Ring domains are essential for GATOR2-dependent mTORC1 activation.

The GATOR2-GATOR1 signaling axis is essential for amino-acid-dependent mTORC1 activation. However, the molecular function of the GATOR2 complex remains unknown. Here, we report that disruption of the Ring domains of Mios, WDR24, or WDR59 completely impedes amino-acid-mediated mTORC1 activation. Mechanistically, via interacting with Ring domains of WDR59 and WDR24, the Ring domain of Mios acts as a hub to maintain GATOR2 integrity, disruption of which leads to self-ubiquitination of WDR24. Physiologically, leucine stimulation dissociates Sestrin2 from the Ring domain of WDR24 and confers its availability to UBE2D3 and subsequent ubiquitination of NPRL2, contributing to GATOR2-mediated GATOR1 inactivation. As such, WDR24 ablation or Ring deletion prevents mTORC1 activation, leading to severe growth defects and embryonic lethality at E10.5 in mice. Hence, our findings demonstrate that Ring domains are essential for GATOR2 to transmit amino acid availability to mTORC1 and further reveal the essentiality of nutrient sensing during embryonic development.

7-Dehydrocholesterol dictates ferroptosis sensitivity.

Ferroptosis, a form of regulated cell death that is driven by iron-dependent phospholipid peroxidation, has been implicated in multiple diseases, including cancer1-3, degenerative disorders4 and organ ischaemia-reperfusion injury (IRI)5,6. Here, using genome-wide CRISPR-Cas9 screening, we identified that the enzymes involved in distal cholesterol biosynthesis have pivotal yet opposing roles in regulating ferroptosis through dictating the level of 7-dehydrocholesterol (7-DHC)-an intermediate metabolite of distal cholesterol biosynthesis that is synthesized by sterol C5-desaturase (SC5D) and metabolized by 7-DHC reductase (DHCR7) for cholesterol synthesis. We found that the pathway components, including MSMO1, CYP51A1, EBP and SC5D, function as potential suppressors of ferroptosis, whereas DHCR7 functions as a pro-ferroptotic gene. Mechanistically, 7-DHC dictates ferroptosis surveillance by using the conjugated diene to exert its anti-phospholipid autoxidation function and shields plasma and mitochondria membranes from phospholipid autoxidation. Importantly, blocking the biosynthesis of endogenous 7-DHC by pharmacological targeting of EBP induces ferroptosis and inhibits tumour growth, whereas increasing the 7-DHC level by inhibiting DHCR7 effectively promotes cancer metastasis and attenuates the progression of kidney IRI, supporting a critical function of this axis in vivo. In conclusion, our data reveal a role of 7-DHC as a natural anti-ferroptotic metabolite and suggest that pharmacological manipulation of 7-DHC levels is a promising therapeutic strategy for cancer and IRI.

A dandelion-like nanomedicine via hierarchical self-assembly for synergistic chemotherapy and photo-dynamic cancer therapy.

The synergistic effect of chemotherapy and photo-dynamic therapy (PDT) is an effective way to improve the efficiency of tumor treatment. However, most synergistic therapeutic drugs have poor water solubility and stability, so it is difficult to achieve high therapeutic effects while avoiding the severe side effects. Herein, a unique dandelion-like nanomedicine (named as cRGDfk-CCPT-mCe6) was successfully synthesized using Ce6-loaded amphiphilic ß-cyclodextrins (ß-CD) doped lipid-based vesicles as the core (receptacle) and ß-CD modified camptothecin (CPT) pro-drug as the flyable dandelion seeds. The ß-CD modified CPT pro-drug was introduced into the core vesicles in succession via host-guest interaction between inter-molecular ß-CD and CPT, and cRGDfk peptides were further introduced as the outermost layer (stigma) to enhance the internalization into cancer cells. CPT interacted with ß-CD through glutathione (GSH)-cleavable disulfide bonds, which led to drug release in glutathione-rich cancer cells, just as spread of dandelion seeds in the wind. GSH consumption further disrupted the intracellular redox homeostasis of cancer cells through combined action of Ce6 with light irradiation and the synergistic anti-tumor effect was thus achieved, resulting in apoptosis of cancer cells. Therefore, the nanomedicine provides a facile and versatile anti-tumor strategy, as well as a persistent anti-cancer effects.

An acetylation-enhanced interaction between transcription factor Sox2 and the steroid receptor coactivators facilitates Sox2 transcriptional activity and function.

SRY-box 2 (Sox2) is a transcription factor with critical roles in maintaining embryonic stem (ES) cell and adult stem cell functions and in tumorigenesis. However, how Sox2 exerts its transcriptional function remains unclear. Here, we used an in vitro protein-protein interaction assay to discover transcriptional regulators for ES cell core transcription factors (Oct4, Sox2, Klf4, and c-Myc) and identified members of the steroid receptor coactivators (SRCs) as Sox2-specific interacting proteins. The SRC family coactivators have broad roles in transcriptional regulation, but it is unknown whether they also serve as Sox2 coactivators. We demonstrated that these proteins facilitate Sox2 transcriptional activity and act synergistically with p300. Furthermore, we uncovered an acetylation-enhanced interaction between Sox2 and SRC-2/3, but not SRC-1, demonstrating it is Sox2 acetylation that promotes the interaction. We identified putative Sox2 acetylation sites required for acetylation-enhanced interaction between Sox2 and SRC-3 and demonstrated that acetylation on these sites contributes to Sox2 transcriptional activity and recruitment of SRC-3. We showed that activation domains 1 and 2 of SRC-3 both display a preferential binding to acetylated Sox2. Finally, functional analyses in mouse ES cells demonstrated that knockdown of SRC-2/3 but not SRC-1 in mouse ES cells significantly downregulates the transcriptional activities of various Sox2 target genes and impairs ES cell stemness. Taken together, we identify specific SRC family proteins as novel Sox2 coactivators and uncover the role of Sox2 acetylation in promoting coactivator recruitment and Sox2 transcriptional function.

Benzoselenazolium-based hemicyanine dye for G-Quadruplex detection.

Benzothiazolium and benzoxazolium are common groups for the construction of hemicyanine dyes; however, their isosteric analogue benzoselenazolium have rarely been studied. Here, we report the development of the first benzoselenazolium-based hemicyanine dye for the selective detection of G-quadruplexes. This molecule, SEMA-1, was validated as a red-emitting and activatable fluorescent probe whose fluorescence would only be activated in the presence of G-quadruplexes in buffer solution. Consistent with this, SEMA-1 was found to accumulate in nucleoli and could be used to detect the high abundance of nucleolar rDNA and rRNA G-quadruplexes in fixed HeLa cells. On the other hand, due to the retained mitochondrial membrane potential in live HeLa cells, SEMA-1 was captured by mitochondria and had the potential to detect the mitochondrial G-quadruplexes. Collectively, this work demonstrates the value of developing G-quadruplex-specific fluorescent probes from novel benzoselenazolium-based hemicyanine scaffold.

A methylation-phosphorylation switch determines Sox2 stability and function in ESC maintenance or differentiation.

Sox2 is a key factor for maintaining embryonic stem cell (ESS) pluripotency, but little is known about its posttranslational regulation. Here we present evidence that the precise level of Sox2 proteins in ESCs is regulated by a balanced methylation and phosphorylation switch. Set7 monomethylates Sox2 at K119, which inhibits Sox2 transcriptional activity and induces Sox2 ubiquitination and degradation. The E3 ligase WWP2 specifically interacts with K119-methylated Sox2 through its HECT domain to promote Sox2 ubiquitination. In contrast, AKT1 phosphorylates Sox2 at T118 and stabilizes Sox2 by antagonizing K119me by Set7 and vice versa. In mouse ESCs, AKT1 activity toward Sox2 is greater than that of Set7, leading to Sox2 stabilization and ESC maintenance. In early development, increased Set7 expression correlates with Sox2 downregulation and appropriate differentiation. Our study highlights the importance of a Sox2 methylation-phosphorylation switch in determining ESC fate.

Development of a Highly Selective and Sensitive Fluorescent Probe for Imaging RNA Dynamics in Live Cells.

RNA imaging is of great importance for understanding its complex spatiotemporal dynamics and cellular functions. Considerable effort has been devoted to the development of small-molecule fluorescent probes for RNA imaging. However, most of the reported studies have mainly focused on improving the photostability, permeability, long emission wavelength, and compatibility with live-cell imaging of RNA probes. Less attention has been paid to the selectivity and detection limit of this class of probes. Highly selective and sensitive RNA probes are still rarely available. In this study, a new set of styryl probes were designed and synthesized, with the aim of upgrading the detection limit and maintaining the selectivity of a lead probe QUID-1 for RNA. Among these newly synthesized compounds, QUID-2 was the most promising candidate. The limit of detection (LOD) value of QUID-2 for the RNA was up to 1.8 ng/mL in solution. This property was significantly improved in comparison with that of QUID-1. Further spectroscopy and cell imaging studies demonstrated the advantages of QUID-2 over a commercially available RNA staining probe, SYTO RNASelect, for highly selective and sensitive RNA imaging. In addition, QUID-2 exhibited excellent photostability and low cytotoxicity. Using QUID-2, the global dynamics of RNA were revealed in live cells. More importantly, QUID-2 was found to be potentially applicable for detecting RNA granules in live cells. Collectively, our work provides an ideal probe for RNA imaging. We anticipate that this powerful tool may create new opportunities to investigate the underlying roles of RNA and RNA granules in live cells.

Structure-Activity Relationship Investigation on Reaction Mechanism between Chlorinated Quinoid Carcinogens and Clinically-Used Aldoxime Nerve-Agent Antidote under Physiological Condition.

Pyridinium aldoximes are best-known therapeutic antidotes used for clinical treatment of poisonings by organophosphorus nerve-agents and pesticides. Recently, we found that pralidoxime (2-PAM, a currently clinically used nerve-agent antidote) could also detoxify tetrachloro-1,4-benzoquinone (TCBQ), which is a carcinogenic quinoid metabolite of the widely used wood preservative pentachlorophenol under normal physiological conditions, via an unusually mild and facile Beckmann fragmentation mechanism accompanied by radical homolysis. However, it is not clear whether the less-chlorinated benzoquinones (CnBQs, n ≤ 3) act similarly; if so, what is the structure-activity relationship? In this study, we found that (1) The stability of reaction intermediates produced by different CnBQs and 2-PAM was dependent not only on the position but also the degree of Cl-substitution on CnBQs, which can be divided into TCBQ- and DCBQ (dichloro-1,4-benzoquinone)-subgroup; (2) The pKa value of hydroxlated quinones (Cn-1BQ-OHs, the hydrolysis products of CnBQs), determined the stability of corresponding intermediates, that is, the decomposition rate of the intermediates depended on the acidity of Cn-1BQ-OHs; (3) The pKa value of the corresponding Cn-1BQ-OHs could also determine the reaction ratio of Beckmann fragmentation to radical homolysis in CnBQs/2-PAM. These new findings on the structure-activity relationship of the halogenated quinoid carcinogens detoxified by pyridinium aldoxime therapeutic agents via Beckmann fragmentation and radical homolysis reaction may have broad implications on future biomedical and environmental research.

First Report of Anthracnose on Rubus rosaefolius Smith Caused by Colletotrichum boninense in China.

Raspberry (Rubus rosaefolius Smith), also called march bubble or milk bubble, is widely distributed and economically important in China. Raspberries are rich in nutrients such as essential amino acids, vitamin C, dietary fiber, superoxide dismutase (SOD) and minerals (Yang et al. 2019). In May 2019, a leaf spot disease was observed on raspberry in Enshi (N29°07'10', E108°23'12'), Hubei province of China. The symptoms were small dark-brown spots (Fig.1) on over 90% of observed plants. To isolate the pathogen, leaf sections (5 mm × 3 mm) from the border of the symptomatic tissue were cut and sterilized with 75% ethanol for 30 s, followed by 2% sodium hypochlorite (NaClO) for 2 min, and then rinsed three times with sterile water. Leaf sections were placed on potato dextrose agar (PDA) medium amended with 25 µg / ml ampicillin and incubated at 25 °C in the dark for 3 days. Isolated colonies were sub-cultured on PDA by hyphal tip transfer. Eight fungal isolates with similar morphology, abundant white aerial hyphae, were collected. Colonies on PDA grew up to 80 mm in diameter by 7 days at 25 °C. The center of each colony became black (Fig.2). Conidia were unicellular, oval and hyaline. Conidia ranged in size from 14.5 to 19.75 µm × 5.80 to 10.20 µm (n=50) in 20% (v/v) V8 vegetable juice medium. No appressoria were observed. Morphological characteristics are similar to those of Colletotrichum spp. (Moriwaki et al. 2003). Total genomic DNA of a representative isolate S1 was extracted with a CTAB method (Stenglein et al. 2006). Internal transcribed spacer (ITS) region of rDNA, actin (ACT) , beta-tubulin (TUB2) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) genes were amplified and sequenced with the primer pairs of ITS4 / ITS5, ACT512F / ACT783R, Bt-2a / Bt-2b and GDF1 / GDR1, respectively (Weir et al. 2012). BLAST results showed that ITS, ACT, TUB2 and GAPDH gene sequences (GenBank accession nos. MN498030, MT780498, MT780496 and MT780497, respectively) were 99% identical to those of Colletotrichum boninense Moriwaki, Sato & Tsukiboshi (GenBank accession nos. MF076598, JX009583, JQ005588 and JX009905, respectively). Concatenated sequences of the four genes were used to conduct a phylogenetic analysis using neighbor-joining method in MEGA7 (Toussaint et al. 2016). The isolate S1 clustered with above C. boninense strains retrieved from NCBI database. Therefore, the present isolate S1 was identified as C. boninense. Pathogenicity tests were performed using one-month-old raspberry plants, 24 controls and 30 inoculated. The plants were sprayed with conidial suspension ( 106 conidia / mL) cultured on 20% (v/v) V8 vegetable juice medium for 15 days. The control plants were sprayed with sterile distilled water. All plants were covered with plastic bags 24h to maintain the relative humidity in the field. Fifteen days after inoculation, typical symptoms of brown spots were observed on leaves similar to the disease on field plants, while the leaves from the control group remained asymptomatic. C. boninense was reisolated and identified from inoculated symptomatic leaves. Anthracnose on raspberry caused by Colletotrichum gloeosporioides (Dai et al. 2013) and C. fioriniae (Schoeneberg et al. 2020) has previously been reported. However, to the best of our knowledge, this is the first report of Colletotrichum boninense causing leaf spot on Raspberry in China. If more reports of this pathogen are found on raspberries, then it may be necessary to develop effective management strategies for controlling this disease.

[The Application Value of True Whole-body PET/CT Scanning Protocol in Patients of Extranodal NK/T Cell Lymphoma].

OBJECTIVE: To assess the staging, restaging, and treatment strategy determination of extranodal NK/T cell lymphoma (ENKT) by PET/CT real body (true whole-body, TWB) imaging, which is superior to PET/CT limitation of the whole body (limited whole-body, LWB, from skull vertex to upper thighs) by adding 'distal lower extremity' images. METHODS: TWB 18F-FDG PET/CT studies performed for staging and follow-up of ENKTL patients between January 2012 and September 2017 were retrospectively reviewed. Patients in staging group received TWB PET/CT evaluation for staging at the first diagnosis. In follow-up group, patients received follow-up evalution with TWB PET/CT and progressive disease (PD) in the LWB range with or without clinical diagnosis or suspicion before follow-up examination, and then divided into four subgroups: staging (+) PD (－), staging (+) PD (+), staging (－) PD (－), staging (－) PD (+). Then the percentage of unexpected ENKTL lesions found at the distal extremity (outside the LWB range) (P1), and the percentage of changes in the staging, restaging/outcome evaluation (P2) in each group were recorded. RESULTS: Among the 225 patients in the staging group, 200 (88.9%) had tumors confined to LWB, while P1 was 11.1% (25 cases) and P2 was 0.4% (1 case). In the follow-up group, the P1 in staging (+) PD (－)( n=85), staging (+) PD (+)( n=4), staging (－) PD (－)( n=43), staging (－) PD (+) goups ( n=15) were 1.2%, 75.0%, 0%, 26.7%, and P2 were 1.2%, 0%, 0%, 13.3%, respectively. In the follow-up group, regardless of whether the TWB PET/CT examination was performed at the initial diagnosis stage, P1 in PD (－) group and PD (+) group was 0.8 vs. 36.8% ( P<0.000 1), and P2 was 0.8% vs. 10.5% ( P<0.000 1). CONCLUSION: It is not recommended that the TWB PET/CT imaging from the top of the head to the bottom of the foot use for the first diagnosis of ENKTL patients. And for follow-up patients with no clinical evidence of tumor progression or with evidence of tumor progression but whose lesions were limited to LWB at the initial diagnosis of TWB PET/CT staging, LWB PET/CT from the top of the head to the middle of the thigh is recommended for routine follow-up. For ENKTL patients, TWB PET/CT was not performed at the initial stage of diagnosis to detect the condition of lower limbs. If the evidence of tumor progression in the LWB range appeared before the follow-up examination, TWB PET/CT was recommended for the follow-up evaluation to evaluate the systemic tumor involvement.

ß-arrestin2 functions as a key regulator in the sympathetic-triggered immunodepression after stroke.

BACKGROUND: Stroke-induced immunodeficiency syndrome (SIDS) is regarded as a protective mechanism for secondary inflammatory injury as well as a contributor to infection complications. Although stroke-induced hyperactivation of the sympathetic system is proved to facilitate SIDS, the involved endogenous factors and pathways are largely elusive. In this study, we aim to investigate the function of beta-arrestin-2 (ARRB2) in the sympathetic-mediated SIDS. METHODS: Splenic ARRB2 expression and the sympathetic system activity were detected after establishing transient models of middle cerebral artery occlusion (MCAO). In addition, a correlation between ARRB2 expression and the sympathetic system activity was analyzed using a linear correlation analysis. Any SIDS reflected in monocyte dysfunction was investigated by measuring inflammatory cytokine secretion and neurological deficit scores and infarct volume were tested to assess neurological outcome. Further, ARRB2 expression in the monocytes was knocked down in vitro by siRNAs. Following the stimulation of noradrenaline and lipopolysaccharide, cytokine secretion and the nuclear factor-κB (NF-κB) pathway were evaluated to gain insight into the mechanisms related to the contribution of ARRB2 to adrenergic-induced monocyte dysfunction. RESULTS: Splenic ARRB2 expression was significantly increased after stroke and also showed a significant positive correlation with the sympathetic system activity. Stroke-induced monocyte dysfunction resulted in an increase of the interleukin-10 (IL-10) level as well as a decrease of the interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) levels. Also, blockade of adrenergic-activity significantly reversed these cytokine levels, and blockade of adrenergic-activity improved stroke-induced neurological results. However, the improved neurological results had no significant correlation with ARRB2 expression. Furthermore, the in vitro results showed that the deficiency of ARRB2 dramatically repealed adrenergic-induced monocyte dysfunction and the inhibition of NF-κB signaling phosphorylation activity. CONCLUSIONS: ARRB2 is implicated in the sympathetic-triggered SIDS, in particular, monocyte dysfunction after stroke. Accordingly, ARRB2 may be a promising therapeutic target for the immunological management of stroke in a clinic.

H3K4 Methyltransferase Set1a Is A Key Oct4 Coactivator Essential for Generation of Oct4 Positive Inner Cell Mass.

Limited core transcription factors and transcriptional cofactors have been shown to govern embryonic stem cell (ESC) transcriptional circuitry and pluripotency, but the molecular interactions between the core transcription factors and cofactors remains ill defined. Here, we analyzed the protein-protein interactions between Oct4, Sox2, Klf4, and Myc (abbreviated as OSKM) and a large panel of cofactors. The data reveal both specific and common interactions between OSKM and cofactors. We found that among the SET1/MLL family H3K4 methyltransferases, Set1a specifically interacts with Oct4 and this interaction is independent of Wdr5. Set1a is recruited to and required for H3K4 methylation at the Oct4 target gene promoters and transcriptional activation of Oct4 target genes in ESCs, and consistently Set1a is required for ESC maintenance and induced pluripotent stem cell generation. Gene expression profiling and chromatin immunoprecipitation-seq analyses demonstrate the broad involvement of Set1a in Oct4 transcription circuitry and strong enrichment at TSS sites. Gene knockout study demonstrates that Set1a is not only required for mouse early embryonic development but also for the generation of Oct4-positive inner cell mass. Together our study provides valuable information on the molecular interactions between OSKM and cofactors and molecular mechanisms for the functional importance of Set1a in ESCs and early development.

Calotropin from Asclepias curasavica induces cell cycle arrest and apoptosis in cisplatin-resistant lung cancer cells.

Calotropin (M11), an active compound isolated from Asclepias curasavica L., was found to exert strong inhibitory and pro-apoptotic activity specifically against cisplatin-induced resistant non-small cell lung cancer (NSCLC) cells (A549/CDDP). Molecular mechanism study revealed that M11 induced cell cycle arrest at the G2/M phase through down-regulating cyclins, CDK1, CDK2 and up-regulating p53 and p21. Furthermore, M11 accelerated apoptosis through the mitochondrial apoptotic pathway which was accompanied by increase Bax/Bcl-2 ratio, decrease in mitochondrial membrane potential, increase in reactive oxygen species production, activations of caspases 3 and 9 as well as cleavage of poly ADP-ribose polymerase (PARP). The activation and phosphorylation of JNK was also found to be involved in M11-induced apoptosis, and SP610025 (specific JNK inhibitor) partially prevented apoptosis induced by M11. In contrast, all of the effects that M11 induce cell cycle arrest and apoptosis in A549/CDDP cells were not significant in A549 cells. Drugs with higher sensitivity against resistant tumor cells than the parent cells are rather rare. Results of this study supported the potential application of M11 on the non-small lung cancer (NSCLC) with cisplatin resistance.

Blocking Sympathetic Nervous System Reverses Partially Stroke-Induced Immunosuppression but does not Aggravate Functional Outcome After Experimental Stroke in Rats.

Stoke results in activation of the sympathetic nervous system (SNS), inducing systemic immunosuppression. However, the potential mechanisms underlying stroke-induced immunosuppression remain unclear. Here, we determined the SNS effects on functional outcome and explored the interactions among SNS, ß-arrestin2 and nuclear factor-κB (NF-κB) after experimental stroke in rats. In the current study, stroke was induced by a transient middle cerebral artery occlusion (MCAO) in rats, and SNS activity was inhibited by intraperitoneal injection of 6-hydroxydopamine HBr (6-OHDA). 7.0 T Micro-MRI and Longa score were employed to assess the functional outcome after stroke. Flow cytometry and ELISA assay were used to measure the expression of MHC class II, tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ). Western blot was conducted to analyze ß-arrestin2 and NF-κB protein expression levels after experimental stroke. We found significantly increased infarct volumes and functional impairment after MCAO at different post-surgery time points, which were not aggravated by 6-OHDA treatment. SNS blockade partially reversed the expression of MHC class II after stroke over time, as well as TNF-α and IFN-γ levels in lipopolysaccharide-stimulated macrophages in vitro. Treatment of MCAO rats with SNS-inhibitor significantly diminished NF-κB activation and enhanced ß-arrestin2 expression after stroke. This study suggests that pharmacological SNS inhibition dose not aggravate functional outcome after stroke. Stroke-induced immunosuppression may be involved in the SNS-ß-arrestin2-NF-κB pathway.

PHD finger protein 2 (PHF2) represses ribosomal RNA gene transcription by antagonizing PHF finger protein 8 (PHF8) and recruiting methyltransferase SUV39H1.

Regulation of rDNA transcription is central to cell growth and proliferation. PHF2 and PHF8 belong to a subfamily of histone demethylases that also possess a PHD domain-dependent di-/trimethylated histone 3 lysine 4 (H3K4me2/3) binding activity and are known to be enriched in the nucleolus. In this study, we show that, unlike PHF8 that activates rDNA transcription, PHF2 inhibits rDNA transcription. Depletion of PHF2 by RNA interference increases and overexpression of PHF2 decreases rDNA transcription, respectively, whereas simultaneous depletion of PHF8 and PHF2 restores the level of rDNA transcription. The inhibition of rDNA transcription by PHF2 depends on its H3K4me2/3 binding activity that is also required for PHF2 association with the promoter of rDNA genes but not its demethylase activity. We provide evidence that PHF2 is likely to repress rDNA transcription by competing with PHF8 for binding of rDNA promoter and by recruiting H3K9me2/3 methyltransferase SUV39H1. We also provide evidence that, whereas PHF8 promotes, PHF2 represses the transcriptional activity of RARα, Oct4, and KLF4 and a few PHF8 target genes tested. Taken together, our study demonstrates a repressive role for PHF2 in transcription by RNA polymerase I and II.

Emerging mechanisms and implications of cGAS-STING signaling in cancer immunotherapy strategies.

The intricate interplay between the human immune system and cancer development underscores the central role of immunotherapy in cancer treatment. Within this landscape, the innate immune system, a critical sentinel protecting against tumor incursion, is a key player. The cyclic GMP-AMP synthase (cGAS) and stimulator of interferon genes (STING) pathway has been found to be a linchpin of innate immunity: activation of this signaling pathway orchestrates the production of type I interferon (IFN-α/ß), thus fostering the maturation, differentiation, and mobilization of immune effectors in the tumor microenvironment. Furthermore, STING activation facilitates the release and presentation of tumor antigens, and therefore is an attractive target for cancer immunotherapy. Current strategies to activate the STING pathway, including use of pharmacological agonists, have made substantial advancements, particularly when combined with immune checkpoint inhibitors. These approaches have shown promise in preclinical and clinical settings, by enhancing patient survival rates. This review describes the evolving understanding of the cGAS-STING pathway's involvement in tumor biology and therapy. Moreover, this review explores classical and non-classical STING agonists, providing insights into their mechanisms of action and potential for optimizing immunotherapy strategies. Despite challenges and complexities, the cGAS-STING pathway, a promising avenue for enhancing cancer treatment efficacy, has the potential to revolutionize patient outcomes.

A Lysosome-Targeted Magnetic Nanotorquer Mechanically Triggers Ferroptosis for Breast Cancer Treatment.

Targeting ferroptosis has attracted exponential attention to eradicate cancer cells with high iron-dependent growth. Increasing the level of intracellular labile iron pool via small molecules and iron-containing nanomaterials is an effective approach to induce ferroptosis but often faces insufficient efficacy due to the fast drug metabolism and toxicity issues on normal tissues. Therefore, developing a long-acting and selective approach to regulate ferroptosis is highly demanded in cancer treatment. Herein, a lysosome-targeted magnetic nanotorquer (T7-MNT) is proposed as the mechanical tool to dynamically induce the endogenous Fe2+ pool outbreak for ferroptosis of breast cancer. T7-MNTs target lysosomes via the transferrin receptor-mediated endocytosis in breast cancer cells. Under the programmed rotating magnetic field, T7-MNTs generate torques to trigger endogenous Fe2+ release by disrupting the lysosomal membrane. This magneto-mechanical manipulation can induce oxidative damage and antioxidant defense imbalance to boost frequency- and time-dependent lipid peroxidization. Importantly, in vivo studies show that T7-MNTs can efficiently trigger ferroptosis under the magnetic field and play as a long-acting physical inducer to boost ferrotherapy efficacy in combination with RSL3. It is anticipated that this dynamic targeted strategy can be coupled with current ferroptosis inducers to achieve enhanced efficacy and inspire the design of mechanical-based ferroptosis inducers for cancer treatment.

Exosome-mediated delivery of artificial circular RNAs for gene therapy of bladder cancer.

Bladder cancer (BCa) is one of the most common malignancies affecting men. Oncogenic transcription factors function as an important regulator in the progression of human cancer. In our study, we aimed to construct artificial circular non-coding RNAs (acircRNAs) consisting of three functional units that mimic the CRISPR-Cas system and elucidate its therapeutic role in bladder cancer. Additionally, the compare of the efficiency in regulating gene expression between acircRNA and CRISPR-dCas systems was performed. We connected the cDNA sequences of TFs aptamer and constructed a circRNA. To demonstrate the platform's practicality, ß-catenin and NF-κB were chosen as functional targets, while T24 and 5637 cell lines served as test models. Real-time Quantitative PCR (qPCR), double luciferase assay and related phenotype assay were used to detect the expression of related genes and the therapeutic effect. To elucidate the functionality of acircRNAs, luciferase vectors capable of detecting ß-catenin and NF-κB expression were employed to assess the inhibitory impact of acircRNA on ß-catenin and NF-κB. Consequently, the optimal combination involving acircRNA-3 was determined. Next, qPCR assay was employed to assess the relative expression levels of target downstream genes following acircRNA treatment. The expression of c-myc and cyclin D1 were used to determine the function of ß-catenin, while Bcl-XL and TRAF1 were used to determine that of NF-κB. The acircRNAs inhibited the ß-catenin and NF-κB related signaling in BCa cells specifically. CD63-HuR fusion protein was used to loading acircRNA into exosomes. The results showed that acircRNA could inhibit the activity of the target transcription factors, and the inhibitory effect was better than that of CRIPSR-dCas9-KRAB. Furthermore, functional experiments demonstrated that the transfection of acircRNA in bladder cells resulted in decreased proliferation, enhanced apoptosis, and suppressed migration. In conclusion, our synthetic gene device exhibited anti-tumor regulatory capabilities and showed greater efficiency in tumor suppression compared to the CRISPR-dCas9-KRAB system. Therefore, our device provides a new strategy for cancer treatment and could be a useful strategy for cancer cells.

CL429 enhances the renewal of intestinal stem cells by upregulating TLR2-YAP1.

Intestinal stem cells (ISCs) play a crucial role in maintaining the equilibrium and regenerative potential of intestinal tissue, thereby ensuring tissue homeostasis and promoting effective tissue regeneration following injury. It has been proven that targeting Toll-like receptors (TLRs) can help prevent radiation-induced damage to the intestine. In this study, we established an intestinal injury model using IR and evaluated the effects of CL429 on ISC regeneration both in vivo and in vitro. Following radiation exposure, mice treated with CL429 showed a significant increase in survival rates (100% survival in the treated group compared to 54.54% in the control group). CL429 also showed remarkable efficacy in inhibiting radiation-induced intestinal damage and promoting ISC proliferation and regeneration. In addition, CL429 protected intestinal organoids against IR-induced injury. Mechanistically, RNA sequencing and Western blot analysis revealed the activation of the Wnt and Hippo signaling pathways by CL429. Specifically, we observed a significant upregulation of YAP1, a key transcription factor in the Hippo pathway, upon CL429 stimulation. Furthermore, knockdown of YAP1 significantly attenuated the radioprotective effect of CL429 on intestinal organoids, indicating that CL429-mediated intestinal radioprotection is dependent on YAP1. In addition, we investigated the relationship between TLR2 and YAP1 using TLR2 knockout mice, and our results showed that TLR2 knockout abolished the activation of CL429 on YAP1. Taken together, our study provides evidence supporting the role of CL429 in promoting ISC regeneration through activation of TLR2-YAP1. And further investigation of the interaction between TLRs and other signaling pathways may enhance our understanding of ISC regeneration after injury.

Scleroglucan protects the intestine from irradiation-induced injury by targeting the IL-17 signaling pathway.

BACKGROUND: Intestinal tissue is extremely sensitive to ionizing radiation (IR), which is easy to cause intestinal radiation sickness, and the mortality rate is very high after exposure. Recent studies have found that intestinal immune cells and intestinal stem cells (ISCs) may play a key role in IR-induced intestinal injury. METHODS: C57BL6 mice matched for age, sex and weight were randomly grouped and intraperitoneal injected with PBS, Scleroglucan (125.0 mg/kg) or Anti-mouse IL-17A -InVivo (10 mg/kg), the number of mice in each group was n ≥ 3.Survival time, body weight, pathology, organoids and immune cell markers of the mice after IR (10.0 Gy) were compared, and the mechanism of action in intestinal tissues was verified by transcriptome sequencing. RESULTS: Scleroglucan has significant radiation protective effects on the intestine, including improving the survival rate of irradiated mice, inhibiting the radiation damage of intestinal tissue, and promoting the proliferation and differentiation of intestinal stem cells (ISCs). The results of RNA sequencing suggested that Scleroglucan could significantly activate the immune system and up-regulate the IL-17 and NF-κB signaling pathways. Flow cytometry showed that Scleroglucan could significantly up-regulate the number of Th17 cells and the level of IL-17A in the gut. IL-17A provides radiation protection. After intraperitoneal injection of Scleroglucan and Anti-mouse IL-17A -InVivo, mice can significantly reverse the radiation protection effect of Scleroglucan, down-regulate the molecular markers of intestinal stem cells and the associated markers of DC, Th1 and Th17 cells, and up-regulate the associated markers of Treg and Macrophage cells. CONCLUSION: Scleroglucan may promote the proliferation and regeneration of ISCs by regulating the activation of intestinal immune function mediated by IL-17 signaling pathway and play a protective role in IR-induced injury.