Structural basis of DNA replication origin recognition by human Orc6 protein binding with DNA.

The six-subunit origin recognition complex (ORC), a DNA replication initiator, defines the localization of the origins of replication in eukaryotes. The Orc6 subunit is the smallest and the least conserved among ORC subunits. It is required for DNA replication and essential for viability in all species. Orc6 in metazoans carries a structural homology with transcription factor TFIIB and can bind DNA on its own. Here, we report a solution structure of the full-length human Orc6 (HsOrc6) alone and in a complex with DNA. We further showed that human Orc6 is composed of three independent domains: N-terminal, middle and C-terminal (HsOrc6-N, HsOrc6-M and HsOrc6-C). We also identified a distinct DNA-binding domain of human Orc6, named as HsOrc6-DBD. The detailed analysis of the structure revealed novel amino acid clusters important for the interaction with DNA. Alterations of these amino acids abolish DNA-binding ability of Orc6 and result in reduced levels of DNA replication. We propose that Orc6 is a DNA-binding subunit of human/metazoan ORC and may play roles in targeting, positioning and assembling the functional ORC at the origins.

The crystal structure of an antiparallel chair-type G-quadruplex formed by Bromo-substituted human telomeric DNA.

Human telomeric guanine-rich DNA, which could adopt different G-quadruplex structures, plays important roles in protecting the cell from recombination and degradation. Although many of these structures were determined, the chair-type G-quadruplex structure remains elusive. Here, we present a crystal structure of the G-quadruplex composed of the human telomeric sequence d[GGGTTAGG8GTTAGGGTTAGG20G] with two dG to 8Br-dG substitutions at positions 8 and 20 with syn conformation in the K+ solution. It forms a novel three-layer chair-type G-quadruplex with two linking trinucleotide loops. Particularly, T5 and T17 are coplanar with two water molecules stacking on the G-tetrad layer in a sandwich-like mode through a coordinating K+ ion and an A6•A18 base pair. While a twisted Hoogsteen A12•T10 base pair caps on the top of G-tetrad core. The three linking TTA loops are edgewise and each DNA strand has two antiparallel adjacent strands. Our findings contribute to a deeper understanding and highlight the unique roles of loop and water molecule in the folding of the G-quadruplex.

In situ preparation of visible-light-driven carbon quantum dots/NaBiO3 hybrid materials for the photoreduction of Cr(VI).

In this study, different carbon quantum dots (CQDs)/NaBiO3 hybrid materials were synthesized as photocatalysts to effectively utilize visible light for the photocatalytic degradation of contaminants effectively. These hybrid materials exhibit an enhanced photocatalytic reduction of hexavalent chromium (Cr(VI)) in the aqueous medium. Zero-dimensional nanoparticles of CQDs were embedded within the two-dimensional NaBiO3 nanosheets by the hydrothermal process. Compared with that of the pure NaBiO3 nanosheets, the photocatalytic performance of the hybrid catalysts was significantly high and 6 wt.% CQDs/NaBiO3 catalyst exhibited better photocatalytic performance. We performed the first-principles density functional theory calculations to study the interfacial properties of pure NaBiO3 nanosheets and hybrid photocatalysts, and confirmed the CQDs played an important role in the CQDs/NaBiO3 composites. The experimental results indicated that the enhanced reduction of Cr(VI) was probably due to the high loading of CQDs (electron acceptor) on NaBiO3, which made NaBiO3 nanomaterials to respond in visible light and significantly improved their electron-hole separation efficiency.

Autophagy inhibits TLR4-mediated invasiveness of oral cancer cells via the NF-κB pathway.

OBJECTIVES: Toll-like receptor 4 (TLR4) is abnormally expressed in oral cancer tissues and promotes cancer cell invasion. The purpose of this study was to clarify the mechanism by which autophagy regulates oral cancer invasion through the TLR4-NF-κB pathway. SUBJECTS AND METHODS: We examined TLR4 expression in oral cancer tissues and analysed the relationship between its expression and clinicopathological features. The invasion and migration of LPS-stimulated oral cancer cells with up- or downregulation of TLR4 expression was detected in addition to NF-κB signalling and autophagy levels. Furthermore, the role of autophagy in regulating TLR4-mediated cell invasiveness was explored by silencing the expression of key autophagy genes ATG7 and p62. RESULTS: We found that TLR4 overexpression was closely related to cervical lymphatic metastasis and poor survival. TLR4 activated the NF-κB pathway to promote the invasiveness of OSCC cells, and autophagy partly inhibited invasiveness by suppressing the NF-κB pathway. We observed that p62 translocated from the cytoplasm to the nucleus when autophagy was activated by LPS. Finally, silencing p62 further promoted LPS-mediated cell invasiveness. CONCLUSION: Toll-like receptor 4 significantly enhanced the invasiveness of OSCC cells. Autophagy may regulate cell invasiveness through the NF-κB pathway by modulating both the cytoplasmic and nuclear levels of p62.

Role of Noxa in proliferation, apoptosis, and autophagy in human adenoid cystic carcinoma.

BACKGROUND: Noxa, which is subset of the Bcl-2 family of proteins, was previously reported to have considerable therapeutic potential in diverse cancers. However, its expression and role in salivary gland adenoid cystic carcinoma (ACC) have not been well studied. This study aimed to elucidate the expression and role of Noxa in salivary gland ACC. MATERIALS AND METHODS: The expression levels of NOXA and its association with overall survival in salivary gland ACC were analyzed by quantitative real-time PCR. We next examined the effects of Noxa overexpression or inhibition on colony formation, proliferation, apoptosis, and autophagy of salivary gland ACC cells. Furthermore, promoter analysis was performed to identify the potential transcriptional activator of NOXA. RESULTS: NOXA was markedly down-regulated and significantly correlated with a more aggressive phenotype and poor overall survival of salivary gland ACC. Ectopic expression of Noxa suppressed the viability and growth of ACC cells, which involved the induction of apoptosis and autophagy. Moreover, the transcriptional activity of NOXA gene could be enhanced by p53. CONCLUSION: The findings of this study indicate that Noxa, activated transcriptionally by p53, suppress the progression of ACC, whereby it regulates proliferation, apoptosis, and autophagy.

Hybrid microbial electrolytic/UV system for highly efficient organic pollutants removal.

This study for the first time proposed an efficient microbial electrolyte/UV system for Methyl Orange decomposition. With an external applied voltage of 0.2 V and cathode aeration of 20 mL/min, H2O2 could be in-situ generated from two-electron reduction of oxygen in cathode, reaching to 8.1 mg/L in 2 hr and continued to increase. The pollutant removal efficiency of approximate 94.7% was achieved at initial neutral pH, with the activation of •OH in the presence of UV illumination. Although the nature of its guiding principles remain on the vista of practical exploration, this proof-of-concept study provides an alternative operation pattern of solar-microbial hybrid technology for future wastewater treatment from a basic but multidisciplinary view.

Amplified Assembly of G-Quadruplex-Decorated DNA Network Nanostructure toward AIE Signaling-Based Sensitive Biosensing.

Aggregation-induced emission (AIE) has offered a promising approach for developing low-background fluorescent methods; however, its applications often suffer from complex probe synthesis and poor biocompatibility. Herein, a novel AIE biosensing method for kanamycin antibiotic assays was developed by utilizing a DNA network nanostructure assembled from an aptamer recognition reaction to capture a large number of tetraphenylethylene fluorogen-labeled signal DNA (DTPE) probes. Due to the excellent hydrophilicity of the oligonucleotides, DTPE exhibited excellent water solubility without obvious background signal emission. Based on an ingenious nucleotide design, an abundance of G-quadruplex blocks neighboring the captured DTPE were formed on the DNA nanostructure. Because of the greatly restricted free motion of DTPE by this unique nanostructure, a strong AIE fluorescence signal response was produced to construct the signal transduction strategy. Together with target recycling and rolling circle amplification-based cascade nucleic acid amplification, this method exhibited a wide linear range from 75 fg mL-1 to 1 ng mL-1 and a detection limit down to 24 fg mL-1. The excellent analytical performance and effective manipulation improvement of the method over previous approaches determine its promising potential for various applications.

Phenol degradation in iron-based advanced oxidation processes through ferric reduction assisted by molybdenum disulfide.

In the iron-based advanced oxidation processes (AOPs), direct use of FeIII can be more convenient than FeII but the reduction of FeIII to FeII is a rate-limiting step. Introducing co-catalysts with abundant reducing sites to Fe-based AOPs can be an efficient way to accelerate the Fe redox process. Herein, molybdenum disulfide (MoS2) was used to enhance the catalytic performance of Fe3+/persulfate (PS) for phenol removal. In the Fe3+/MoS2/PS system, 99.6 ± 0.1% of phenol was removed in 60 min, comparable to that of the Fe2+/PS/MoS2 system (99.1 ± 0.3%). With the help of MoS2, 99.3 ± 4.2% of Fe3+ was transformed to Fe2+ in 10 min, and the Fe2+/Fe ratio was able to be maintained at 70.0 ± 1.4% after 60 min. The rapid and complete reduction of Fe3+ with MoS2 made it possible to replace Fe2+ by Fe3+, which is easier to store, transport, and use. The decrease in XPS peak area percentage of Mo(IV) and the lower valent S after reaction revealed that MoS2 acted as an electron provider in the Fe redox cycle. Quenching experiment results indicated that the phenol removal was highly depended on the surface-bound radicals, including both SO4•- and •OH. Those results have demonstrated that ferric salts can be directly used in the Fe-based AOPs and the redox cycle could be sustained with the assistance of MoS2.

Activation of persulfate for degradation of sodium dodecyl sulfate by a hybrid catalyst hematite/cuprous sulfide with enhanced FeIII/FeII redox cycling.

Surfactants are recalcitrant compounds that require advanced treatment for their degradation. Heterogeneous advanced oxidation processes (AOPs) using iron-based catalysts can be a promising method for surfactant degradation. The acceleration of the FeIII/FeII redox cycling is the key to enhance the catalytic degradation. Herein, a hybrid catalyst composed of α-Fe2O3 and Cu2S was synthesized to improve the reduction of FeIII in a heterogeneous persulfate-AOP system. The results of XRD, Raman and TEM demonstrated the successful preparation of the hybrid catalyst. Because of the optimized FeII regeneration, the AOP containing the catalyst FC75 achieved 100.0% removal of sodium dodecyl sulfate (SDS) in a neutral aquatic environment, significantly higher than 22.9 ± 2.4% with pure α-Fe2O3 or 39.6 ± 2.5% with pure Cu2S. The catalyst FC75 demonstrated effective SDS removal in the recycling test (82.7 ± 7.0% after six recycling test) and in actual wastewater (84.4 ± 4.5%). The regeneration of FeII was confirmed by the increased proportion of FeII from 39.5% in the fresh catalyst to 42.6% in the used catalyst. The main active species was revealed to be sulfate radicals under an acidic condition and shifted to hydroxyl radicals under a basic condition. In the hybrid catalyst, α-Fe2O3 provided FeII to activate persulfate to radicals, with an oxidation product of FeIII, which was then reduced to FeII by CuI provided by Cu2S, coupling with the oxidation of CuI to CuII. The S element in Cu2S could directly or indirectly facilitate the FeIII/FeII redox cycling as an electron donor. Those results have demonstrated that the developed hybrid catalyst is able to promote FeII regeneration for effective SDS removal.

Hematite/selenium disulfide hybrid catalyst for enhanced Fe(III)/Fe(II) redox cycling in advanced oxidation processes.

Regeneration of Fe(II) is a key issue for heterogeneous advanced oxidation processes (AOPs) using iron-based catalysts. Herein, a hybrid catalyst was developed from α-Fe2O3 and SeS2 to enhance the Fe(III)/Fe(II) redox cycling in both hydrogen peroxide (H2O2) system and persulfate (PS) system. The regeneration of Fe(II) was evidenced by the increased Fe(II)/Fe(III) ratio in the used catalyst (205.8% in the H2O2 system or 125.4% in the PS system), compared to 68.4% in the fresh hybrid catalyst Fe/Se-3. Methyl orange was used as a model pollutant to evaluate the degradation performance of the hybrid catalyst. Owing to the promotion of Fe(II) regeneration, Fe/Se-3 achieved a pollutant removal efficiency of 100.0% in 12 min in both systems, significantly higher than that with pure α-Fe2O3 (33.9 ± 3.6% in the H2O2 system or 30.7 ± 2.8% in the PS system). The dominant active species were identified as hydroxyl radicals in the H2O2 system and sulfate radicals in the PS system. In the proposed mechanism, soluble and surface-bound Fe species are provided by α-Fe2O3 to activate H2O2 or PS to radicals, and SeS2 participates in the reactions via Se(IV) reducing Fe(III) to Fe(II) and S atoms being released through protonation to expose more active Se sites.

Self-Assembled Networks for Regulating Lithium Polysulfides in Lithium-Sulfur Batteries.

Inhibiting the shuttle effect caused by soluble lithium polysulfides (LiPSs) is of importance for lithium-sulfur (Li-S) batteries. Here, a strategy was developed to construct protective layers by self-assembly networks to regulate the LiPSs. 2,5-Dichloropyridine (25DCP) holds two kinds of functional groups. Among them, the two C-Cl bonds were nucleophilic substituted by S in LiPSs to form long chains. The pyridine N interacted with Li in other LiPSs via Li bonds to form a short chain. As a result, the long chains were cross-linked by the short chain to form an insoluble network. The as-prepared network covered the sulfur electrode interface to suppress the shuttle effect of the subsequently generated LiPSs. Furthermore, 25DCP improved the redox dynamics by changing the energy level and electronic structure of the sulfur species. Therefore, the Li-S batteries with 25DCP exhibited good electrochemical performance. This work provides a feasible strategy for regulating the LiPSs.

Characteristics of microbial denitrification under different aeration intensities: Performance, mechanism, and co-occurrence network.

This study aimed to explore how dissolved oxygen (DO) affected the characteristics and mechanisms of denitrification in mixed bacterial consortia. We analyzed denitrification efficiency, intracellular nicotinamide adenine dinucleotide (NADH), relative expression of functional genes, and potential co-occurrence network of microorganisms. Results showed that the total nitrogen (TN) removal rates at different aeration intensities (0.00, 0.25, 0.63, and 1.25 L/(L·min)) were 0.93, 1.45, 0.86, and 0.53 mg/(L·min), respectively, which were higher than previously reported values for pure culture. The optimal aeration intensity was 0.25 L/(L·min), at which the maximum NADH accumulation rate and highest relative abundance of napA, nirK, and nosZ were achieved. With increased aeration intensity, the amount of electron flux to nitrate decreased and nitrate assimilation increased. On one hand, nitrate reduction was primarily inhibited by oxygen through competition for electron donors of a certain single strain. On the other hand, oxygen was consumed rapidly by bacteria by stimulating carbon metabolism to create an optimal denitrification niche for denitrifying microorganisms. Denitrification was performed via inter-genus cooperation (competitive interactions and symbiotic relationships) between keystone taxa (Azoarcus, Paracoccus, Thauera, Stappia, and Pseudomonas) and other heterotrophic bacteria (OHB) in aeration reactors. However, in the non-aeration case, which was primarily carried out based on intra-genus syntrophy within genus Propionivibrio, the co-occurrence network constructed the optimal niche contributing to the high TN removal efficiency. Overall, this study enhanced our knowledge about the molecular ecological mechanisms of aerobic denitrification in mixed bacterial consortia and has theoretical guiding significance for further practical application.

Simultaneous ammonium and sulfate biotransformation driven by aeration: Nitrogen/sulfur metabolism and metagenome-based microbial ecology.

The present study aimed to clarify the effect of oxygen respiration on biotransformation of alternative electron acceptors (e.g., nitrate and sulfate) underlying the simultaneous removal of ammonium and sulfate in a single aerated sequencing batch reactor. Complete nitrification was achieved in feast condition, while denitrification was carried out in both feast and famine conditions when aeration intensity (AI) was higher than 0.22 L/(L·min). Reactors R1 [0.56 L/(L·min)], R2 [0.22 L/(L·min)], and R3 [0.08 L/(L·min)] achieved 72.39% sulfate removal efficiency in feast condition, but H2S release occurred in R3. Following exogenous substrate depletion, sulfate concentration increased again and exceeded the influent value in R1, indicating that sulfate transformation was affected by oxygen intrusion. Metagenomic analysis showed that a higher AI promoted sulfate reduction by switching from dissimilatory to assimilatory pathway. Lower AI-acclimated microorganisms (R3) produced H2S and ammonium, while higher AI-acclimated microorganisms (R1) accumulated nitrite, which confirmed that biotransformation of N and S was strongly regulated by redox imbalance driven by aeration. This implied that respiration control, a microbial self-regulation mechanism, was linked to the dynamic imbalance between electron donors and electron acceptors. Aerobic nitrate (sulfate) reduction, as one of the effects of respiration control, could be used as an alternative strategy to compensate for dynamic imbalance, when supported by efficient endogenous metabolism. Moderate aeration induced microorganisms to change their energy conservation and survival strategy through respiration control and inter-genus protection of respiratory activity among keystone taxa (including Azoarcus in R1, Thauera in R2, and Thiobacillus, Ottowia, and Geoalkalibacter in R3) to form an optimal niche in response to oxygen intrusion and achieve benign biotransformation of C, N, and S without toxic intermediate accumulation. This study clarified the biotransformation mechanism of ammonium and sulfate driven by aeration and provided theoretical guidance for optimizing existing aeration-based techniques.

Preliminary evidence from a multicenter prospective observational study of the safety and efficacy of chloroquine for the treatment of COVID-19.

Effective therapies are urgently needed for the SARS-CoV-2 pandemic. Chloroquine has been proved to have antiviral effect against coronavirus in vitro. In this study, we aimed to assess the efficacy and safety of chloroquine with different doses in COVID-19. In this multicenter prospective observational study, we enrolled patients older than 18 years old with confirmed SARS-CoV-2 infection excluding critical cases from 12 hospitals in Guangdong and Hubei Provinces. Eligible patients received chloroquine phosphate 500 mg, orally, once (half dose) or twice (full dose) daily. Patients treated with non-chloroquine therapy were included as historical controls. The primary endpoint is the time to undetectable viral RNA. Secondary outcomes include the proportion of patients with undetectable viral RNA by day 10 and 14, hospitalization time, duration of fever, and adverse events. A total of 197 patients completed chloroquine treatment, and 176 patients were included as historical controls. The median time to achieve an undetectable viral RNA was shorter in chloroquine than in non-chloroquine (absolute difference in medians -6.0 days; 95% CI -6.0 to -4.0). The duration of fever is shorter in chloroquine (geometric mean ratio 0.6; 95% CI 0.5 to 0.8). No serious adverse events were observed in the chloroquine group. Patients treated with half dose experienced lower rate of adverse events than with full dose. Although randomized trials are needed for further evaluation, this study provides evidence for safety and efficacy of chloroquine in COVID-19 and suggests that chloroquine can be a cost-effective therapy for combating the COVID-19 pandemic.

Facile construction of reduced graphene oxide supported three-dimensional polyaniline/WO2.72 nanobelt-flower as a full solar spectrum light response catalyst for efficient photocatalytic conversion of bromate.

As a carcinogenic byproduct in drinking water treatment, bromate has raised global concerns on environmental and health hazard, calling for effective treatments. In the current work, a novel reduced graphene oxide supported polyaniline/WO2.72 nanobelt-flower (RGO/PANI/WO2.72) ternary composite was prepared through a solvent volatilization method for photocatalytic reduction of bromate. The prepared sample was characterized, and the influence of aqueous pH, ions and dissolved oxygen on the bromate reduction was explored. As expected, the introduction of RGO and PANI on the WO2.72 exhibited great synergistic effects on the separation of photogenerated carriers. The calculated reduction rate constant of RGO/PANI/WO2.72 was about six times as high as that of pure WO2.72. Specially, the prepared photocatalyst possessed strong optical absorption in a broad range of 250-2500 nm, and thus displaying excellent catalytic performance in utilization of all solar spectrum energy. Moreover, the RGO/PANI/WO2.72 exhibited stable photocatalytic activity in cycling test. Considered holistically, the present study offered a valuable approach for rational construction of heterogeneous structure in the development of bromate-catalyzed reduction.

An individual patient data meta-analysis on the effect of chemotherapy on survival in patients with craniofacial osteosarcoma.

Chemotherapy improves the survival of patients with long bone osteosarcomas. However, the benefits of chemotherapy in the treatment of craniofacial osteosarcoma (CFOS) are still controversial. We searched PubMed and EMBASE from February 1997 to December 2016 to identify studies on CFOS. The individual patient data of these studies were pooled into a meta-analysis. Univariate and multivariate survival analyses were performed. Thirteen studies with a total of 184 patients met our inclusion criteria. Positive resection margin was a poor prognostic factor for CFOS in the univariate and multivariate survival analyses. Chemotherapy improved overall survival (OS) and disease-specific survival (DSS) in patients with CFOS who had tumors in the maxilla, positive resection margins, or high-grade tumors. Patients with local tumor recurrence had better OS and DSS when treated with chemotherapy. Chemotherapy improves survival in patients with CFOS with adverse factors, such as tumors with positive margins, high-grade tumors, and recurrent tumors.

The design of a polyaniline-decorated three dimensional W18O49 composite for full solar spectrum light driven photocatalytic removal of aqueous nitrite with high N2 selectivity.

Photocatalysis using solar energy is the most promising green technology for nitrite removal. However, effective photocatalytic performance is often challenged by the limited light absorption, utilization of expensive noble metals and undesired products (nitrate and ammonium). Here, we report for the first time that a full solar light response polyaniline-decorated three dimensional W18O49 composite (PANI@W18O49), a noble metal-free photocatalyst, possesses excellent photocatalytic activity for aqueous nitrite removal with high N2 selectivity. The prepared sample was thoroughly identified via XRD, Raman, FTIR, SEM, TEM, UV-vis DRS and PL. The catalytic results demonstrated that over 80% N2 selectivity (initial concentration 1.0 mM) was achieved through the PANI@W18O49 without sacrificial agent under 300 W Xe lamp irradiation for 60 min. Such advantages were attributed to the built-in junction between n-type W18O49 and p-type PANI, offering suitable redox levels of electron-hole pairs for NO2- reaction. The modification of PANI also benefited the light harvesting ability and activated carriers migration, the calculated rate constant of PANI@W18O49 is about four times as high as that of W18O49. The current study not only prepared a promising photocatalyst, but also provides new insights into improving the photocatalytic activity and N2 selectivity for nitrite treatment.

A chair-type G-quadruplex structure formed by a human telomeric variant DNA in K+ solution.

Guanine tracts of human telomeric DNA sequences are known to fold into eight different four-stranded structures that vary by the conformation of guanine nucleotides arranged in the stack of G-tetrads in their core and by different kinds and orders of connecting loops, called G-quadruplexes. Here, we present a novel G-quadruplex structure formed in K+ solution by a human telomeric variant d[(GGGTTA)2GGGTTTGGG], htel21T18. This variant DNA is located in the subtelomeric regions of human chromosomes 8, 11, 17, and 19 as well as in the DNase hypersensitive region and in the subcentromeric region of chromosome 5. Interestingly, single A18T substitution that makes htel21T18 different from the human telomeric sequence results in the formation of a three-layer chair-type G-quadruplex, a fold previously unknown among human telomeric repeats, with two loops interacting through the reverse Watson-Crick A6·T18 base pair. The loops are edgewise; glycosidic conformation of guanines is syn·anti·syn·anti around each tetrad, and each strand of the core has two antiparallel adjacent strands. Our results expand the repertoire of known G-quadruplex folding topologies and may provide a potential target for structure-based anticancer drug design.

Investigation of cancer-associated fibroblasts and p62 expression in oral cancer before and after chemotherapy.

PURPOSE: The aim of this study is to investigate the expression of the autophagy protein p62 in oral squamous cell carcinoma (OSCC) cells before and after chemotherapy. We also detected cancer-associated fibroblasts (CAFs) in these OSCC samples to explore the roles of p62 and CAFs in chemotherapy. MATERIALS AND METHODS: Immunohistochemistry was used to analyze the expression of p62 and α-SMA in 26 paired OSCC samples before and after chemotherapy. The relationships between clinicopathological features, clinical outcome and the expression of these proteins were analyzed. RESULTS: Our results indicated an increased stromal α-SMA expression after chemotherapy in OSCC samples. High p62 expression of OSCC cells closely correlated with stromal α-SMA expression after chemotherapy. Furthermore, the post-chemotherapy p62 expression was associated with the prognosis for OSCC patients. CONCLUSION: These results suggest that chemotherapy may increase CAFs in OSCC. High cytoplasmic p62 expression may serve as a poor prognostic marker for OSCC patients.

Characterizations of distinct parallel and antiparallel G-quadruplexes formed by two-repeat ALS and FTD related GGGGCC sequence.

The large expansion of GGGGCC (G4C2) repeats of the C9orf72 gene have been found to lead to the pathogenesis of devastating neurological diseases, amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). The structural polymorphisms of C9orf72 HRE DNA and RNA may cause aberrant transcription and contribute to the development of ALS and FTD. Here we showed that the two-repeat G4C2 DNA, d(G4C2)2, simultaneously formed parallel and antiparallel G-quadruplex conformations in the potassium solution. We separated different folds of d(G4C2)2 by anion exchange chromatography, followed with characterizations by circular dichroism and nuclear magnetic resonance spectroscopy. The parallel d(G4C2)2 G-quadruplex folded as a symmetric tetramer, while the antiparallel d(G4C2)2 adopted the topology of an asymmetric dimer. These folds are distinct from the antiparallel chair-type conformation we previously identified for the d(G4C2)4 G-quadruplex. Our findings have demonstrated the conformational heterogeneity of the C9orf72 HRE DNA, and provided new insights into the d(G4C2)n folding. Meanwhile, the purified d(G4C2)2 G-quadruplex samples are suitable for further three-dimensional structure characterizations, which are required for the structure-based design of small molecules targeting ALS and FTD related C9orf72 HRE.