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The ubiquitin-specific protease 7 (USP7), as a member of deubiquitination enzymes, represents an attractive therapeutic target for various cancers, including prostate cancer and liver cancer. The change of the inhibitor stereocenter from the S to R stereochemistry (S-ALM → R-ALM34) markedly improved USP7 inhibitory activity. However, the molecular mechanism for the stereo-selectivity of enantiomeric inhibitors to USP7 is still unclear. In this work, molecular docking, molecular dynamics (MD) simulations, molecular mechanics/Generalized-Born surface area (MM/GBSA) calculations, and free energy landscapes were performed to address this mystery. MD simulations revealed that S-ALM34 showed a high degree of conformational flexibility compared to the R-ALM34 counterpart, and S-ALM34 binding led to the enhanced intradomain motions of USP7, especially the BL1 and BL2 loops and the two helices α4 and α5. MM/GBSA calculations showed that the binding strength of R-ALM34 to USP7 was stronger than that of S-ALM34 by - 4.99 kcal/mol, a similar trend observed by experimental data. MM/GBSA free energy decomposition was further performed to differentiate the ligand-residue spectrum. These analyses not only identified the hotspot residues interacting with R-ALM34, but also revealed that the hydrophobic interactions from F409, K420, H456, and Y514 play the major determinants in the binding of R-ALM34 to USP7. This result is anticipated to shed light on energetic basis and conformational dynamics information to aid in the design of more potent and selective inhibitors targeting USP7., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Citrus is one of the most important fruit crop genera in the world, but many Citrus species are vulnerable to cold stress. Ichang papeda (Citrus ichangensis), a cold-hardy citrus species, holds great potential for identifying valuable metabolites that are critical for cold tolerance in Citrus. However, the metabolic changes and underlying mechanisms that regulate Ichang papeda cold tolerance remain largely unknown. In this study, we compared the metabolomes and transcriptomes of Ichang papeda and HB pummelo (Citrus grandis "Hirado Buntan", a cold-sensitive species) to explore the critical metabolites and genes responsible for cold tolerance. Metabolomic analyses led to the identification of common and genotype-specific metabolites, consistent with transcriptomic alterations. Compared to HB pummelo under cold stress, Ichang papeda accumulated more sugars, flavonoids, and unsaturated fatty acids, which are well-characterized metabolites involved in stress responses. Interestingly, sphingosine and chlorogenic acid substantially accumulated only in Ichang papeda. Knockdown of CiSPT (C. ichangensis serine palmitoyltransferase) and CiHCT2 (C. ichangensis hydroxycinnamoyl-CoA: shikimate hydroxycinnamoyltransferase2), two genes involved in sphingosine and chlorogenic acid biosynthesis, dramatically decreased endogenous sphingosine and chlorogenic acid levels, respectively. This reduction in sphingosine and chlorogenic acid notably compromised the cold tolerance of Ichang papeda, whereas exogenous application of these metabolites increased plant cold tolerance. Taken together, our findings indicate that greater accumulation of a spectrum of metabolites, particularly sphingosine and chlorogenic acid, promotes cold tolerance in cold-tolerant citrus species. These findings broaden our understanding of plant metabolic alterations in response to cold stress and provide valuable targets that can be manipulated to improve Citrus cold tolerance., Competing Interests: Conflict of interest statement. The authors declare no conflicts of interest associated with this work., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Dark photons have emerged as promising candidates for dark matter, and their search is a top priority in particle physics, astrophysics, and cosmology. We report the first use of a tunable niobium superconducting radio-frequency cavity for a scan search of dark photon dark matter with innovative data analysis techniques. We mechanically adjusted the resonant frequency of a cavity submerged in liquid helium at a temperature of 2 K, and scanned the dark photon mass over a frequency range of 1.37 MHz centered at 1.3 GHz. Our study leveraged the superconducting radio-frequency cavity's remarkably high quality factors of approximately 10^{10}, resulting in the most stringent constraints to date on a substantial portion of the exclusion parameter space on the kinetic mixing coefficient ε between dark photons and electromagnetic photons, yielding a value of ε<2.2×10^{-16}.

Vacuolar malic acid accumulation largely determines fruit acidity, a key trait for the taste and flavor of apple and other fleshy fruits. Aluminum-activated malate transporter 9 (ALMT9/Ma1) underlies a major genetic locus, Ma, for fruit acidity in apple, but how the protein transports malate across the tonoplast is unclear. Here, it is shown that overexpression of the coding sequence of Ma1 (Ma1α) drastically decreases fruit acidity in "Royal Gala" apple, leading to uncovering alternative splicing underpins Ma1's function. Alternative splicing generates two isoforms: Ma1β is 68 amino acids shorter with much lower expression than the full-length protein Ma1α. Ma1β does not transport malate itself but interacts with the functional Ma1α to form heterodimers, creating synergy with Ma1α for malate transport in a threshold manner (When Ma1β/Ma1α ≥ 1/8). Overexpression of Ma1α triggers feedback inhibition on the native Ma1 expression via transcription factor MYB73, decreasing the Ma1β level well below the threshold that leads to significant reductions in Ma1 function and malic acid accumulation in fruit. Overexpression of Ma1α and Ma1β or genomic Ma1 increases both isoforms proportionally and enhances fruit malic acid accumulation. These findings reveal an essential role of alternative splicing in ALMT9-mediated malate transport underlying apple fruit acidity., (© 2024 The Authors. Advanced Science published by Wiley‐VCH GmbH.)

Exploiting effective, stable, and cost-efficient electrocatalysts for the water oxidation reaction is highly desirable for renewable energy conversion techniques. Constructional design and compositional manipulation are widely used approaches to efficaciously boost the electrocatalytic performance. Herein, we designed a NiFe-bimetallic sulfide/N-doped carbon composite via a two-step thermal treatment of Prussian blue analogues/cellulose nanofibers (PBA/CNFs) film. The NiFe-bimetallic sulfide/N-doped carbon composite displayed enhanced OER performance in an alkaline environment, with an overpotential of 282 mV at 10 mA cm -2 , a Tafel slope of 59.71 mV dec -1 , and good stability, making the composite a candidate electrocatalyst for OER-related energy equipment. The introduction of CNFs in the precursor prevented the aggregation of PBA nanoparticles (NPs), exposed more active sites, and the resulting carbon substrate enhanced the electroconductivity of the composite. Moreover, the synergistic effect of Ni and Fe in the bimetallic sulfide could modulate the configuration of electrons, enrich the catalytically active sites, and augment the electric conductivity, thus ameliorating the OER performance. This study broadens the application of MOF-CNF composites to construct hierarchical structures of metal compounds and provides some thoughts for the development of cost-effective precious-metal-free catalysts for electrocatalysis.

Vacuole largely dictates the fruit taste and flavor, as most of the sugars and organic acids are stored in the vacuoles of the fruit. However, difficulties associated with vacuole separation severely hinder identification and characterization of vacuolar proteins in fruit species. In this study, we established an effective approach for separating vacuoles and successfully purified vacuolar protein from six types of citrus fruit with varying patterns of sugar and organic acid contents. By using label-free LC-MS/MS proteomic analysis, 1443 core proteins were found to be associated with the essential functions of vacuole in citrus fruit. Correlation analysis of metabolite concentration with proteomic data revealed a transporter system for the accumulation of organic acid and soluble sugars in citrus. Furthermore, we characterized the physiological roles of selected key tonoplast transporters, ABCG15, Dict2.1, TMT2, and STP7 in the accumulation of citric acid and sugars. These findings provide a novel perspective and practical solution for investigating the transporters underlying the formation of citrus taste and flavor., Competing Interests: The authors declare no conflict of interest., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nanjing Agricultural University.)

The orange subfamily (Aurantioideae) contains several Citrus species cultivated worldwide, such as sweet orange and lemon. The origin of Citrus species has long been debated and less is known about the Aurantioideae. Here, we compiled the genome sequences of 314 accessions, de novo assembled the genomes of 12 species and constructed a graph-based pangenome for Aurantioideae. Our analysis indicates that the ancient Indian Plate is the ancestral area for Citrus-related genera and that South Central China is the primary center of origin of the Citrus genus. We found substantial variations in the sequence and expression of the PH4 gene in Citrus relative to Citrus-related genera. Gene editing and biochemical experiments demonstrate a central role for PH4 in the accumulation of citric acid in citrus fruits. This study provides insights into the origin and evolution of the orange subfamily and a regulatory mechanism underpinning the evolution of fruit taste., (© 2023. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Sugar transporter proteins (STPs) play vital roles in sugar transport and allocation of carbon sources in plants. However, the evolutionary dynamics of this important gene family and their functions are still largely unknown in citrus, which is the largest fruit crop in the world. In this study, fourteen non-redundant CgSTP family members were identified in pummelo ( Citrus grandis ). A comprehensive analysis based on the biochemical characteristics, the chromosomal location, the exon-intron structures and the evolutionary relationships demonstrated the conservation and the divergence of CgSTPs. Moreover, CgSTP4, 11, 13, 14 were proofed to be localized in plasma membrane and have glucose transport activity in yeast. The hexose content were significantly increased with the transient overexpression of CgSTP11 and CgSTP14 . In addition, antisense repression of CgSTP4 induced the shorter pollen tube length in vitro , implying the potential role of CgSTP4 in pummelo pollen tube growth. Taken together, this work explored a framework for understanding the physiological role of CgSTPs and laid a foundation for future functional studies of these members in citrus species., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Xu, Liu, Zhao, Zhang, Li, Guo, Liu and Li.)

β-Amylase (BAM)-mediated starch degradation is a main source of soluble sugars that help plants adapt to environmental stresses. Here, we demonstrate that dehydration-induced expression of PtrBAM3 in trifoliate orange (Poncirus trifoliata (L.) Raf.) functions positively in drought tolerance via modulation of starch catabolism. Two transcription factors, PtrABF4 (P. trifoliata abscisic acid-responsive element-binding factor 4) and PtrABR1 (P. trifoliata ABA repressor 1), were identified as upstream transcriptional activators of PtrBAM3 through yeast one-hybrid library screening and protein-DNA interaction assays. Both PtrABF4 and PtrABR1 played a positive role in plant drought tolerance by modulating soluble sugar accumulation derived from BAM3-mediated starch decomposition. In addition, PtrABF4 could directly regulate PtrABR1 expression by binding to its promoter, leading to a regulatory cascade to reinforce the activation of PtrBAM3. Moreover, PtrABF4 physically interacted with PtrABR1 to form a protein complex that further promoted the transcriptional regulation of PtrBAM3. Taken together, our finding reveals that a transcriptional cascade composed of ABF4 and ABR1 works synergistically to upregulate BAM3 expression and starch catabolism in response to drought condition. The results shed light on the understanding of the regulatory molecular mechanisms underlying BAM-mediated soluble sugar accumulation for rendering drought tolerance in plants., (© American Society of Plant Biologists 2022. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

There is a continuously rising incidence of non-alcoholic fatty liver disease (NAFLD) around the world, which parallels the increasing incidence of metabolic diseases. NAFLD is a range of liver conditions that contains simple non-alcoholic fatty liver and advanced non-alcoholic steatohepatitis. In serious cases, NAFLD may develop into cirrhosis or even liver cancer. NAFLD has an intense relationship with metabolic syndrome, type 2 diabetes mellitus. It is known that gut microbiota, and functional molecules such as adenosine monophosphate-activated protein kinase JNK, and peroxisome proliferator-activated receptors (PPARs) in progressing and treating NAFLD. Traditionally, the conventional and effective therapeutic strategy is lifestyle intervention. Nowadays, new medicines targeting specific molecules, such as farnesoid X receptor, PPARs, and GLP-1 receptor, have been discovered and shown beneficial effects on patients with NAFLD. In this article, we focus on the molecular mechanisms and therapeutic approaches to NAFLD., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Che, Zhao, Li, Li, Cho and Yu.)

Introduction: In recent years, people have gained a profound understanding of chronic insomnia disorder (CID), but the pathophysiological mechanism of CID is still unclear. There is some evidence that the locus coeruleus (LC) is involved in the regulation of wakefulness in CID, but there have been few studies using brain functional imaging. The purpose of this study was to evaluate the resting-state functional connectivity (FC) between the LC and other brain voxels in CID and whether these abnormal FC are involved in the regulation of wakefulness., Methods: A total of 49 patients with chronic insomnia disorder and 47 healthy controls (HC) matched for gender, age, and education were examined with rs-fMRI in this study. The LC was selected as the region of interest, and then seed-based analysis was conducted on the LC and other voxels to obtain the brain regions with abnormal FC. The correlation between the FC value of the abnormal connection area and the clinical scale score was analyzed., Results: Compared with the HC, the FC between the LC and right precuneus, right posterior cingulate cortex, left middle temporal gyrus, left calcarine, and right superior orbitofrontal cortex was significantly enhanced (p < 0.05, FDR correction), and the functional connectivity signal value between the locus coeruleus and left middle temporal gyrus was positively correlated with the Self-Rating Depression Scale (p = 0.021)., Conclusion: The abnormal FC between the LC and multiple brain regions may contribute to a better understanding of the neurobiological mechanism of CID., Competing Interests: We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the position presented in, or the review of., (© 2022 Li et al.)

The phytohormone abscisic acid (ABA) regulates ion channel activity and stomatal movement in response to drought stress. Cellular ABA levels change depending on cellular and environmental conditions via modulation of its biosynthesis, catabolism and transport. Although factors involved in ABA biosynthesis and degradation have been studied extensively, how ABA transporters are modulated to fine-tune ABA levels, especially under drought stress, remains elusive. Here, we show that Arabidopsis thaliana SORTING NEXIN 2 (SNX2) proteins play a critical role in endosomal trafficking of the ABA exporter ATP BINDING CASETTE G25 (ABCG25) via direct interaction at endosomes, leading to its degradation in the vacuole. In agreement, snx2a and snx2b mutant plants showed enhanced recycling of GFP-ABCG25 from early endosomes to the plasma membrane and higher accumulation of GFP-ABCG25. Phenotypically, snx2a and snx2b plants were highly sensitive to exogenous ABA and displayed enhanced ABA-mediated inhibition of inward K + currents and ABA-mediated activation of slow anion currents in guard cells, resulting in an increased tolerance to drought stress. Based on these results, we propose that SNX2 proteins play a crucial role in stomatal movement and tolerance to drought stress by modulating the endosomal trafficking of ABCG25 and thus cellular ABA levels., (© 2022 Society for Experimental Biology and John Wiley & Sons Ltd.)

Voltage controlled magnetic anisotropy (VCMA) has been considered as an effective method in traditional magnetic devices with lower power consumption. In this article, we have investigated the dual-axis control of magnetic anisotropy in Co 2 MnSi/GaAs/PZT hybrid heterostructures through piezo-voltage-induced strain using longitudinal magneto-optical Kerr effect (LMOKE) microscopy. The major modification of in-plane magnetic anisotropy of the Co 2 MnSi thin film is controlled obviously by the piezo-voltages of the lead zirconate titanate (PZT) piezotransducer, accompanied by the coercivity field and magnetocrystalline anisotropy significantly manipulated. Because in-plane cubic magnetic anisotropy and uniaxial magnetic anisotropy coexist in the Co 2 MnSi thin film, the initial double easy axes of cubic split to an easiest axis (square loop) and an easier axis (two-step loop). While the stress direction is parallel to the [1-10] easiest axis (sample I), the square loop of the [1-10] direction could transform to a two-step loop under the negative piezo-voltages (compressed state). At the same time, the initial two-step loop of the [110] axis simultaneously changes to a square loop (the easiest axis). Otherwise, we designed and fabricated the sample II in which the PZT stress is parallel to the [110] two-step axis. The phenomenon of VCMA was also obtained along the [110] and [1-10] directions. However, the manipulated results of sample II were in contrast to those of the sample I under the piezo-voltages. Thus, an effective dual-axis regulation of the in-plane magnetization rotation was demonstrated in this work. Such a finding proposes a more optimized method for the magnetic logic gates and memories based on voltage-controlled magnetic anisotropy in the future., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Lead (Pb)-free halide hybrid materials have received a great deal of attention because of their potential in optoelectronic applications. However, heteroatom-based amine lead-free tin halide hybrid single crystals have not been well investigated yet. Detailed synthetic processes, growth, crystal structures, and stability of (ACH 2 CH 2 NH 3 ) 2 SnBr 6 (A = OH or SH) and (BCH 2 CH 2 NH 3 ) 2 SnI 4 (B = I or SH) single crystals were investigated. Interestingly, (IH 3 NCH 2 CH 2 SSCH 2 CH 2 NH 3 ) 2 HPO 3 exhibited orange-red photoluminescence (PL) at about 620 nm with an average PL lifetime of about 912 ns. (HSCH 2 CH 2 NH 3 ) 2 SnI 4 single crystals exhibited a PL peak at 620 nm with an average PL lifetime of about 0.607 ns. More importantly, (HSCH 2 CH 2 NH 3 ) 2 SnI 4 single crystals exhibited reversible red-black color transformations when exposed to a H 3 PO 2 solution and an ambient atmosphere, which was attributed to oxidation from Sn 2+ to Sn 4+ , rather than from I - to I 3 - (I 2 ). The intriguing characteristics should provide guidance for further optoelectronic applications of these Pb-free halide hybrid materials.

Solar and wind energy harvesting technology is increasingly an economical and efficient energy form and receives excellent support from government policies worldwide. Various functional and structural nanogenerators based on multi-effects named hybridized nanogenerators have been reported separately or simultaneously to effectively generate the wasted mechanical and solar energy in our daily life. We review the development of hybridized nanogenerators, including the working mechanism of solar and mechanical energies. Moreover, the classification of nanogenerators for scavenging mechanical and solar energies is discussed. The potential applications of hybridized nanogenerators are reviewed. Finally, the challenge and prospective of hybridized nanogenerators and the future explored improvements of output performance, stability, preparation, large-scale utilizing, and efficiency are discussed. The hybridized nanogenerator as the energy technology will be popularized in energy and self-powered sensor systems., Competing Interests: The authors declare no competing interests., (© 2021 The Authors.)

Malate accumulation in the vacuole largely determines apple ( Malus domestica ) fruit acidity, and low fruit acidity is strongly associated with truncation of Ma1 , an ortholog of ALUMINUM-ACTIVATED MALATE TRANSPORTER9 ( ALMT9 ) in Arabidopsis ( Arabidopsis thaliana ). A mutation at base 1,455 in the open reading frame of Ma1 leads to a premature stop codon that truncates the protein by 84 amino acids at its C-terminal end. Here, we report that both the full-length protein, Ma1, and its naturally occurring truncated protein, ma1, localize to the tonoplast; when expressed in Xenopus laevis oocytes and Nicotiana benthamiana cells, Ma1 mediates a malate-dependent inward-rectifying current, whereas the ma1-mediated transmembrane current is much weaker, indicating that ma1 has significantly lower malate transport activity than Ma1. RNA interference suppression of Ma1 expression in 'McIntosh' apple leaves, 'Empire' apple fruit, and 'Orin' apple calli results in a significant decrease in malate level. Genotyping and phenotyping of 186 apple accessions from a diverse genetic background of 17 Malus species combined with the functional analyses described above indicate that Ma1 plays a key role in determining fruit acidity and that the truncation of Ma1 to ma1 is genetically responsible for low fruit acidity in apple. Furthermore, we identified a C-terminal domain conserved in all tonoplast-localized ALMTs essential for Ma1 function; protein truncations into this conserved domain significantly lower Ma1 transport activity. We conclude that the truncation of Ma1 to ma1 reduces its malate transport function by removing a conserved C-terminal domain, leading to low fruit acidity in apple., (© 2020 American Society of Plant Biologists. All Rights Reserved.)

Rapid pollen tube growth requires uptake of Suc or its hydrolytic products, hexoses, from the apoplast of surrounding tissues in the style. Due to species-specific sugar requirements, reliance of pollen germination and tube growth on cell wall invertase and Suc or hexose transporters varies between species, but it is not known if plants have a sugar transporter that mediates the uptake of both hexose and Suc for pollen tube growth. Here, we show that a sugar transporter protein in apple ( Malus domestica ), MdSTP13a, takes up both hexose and Suc when expressed in yeast, and is essential for pollen tube growth on Glc and Suc but not on maltose. MdSTP13a-mediated direct uptake of Suc is primarily responsible for apple pollen tube growth on Suc medium. Sorbitol, a major photosynthate and transport carbohydrate in apple, modulates pollen tube growth via the MYB transcription factor MdMYB39L, which binds to the promoter of MdSTP13a to activate its expression. Antisense repression of MdSTP13a blocks sorbitol-modulated pollen tube growth. These findings demonstrate that MdSTP13a takes up both hexose and Suc for sorbitol-modulated pollen tube growth in apple, revealing a situation where acquisition of sugars for pollen tube growth is regulated by a sugar alcohol., (© 2020 American Society of Plant Biologists. All rights reserved.)

The aim of current study was to investigate the metabolic changes associated with histidine supplementation in serum and urine metabolic signatures and serum amino acid (AA) profiles. Serum and urine 1 H NMR-based metabolomics and serum AA profiles were employed in 32 and 37 obese women with metabolic syndrome (MetS) intervened with placebo or histidine for 12 weeks. Multivariable statistical analysis were conducted to define characteristic metabolites. In serum 1 H NMR metabolic profiles, increases in histidine, glutamine, aspartate, glycine, choline, and trimethylamine-N-oxide (TMAO) were observed; meanwhile, decreases in cholesterol, triglycerides, fatty acids and unsaturated lipids, acetone, and α/β-glucose were exhibited after histidine supplement. In urine 1 H NMR metabolic profiles, citrate, creatinine/creatine, methylguanidine, and betaine + TMAO were higher, while hippurate was lower in histidine supplement group. In serum AA profiles, 10 AAs changed after histidine supplementation, including increased histidine, glycine, alanine, lysine, asparagine, and tyrosine and decreased leucine, isoleucine, ornithine, and citrulline. The study showed a systemic metabolic response in serum and urine metabolomics and AA profiles to histidine supplementation, showing significantly changed metabolism in AAs, lipid, and glucose in obese women with MetS.

Mid-infrared (MIR) nonlinear optical crystals of LiInSe 2 (LISe) were grown by a modified Bridgman technique on a (001)-seed. A 7-12 μm widely tunable picosecond (ps) MIR optical parametric amplifier (OPA) based on a LISe crystal was demonstrated for the first time, to the best of our knowledge. The MIR OPA was pumped by a 30 ps 1064 nm Nd:YAG laser and injected by a KTiOPO 4 (KTP)-based widely tunable near-infrared seed. The idler operating at 7.5 μm with the highest pulse energy of 170 μJ was obtained under a pump energy of 14 mJ. The corresponding energy conversion efficiency is ∼1.21%, and the photon conversion efficiency is 8.6%. The output energies were measured to be ∼121  μJ at 7 μm and ∼21  μJ at 12 μm.

Tanshinone IIA (T2A), a derivative of phenanthrenequinone and also the major active ingredient of Danshen, has been paid extensive attention as a promising cancer therapy for its potential anti-cancer activities. In this study, the apoptosis and autophagy of human prostate cancer PC-3 cells were observed after 5 μM T2A treatment, as well as their relevance. Mitochondrial-dependent apoptosis was firstly detected through morphological observation and biochemical analysis. Meanwhile, 5 μM T2A successfully triggered the autophagy of PC-3 cells, indicated by increased expression of Beclin1, and LC3 II. Validation experiments were conducted to further consolidate T2A's contribution to autophagy: Pretreatment with autophagy inhibitor 3-methyladenine (3-MA) provided protection against autophagy and enhanced T2A-induced apoptosis. Besides, the apoptosis suppressor z-VAD-fmk failed to facilitate the formation of autophagic vacuoles, which also proved the T2A-induced autophagy independent of apoptosis. Moreover, the reactive oxygen species (ROS) scavenger N-acetyl-L-cysteine (NAC) efficiently inhibited the expression of Beclin1, LC3-II, and cleaved caspase-3, which indicated apoptosis and autophagy with dependence on intracellular ROS production. Taken together, these results demonstrated that autophagy is the cytoprotective mechanism in this experimental system, and the ROS resulted from T2A treatment played a critical role in apoptosis and autophagy initiation.