted cells showed altered attachment rates of JAr spheroids. Collectively, our findings indicate that miR-182-5p-mediated NDRG1 downregulation impaired embryo implantation by upregulating the NF-κΒ/ZEB1/E-cadherin pathway. Hence, miR-182-5p is a potential biomarker for negative selection in endometrial receptivity and a therapeutic target for successful embryo implantation.

Practical solutions with PCM for providing thermal stability of temporary house, school and hospital in disaster situations.

Globally, humanity is at risk from the coronavirus disease (COVID-19). To address the shortage of beds in quarantining those infected with COVID-19, hospitals have prepared temporary beds. However, for temporary hospital beds, it is difficult to maintain a comfortable temperature due to lack of insulation and heat storage. Phase change materials (PCMs) are used to provide temperature stability and control for temporary structure. Therefore, this study aimed to conduct experiments that analyze the effect of room temperature stabilization using a PCM. The method of macro packed PCM (MPPCM) was used to apply the PCM to buildings. The MPPCM installation location was selected and the effect of reducing the box temperature was analyzed, according to the strength of the heat source. As a result, a maximum reduction of 4.9 °C in the box temperature was achieved. Therefore, the application of MPPCM to buildings give to stabilize the box temperature. And the result showed the possibility of providing a comfortable indoor space for temporary hospital beds.

LncRNA LINC00240 suppresses invasion and migration in non-small cell lung cancer by sponging miR-7-5p.

BACKGROUND: lncRNAs have important roles in regulating cancer biology. Accumulating evidence has established a link between the dysregulation of lncRNAs and microRNA in cancer progression. In previous studies, miR-7-5p has been found to be significantly down-regulated in mesenchymal-like lung cancer cell lines and directly regulated EGFR. In this work, we investigated the lncRNA partner of miR-7-5p in the progression of lung cancer. METHODS: We investigated the expression of miR-7-5p and the lncRNA after transfection with an miR-7-5p mimics using a microarray. The microarray results were validated using quantitative real time-polymerase Chain Reaction (qRT-PCR). The regulatory effects of lncRNA on miR-7-5p and its target were evaluated by changes in the expression of miR-7-5p after transfection with siRNAs for lncRNA and the synthesis of full-length lncRNA. The effect of miR-7-5p on lncRNA and the miRNA target was evaluated after transfection with miRNA mimic and inhibitor. The role of lncRNA in cancer progression was determined using invasion and migration assays. The level of lncRNA and EGFR in lung cancer and normal lung tissue was analyzed using TCGA data. RESULTS: We found that LINC00240 was downregulated in lung cancer cell line after miR-7-5p transfection with an miR-7-5p mimic. Further investigations revealed that the knockdown of LINC00240 induced the overexpression of miR-7-5p. The overexpression of miR-7-5p diminished cancer invasion and migration. The EGFR expression was down regulated after siRNA treatment for LINC00240. Silencing LINC00240 suppressed the invasion and migration of lung cancer cells, whereas LINC00240 overexpression exerted the opposite effect. The lower expression of LINC00240 in squamous lung cancer was analyzed using TCGA data. CONCLUSIONS: Taken together, LINC00240 acted as a sponge for miR-7-5p and induced the overexpression of EGFR. LINC00240 may represent a potential target for the treatment of lung cancer.

GATA-1-dependent histone H3K27 acetylation mediates erythroid cell-specific chromatin interaction between CTCF sites.

CCCTC-binding factor (CTCF) sites interact with each other in the chromatin environment, establishing chromatin domains. Our previous study showed that interaction between CTCF sites is cell type-specific around the ß-globin locus and is dependent on erythroid-specific activator GATA-1. To find out molecular mechanisms of the cell type-specific interaction, we directly inhibited GATA-1 binding to the ß-globin enhancers by deleting its binding motifs and found that histone H3K27 acetylation (H3K27ac) was decreased at CTCF sites surrounding the ß-globin locus, even though CTCF binding itself was maintained at the sites. Forced H3K27ac by Trichostatin A treatment or CBP/p300 KD affected the interactions between CTCF sites around the ß-globin locus without changes in CTCF binding. Analysis of public ChIA-PET data revealed that H3K27ac is higher at CTCF sites forming short interactions than long interactions. GATA-1 was identified as a representative transcription factor that relates with genes present inside the short interactions in erythroid K562 cells. Depletion of GATA-1-reduced H3K27ac at CTCF sites near erythroid-specific enhancers. These results indicate that H3K27ac at CTCF sites is required for cell type-specific chromatin interactions between them. Tissue-specific activator GATA-1 appears to play a role in H3K27ac at CTCF sites in erythroid cells.

A comparative analysis of biochar, activated carbon, expanded graphite, and multi-walled carbon nanotubes with respect to PCM loading and energy-storage capacities.

To obtain high thermal performance composite phase change materials (PCMs), various other supporting materials have been utilized to encapsulate organic PCMs. In this study, four carbon materials (biochar, activated carbon, carbon nanotubes, and expanded graphite) were introduced to support heptadecane. The composite PCMs were designed using vacuum impregnation techniques. The structural stability, chemical compatibility, thermal stability, and thermal energy storage capacity of the as-prepared materials were systematically characterized using differential scanning calorimetry, Fourier-transform infrared spectroscopy, etc. Among the supporting materials, expanded graphite had a high PCM content of 94.5%, whereas it was low for biochar-supported PCM (25.7%). Meanwhile, the latent heat storage capacity ranged from 53.3 J/g to 195.9 J/g. It was observed that the intermolecular interactions between the PCM and supporting materials and the surface functionality of the encapsulating agents play a leading role in the thermal performance of the composite PCMs. Furthermore, pore structures such as specific surface area, total pore volume, and pore size distribution have a combined effect on the crystallinity of heptadecane in the composite PCMs. The study will provide insight into developing and designing carbon-based composite PCMs for heat-storage purposes.

High-throughput single-molecule imaging system using nanofabricated trenches and fluorescent DNA-binding proteins.

DNA curtain is a high-throughput system, integrating a lipid bilayer, fluorescence imaging, and microfluidics to probe protein-DNA interactions in real-time and has provided in-depth understanding of DNA metabolism. Especially, the microfluidic platform of a DNA curtain is highly suitable for a biochip. In the DNA curtain, DNA molecules are aligned along chromium nanobarriers, which are fabricated on a slide surface, and visualized using an intercalating dye, YOYO-1. Although the chromium barriers confer precise geometric alignment of DNA, reuse of the slides is limited by wear of the barriers during cleaning. YOYO-1 is rapidly photobleached and causes photocleavage of DNA under continuous laser illumination, restricting DNA observation to a brief time window. To address these challenges, we developed a new nanopatterned slide, upon which carved nanotrenches serve as diffusion barriers. The nanotrenches were robust under harsh cleaning conditions, facilitating the maintenance of surface cleanliness that is essential to slide reuse. We also stained DNA with a fluorescent protein with a DNA-binding motif, fluorescent protein-DNA binding peptide (FP-DBP). FP-DBP was slowly photobleached and did not cause DNA photocleavage. This new DNA curtain system enables a more stable and repeatable investigation of real-time protein-DNA interactions and will serve as a good platform for lab-on-a-chip.

LRF acts as an activator and repressor of the human ß-like globin gene transcription in a developmental stage dependent manner.

Leukemia/lymphoma-related factor (LRF; a hematopoietic transcription factor) has been suggested to repress fetal γ-globin genes in the human adult stage ß-globin locus. Here, to study the role of LRF in the fetal stage ß-globin locus, we knocked out its expression in erythroid K562 cells, in which the γ-globin genes are mainly transcribed. The γ-globin transcription was reduced in LRF knock-out cells, and transcription factor binding to the ß-globin locus control region hypersensitive sites (LCR HSs) and active histone organization in the LCR HSs were disrupted by the depletion of LRF. In contrast, LRF loss in the adult stage ß-globin locus did not affect active chromatin structure in the LCR HSs and induced the fetal γ-globin transcription. These results indicate that LRF may act as an activator and repressor of the human ß-like globin gene transcription in a manner dependent on developmental stage.

Increase of Hspa1a and Hspa1b genes in the resting B cells of Sirt1 knockout mice.

Sirt1, also known as the longevity gene, is an NAD+-dependent class III histone deacetylase that has been extensively studied in multiple areas of research including cellular metabolism, longevity, cancer, autoimmunity, and immunity. However, little is known about the function of Sirt1 in B cells. This study aimed to investigate the role of Sirt1 in the expression pattern of mRNAs in the resting B cells of mice. CD19+ B cell-specific inducible Sirt1 knockout (KO) mice were divided into tamoxifen-treated Sirt1 KO group (S19T) or control group (S19). mRNAs extracted from resting B cells of both groups were analyzed for differentially expressed genes (DEG) using microarray. DEG analysis showed significant differential expression of 20 genes, of which Hspa1a and Hspa1b showed the highest fold change (FC) in S19T compared with S19 (p value < 0.01 and FC > 3). Further, Kyoto Encyclopedia of Genes and Genomes analysis identified pathways associated with diseases, organismal systems, and antigen processing and presentation. Additionally, the pathways known to involve Hspa1a and Hspa1b were also activated in the S19T group. On the other hand, after in vitro stimulation with lipopolysaccharide, cell viability and IgM production were significantly decreased in Sirt1 KO B cells, while expressions of TNF-α, IL-6, and IL-10 were increased. In summary, our study reveals that Sirt1 may maintain the quiescent state in resting B cells by suppressing the increase of Hspa1a and Hspa1b. This work provides a foundation for further studies on the functional roles of Sirt1 in B cells.

KLF1 stabilizes GATA-1 and TAL1 occupancy in the human ß-globin locus.

KLF1 is an erythroid specific transcription factor that binds to regulatory regions of erythroid genes. Binding sites of KLF1 are often found near binding sites of GATA-1 and TAL1. In the ß-globin locus, KLF1 is required for forming active chromatin structure, although its role is unclear. To explore the role of KLF1 in transcribing the human γ-globin genes, we stably reduced the expression of KLF1 in erythroid K562 cells, compromising its association in the ß-globin locus. The γ-globin transcription was reduced with disappearance of active chromatin structure of the locus in the KLF1 knockdown cells. Interestingly, GATA-1 and TAL1 binding was reduced in the ß-globin locus, even though their expressions were not affected by KLF1 knockdown. The KLF1-dependent GATA-1 and TAL1 binding was observed in the adult locus transcribing the ß-globin gene and in several erythroid genes, where GATA-1 occupancy is independent from TAL1. These results indicate that KLF1 plays a role in facilitating and/or stabilizing GATA-1 and TAL1 occupancy in the erythroid genes, contributing to the generation of active chromatin structure such as histone acetylation and chromatin looping.

Analysis of alcohol-induced DNA damage in Escherichia coli by visualizing single genomic DNA molecules.

Consumption of alcohol injures DNA, and such damage is considered to be a primary cause for the development of cancer and many other diseases essentially due to reactive oxygen species generated from alcohol. To sensitively detect alcohol-induced DNA lesions in a biological system, we introduced a novel analytical platform for visualization of single genomic DNA molecules using E. coli. By fluorescently labelling the DNA lesions, our approach demonstrated, with the highest sensitivity, that we could count the number of DNA lesions induced by alcohol metabolism in a single bacterial cell. Moreover, our results showed a linear relationship between ethanol concentration and the number of DNA lesions: 0.88 lesions per 1% ethanol. Using this approach, we quantitatively analysed the DNA damage induced by exposure to alcoholic beverages such as beer (5% ethanol), rice wine (13%), soju (20%), and whisky (40%).

The hematopoietic regulator TAL1 is required for chromatin looping between the ß-globin LCR and human γ-globin genes to activate transcription.

TAL1 is a key hematopoietic transcription factor that binds to regulatory regions of a large cohort of erythroid genes as part of a complex with GATA-1, LMO2 and Ldb1. The complex mediates long-range interaction between the ß-globin locus control region (LCR) and active globin genes, and although TAL1 is one of the two DNA-binding complex members, its role is unclear. To explore the role of TAL1 in transcription activation of the human γ-globin genes, we reduced the expression of TAL1 in erythroid K562 cells using lentiviral short hairpin RNA, compromising its association in the ß-globin locus. In the TAL1 knockdown cells, the γ-globin transcription was reduced to 35% and chromatin looping of the (G)γ-globin gene with the LCR was disrupted with decreased occupancy of the complex member Ldb1 and LMO2 in the locus. However, GATA-1 binding, DNase I hypersensitive site formation and several histone modifications were largely maintained across the ß-globin locus. In addition, overexpression of TAL1 increased the γ-globin transcription and increased interaction frequency between the (G)γ-globin gene and LCR. These results indicate that TAL1 plays a critical role in chromatin loop formation between the γ-globin genes and LCR, which is a critical step for the transcription of the γ-globin genes.