A Mettl16/m6A/mybl2b/Igf2bp1 axis ensures cell cycle progression of embryonic hematopoietic stem and progenitor cells.

Prenatal lethality associated with mouse knockout of Mettl16, a recently identified RNA N6-methyladenosine (m6A) methyltransferase, has hampered characterization of the essential role of METTL16-mediated RNA m6A modification in early embryonic development. Here, using cross-species single-cell RNA sequencing analysis, we found that during early embryonic development, METTL16 is more highly expressed in vertebrate hematopoietic stem and progenitor cells (HSPCs) than other methyltransferases. In Mettl16-deficient zebrafish, proliferation capacity of embryonic HSPCs is compromised due to G1/S cell cycle arrest, an effect whose rescue requires Mettl16 with intact methyltransferase activity. We further identify the cell-cycle transcription factor mybl2b as a directly regulated by Mettl16-mediated m6A modification. Mettl16 deficiency resulted in the destabilization of mybl2b mRNA, likely due to lost binding by the m6A reader Igf2bp1 in vivo. Moreover, we found that the METTL16-m6A-MYBL2-IGF2BP1 axis controlling G1/S progression is conserved in humans. Collectively, our findings elucidate the critical function of METTL16-mediated m6A modification in HSPC cell cycle progression during early embryonic development.

Deubiquitinase USP9X stabilizes RNA m6A demethylase ALKBH5 and promotes acute myeloid leukemia cell survival.

Post-translational modifications including protein ubiquitination regulate a plethora of cellular processes in distinct manners. RNA N6-methyladenosine is the most abundant post-transcriptional modification on mammalian mRNAs and plays important roles in various physiological and pathological conditions including hematologic malignancies. We previously determined that the RNA N6-methyladenosine eraser ALKBH5 is necessary for the maintenance of acute myeloid leukemia (AML) stem cell function, but the post-translational modifications involved in ALKBH5 regulation remain elusive. Here, we show that deubiquitinase ubiquitin-specific peptidase 9X (USP9X) stabilizes ALKBH5 and promotes AML cell survival. Through the use of mass spectrometry as an unbiased approach, we identify USP9X and confirm that it directly binds to ALKBH5. USP9X stabilizes ALKBH5 by removing the K48-linked polyubiquitin chain at K57. Using human myeloid leukemia cells and a murine AML model, we find that genetic knockdown or pharmaceutical inhibition of USP9X inhibits leukemia cell proliferation, induces apoptosis, and delays AML development. Ectopic expression of ALKBH5 partially mediates the function of USP9X in AML. Overall, this study uncovers deubiquitinase USP9X as a key for stabilizing ALKBH5 expression and reveals the important role of USP9X in AML, which provides a promising therapeutic strategy for AML treatment in the clinic.

Cation Modifies Interfacial Water Structures on Platinum during Alkaline Hydrogen Electrocatalysis.

The molecular details of an electrocatalytic interface play an essential role in the production of sustainable fuels and value-added chemicals. Many electrochemical reactions exhibit strong cation-dependent activities, but how cations affect reaction kinetics is still elusive. We report the effect of cations (K+, Li+, and Ba2+) on the interfacial water structure using second-harmonic generation (SHG) and classical molecular dynamics (MD) simulation. The second- (χH2O(2)) and third-order (χH2O(3)) optical susceptibilities of water on Pt are smaller in the presence of Ba2+ compared to those of K+, suggesting that cations can affect the interfacial water orientation. MD simulation reproduces experimental SHG observations and further shows that the competition between cation hydration and interfacial water alignment governs the net water orientation. The impact of cations on interfacial water supports a cation hydration-mediated mechanism for hydrogen electrocatalysis; i.e., the reaction occurs via water dissociation followed by cation-assisted hydroxide/water exchange on Pt. Our study highlights the role of interfacial water in electrocatalysis and how innocent additives (such as cations) can affect the local electrochemical environment.

Association of estimated glomerular filtration rate with prostate cancer risk in a cross-ethnic population: a Mendelian randomization study.

OBJECTIVE: To investigate whether a causal relationship exists between the estimated glomerular filtration rate (EGFR) and the occurrence of prostate cancer in East Asian and European populations and to determine if genetic factors influence the association between the EGFR and prostate cancer risk. METHODS: In this Mendelian randomization study, the existence of a causal relationship between the EGFR and prostate cancer occurrence was assessed using five analytical techniques, including Mendelian randomization-Egger regression (MR-Egger), calculation of the weighted median estimator (WME), the maximum likelihood ratio method, the linear median weighting method and the random-effects inverse-variance weighting (IVW) method. RESULTS: In the IVW model, no causal relationship was observed between the EGFR and prostate cancer in either the East Asian or European populations. CONCLUSIONS: After excluding confounding factors and reverse causal associations using two-sample Mendelian randomization, unbiased estimates were obtained, and there was no causal relationship between prostate cancer and the EGFR in the East Asian or European populations. Therefore, for patients with suspected prostate cancer, it is considered unnecessary to improve the detection of glomerular filtration rate, which will effectively reduce the economic burden of patients.

Biotransformation of antioxidant eriocitrin into characteristic metabolites by the gut microbiota.

Eriocitrin is a flavonoid glycoside with strong antioxidant capacity that has a variety of pharmacological activities, such as hypolipidemic, anticancer and anti-inflammatory effects. We found that the gut microbiota could rapidly metabolize eriocitrin. By using LC/MSn-IT-TOF, we identified three metabolites of eriocitrin metabolized in the intestinal microbiota: eriodictyol-7-O-glucoside, eriodictyol, and dihydrocaffeic acid. By comparing these two metabolic pathways of eriocitrin (the gut microbiota and liver microsomes), the intestinal microbiota may be the primary metabolic site of eriocitrin metabolism. These findings provide a theoretical foundation for the study of pharmacologically active substances.

The changes of intestinal microbiota and metabolomics during the inhibition of bladder cancer by liquiritigenin.

Liquiritigenin is a natural medicine. However, its inhibitory effect and its potential mechanism on bladder cancer (BCa) remain to be explored. It was found that it could be visualized that the transplanted tumours in the low-dose liquiritigenin -treated group and the high-dose liquiritigenin -treated group were smaller than those in the model group. Liquiritigenin treatment led to alterations in Lachnoclostridium, Escherichia-Shigella, Alistipes and Akkermansia. Non-targeted metabolomics analysis showed that a total of multiple differential metabolites were identified between the model group and the high-dose liquiritigenin-treated group. This provides a new direction and rationale for the antitumour effects of liquiritigenin.

YBX1 is required for maintaining myeloid leukemia cell survival by regulating BCL2 stability in an m6A-dependent manner.

RNA-binding proteins (RBPs) are critical regulators of transcription and translation that are often dysregulated in cancer. Although RBPs are increasingly recognized as being important for normal hematopoiesis and for hematologic malignancies as oncogenes or tumor suppressors, RBPs that are essential for the maintenance and survival of leukemia remain elusive. Here we show that YBX1 is specifically required for maintaining myeloid leukemia cell survival in an N6-methyladenosine (m6A)-dependent manner. We found that expression of YBX1 is significantly upregulated in myeloid leukemia cells, and deletion of YBX1 dramatically induces apoptosis and promotes differentiation coupled with reduced proliferation and impaired leukemic capacity of primary human and mouse acute myeloid leukemia cells in vitro and in vivo. Loss of YBX1 has no obvious effect on normal hematopoiesis. Mechanistically, YBX1 interacts with insulin-like growth factor 2 messenger RNA (mRNA)-binding proteins (IGF2BPs) and stabilizes m6A-tagged RNA. Moreover, YBX1 deficiency dysregulates the expression of apoptosis-related genes and promotes mRNA decay of MYC and BCL2 in an m6A-dependent manner, which contributes to the defective survival that results from deletion of YBX1. Thus, our findings have uncovered a selective and critical role of YBX1 in maintaining myeloid leukemia survival, which might provide a rationale for the therapeutic targeting of YBX1 in myeloid leukemia.

Pax transactivation domain-interacting protein is required for preserving hematopoietic stem cell quiescence via regulating lysosomal activity.

Hematopoietic stem cells (HSC) maintain lifetime whole blood hematopoiesis through self-renewal and differentiation. In order to sustain HSC stemness, most HSC reside in a quiescence state, which is affected by diverse cellular stress and intracellular signal transduction. How HSC accommodate those challenges to preserve lifetime capacity remains elusive. Here we show that Pax transactivation domain-interacting protein (PTIP) is required for preserving HSC quiescence via regulating lysosomal activity. Using a genetic knockout mouse model to specifically delete Ptip in HSC, we find that loss of Ptip promotes HSC exiting quiescence, and results in functional exhaustion of HSC. Mechanistically, Ptip loss increases lysosomal degradative activity of HSC. Restraining lysosomal activity restores the quiescence and repopulation potency of Ptip-/- HSC. Additionally, PTIP interacts with SMAD2/3 and mediates transforming growth factor-ß signaling-induced HSC quiescence. Overall, our work uncovers a key role of PTIP in sustaining HSC quiescence via regulating lysosomal activity.

RNA Modifications in Hematologic Malignancies.

Chemical modifications on macromolecules such as DNA, RNA and proteins play important roles in almost all biological processes. The revival of RNA modification research began with the discovery of RNA modification machineries, and with the development of better techniques for characterizing and profiling these modifications at the transcriptome-wide level. Hematopoietic system is maintained by hematopoietic stem cells that possess efficient self-renewal capacity and the potential of differentiation into all lineages of blood cells, and the imbalance of this homeostasis frequently causes hematologic malignancies such as leukemia. Recent studies reveal that dysregulated RNA modifications play essential roles in hematologic malignancies. Herein, we summarize recent advances in some major RNA modifications, the detection methods, roles and mechanisms of these RNA modifications in hematologic malignancies.

Metabolites Analysis of Anti-Myocardial Ischemia Active Components of Saussurea involucrata Based on Gut Microbiota-Drug Interaction.

Saussurea involucrata has been reported to have potential therapeutic effects against myocardial ischemia. The pharmacological effects of oral natural medicines may be influenced by the participation of gut microbiota. In this study, we aimed to investigate the bidirectional regulation of gut microbiota and the main components of Saussurea involucrata. We first established a quantitative method for the four main components (chlorogenic acid, syringin, acanthoside B, rutin) which were chosen by fingerprint using liquid chromatography tandem mass spectrometry (LC-MS/MS), and found that gut microbiota has a strong metabolic effect on them. Meanwhile, we identified five major rat gut microbiota metabolites (M1-M5) using liquid chromatography tandem time-of-flight mass spectrometry (LC/MSn-IT-TOF). The metabolic properties of metabolites in vitro were preliminarily elucidated by LC-MS/MS for the first time. These five metabolites of Saussurea involucrata may all have potential contributions to the treatment of myocardial ischemia. Furthermore, the four main components (10 µg/mL) can significantly stimulate intestinal bacteria to produce short chain fatty acids in vitro, respectively, which can further contribute to the effect in myocardial ischemia. In this study, the therapeutic effect against myocardial ischemia of Saussurea involucrata was first reported to be related to the intestinal flora, which can be useful in understanding the effective substances of Saussurea involucrata.

Biotransformation of Liquiritigenin into Characteristic Metabolites by the Gut Microbiota.

The bioavailability of flavonoids is generally low after oral administration. The metabolic transformation of flavonoids by the gut microbiota may be one of the main reasons for this, although these metabolites have potential pharmacological activities. Liquiritigenin is an important dihydroflavonoid compound found in Glycyrrhiza uralensis that has a wide range of pharmacological properties, such as antitumor, antiulcer, anti-inflammatory, and anti-AIDS effects, but its mechanism of action remains unclear. This study explored the metabolites of liquiritigenin by examining gut microbiota metabolism and hepatic metabolism in vitro. Using LC-MS/MS and LC/MSn-IT-TOF techniques, three possible metabolites of liquiritigenin metabolized by the gut microbiota were identified: phloretic acid (M3), resorcinol (M4), and M5. M5 is speculated to be davidigenin, which has antitumor activity. By comparing these two metabolic pathways of liquiritigenin (the gut microbiota and liver microsomes), this study revealed that there are three main metabolites of liquiritigenin generated by intestinal bacteria, which provides a theoretical basis for the study of pharmacologically active substances in vivo.

PESV represses non-small cell lung cancer cell malignancy through circ_0016760 under hypoxia.

BACKGROUND: Non-small cell lung cancer (NSCLC) accounts for more than 80% of lung cancers, which is the most common malignant tumor worldwide. Polypeptide extract from scorpion venom (PESV) has been reported to inhibit NSCLC process. The present study aims to reveal the roles of PESV in NSCLC progression under hypoxia and the inner mechanism. METHODS: The expression levels of circular RNA 0016760 (circ_0016760) and microRNA-29b (miR-29b) were detected by quantitative real-time polymerase chain reaction (qRT-PCR). Protein expression was determined by western blot and immunohistochemistry assays. Cell migration, invasion, proliferation and tube formation were investigated by transwell, cell colony formation, 3-(4,5-Dimethylthazol-2-yl)-2,5-diphenyltetrazolium bromide and tube formation assays. The impacts between PESV and circ_0016760 overexpression on tumor growth in vivo were investigated by in vivo tumor formation assay. RESULTS: Circ_0016760 expression was dramatically upregulated in NSCLC tissues and cells, compared with adjacent lung tissues and cells, respectively. PESV treatment downregulated circ_0016760 expression. Circ_0016760 silencing or PESV treatment repressed cell migration, invasion, proliferation and tube formation under hypoxia in NSCLC cells. Circ_0016760 overexpression restored the effects of PESV treatment on NSCLC process under hypoxia. Additionally, circ_0016760 acted as a sponge of miR-29b, and miR-29b bound to HIF1A. Meanwhile, miR-29b inhibitor impaired the influences of circ_0016760 knockdown on NSCLC process under hypoxia. Further, ectopic circ_0016760 expression restrained the effects of PESV exposure on tumor formation in vivo. CONCLUSION: Circ_0016760 overexpression counteracted PESV-induced repression of NSCLC cell malignancy and angiogenesis under hypoxia through miR-29b/HIF1A axis.

Transcription factor OsMADS25 improves rice tolerance to cold stress.

Cold stress is the limiting factor of rice growth and production, and it is important to clone cold stress tolerant genes and cultivate cold tolerance rice varieties. The MADS transcription factors play an important role in abiotic stress signaling in rice. This study showed that OsMADS25 was up-regulated by low temperature and abscisic acid (ABA), suggesting that OsMADS25 may be involved in ABA-dependent signaling. The OsMADS25 overexpression vector, pCambia1300-221-OsMADS25-Flag, was constructed and introduced into the rice variety Zhonghua 11 (ZH11) through Agrobacterium tumefacian-mediated genetic transformation. Two homozygous lines with high expression levels were selected for phenotypic identification. OsMADS25 overexpression lines show significantly improved cold stress tolerance and the sensitivity to ABA at the seedling stage of rice. Reactive oxygen species (ROS) was detected by diaminobenzidine (DAB) staining and nitroblue tetrazolium (NBT) staining. After treatment with cold stress, little ROS accumulation was observed in OsMADS25 overexpression lines compared to wild-type ZH11. In conclusion, OsMADS25 plays a role in scavenging reactive oxygen species (ROS) and could improve rice tolerance to cold stress involved in ABA-dependent pathway.

Pyrimidine Nucleosides from Streptomyces sp. SSA28.

Eleven new pyrimidine nucleosides (1-11) and 12 known analogues (12-23) were isolated from the marine-derived Streptomyces sp. SSA28. All of the new structures were elucidated by extensive NMR spectroscopic analysis and HRESIMS data. The absolute configurations of compound 1 were determined by X-ray diffraction. The configurations of 2-16 were investigated by ECD calculations. Compounds 11-16 showed cytotoxicity against HCT-116 human colon cancer cell lines with IC50 values from 0.39 ± 0.03 to 6.63 ± 0.47 µM.

Bioactive staurosporine derivatives from the Streptomyces sp. NB-A13.

Six new (1-6) and nine known (7-15) staurosporine derivatives were isolated from the rice solid fermentation of the marine-derived Streptomyces sp. NB-A13. The structures of the new staurosporine derivatives were established by extensive spectroscopic data interpretation. The absolute configurations of 1 and 2 were assigned by quantum chemical calculations of the electronic circular dichroism (ECD) spectra. All of these compounds were screened for their cytotoxic activities against PC-3 and SW-620 cell lines. Compound 7 exhibited stronger inhibitory activity against SW-620 cell lines than the positive control staurosporine (25.10 nM), with IC50 values of 9.99 nM. Moreover, compounds 1-5, 8-13 and 15 also showed significant cytotoxicities with IC50 values ranging from 0.02 to 16.60 µM, while 6 exhibited no cytotoxic potency. Additionally, compounds 1-7 were also tested for enzyme inhibition activities of Protein kinase C theta (PKC-θ), and showed activity with IC50 values ranging from 0.06 to 9.43 µM except for compound 6, which has no inhibition activity.

Cellular and Molecular State of Myeloid Leukemia Stem Cells.

Leukemia stem cells (LSCs) are leukemia-initiating population with the capacity to self-renew, differentiate, and stay quiescent. Human hematopoietic malignancies such as chronic myeloid leukemia (CML) and acute myeloid leukemia (AML) are derived from this cell population. LSCs are also responsible for disease relapse due to its resistance to drug treatment. This rare cell population is phenotypically and functionally heterogeneous. Increasing evidence indicates that this heterogeneous cellular state of LSCs might determine the different drug sensitivity and is the major reason for disease relapse. In here, focusing on myeloid leukemia stem cells, we describe the biological features including cellular and molecular state, heterogeneity of LSCs, and the dynamic cross talk between LSCs and bone marrow microenvironment. These specific features of LSCs highlight the dynamic cellular state of LSCs, and further exploring on it might provide potential therapeutic targets that are important for eliminating LSCs.

Bioactive Indolocarbazoles from the Marine-Derived Streptomyces sp. DT-A61.

Nine new indolocarbazoles (1-9) were isolated from the marine-derived Streptomyces sp. DT-A61. Among them compounds 1-8 featured a hydroxy group at the C-3 or C-9 position. All purified compounds were identified by 1D and 2D NMR and HRESIMS data. The absolute configurations of 4-6, 8, and 9 were determined by electronic circular dichroism spectroscopic data. Compound 7 exhibited significant activity against human prostate PC-3 cancer cells with an IC50 value of 0.16 µM. Compounds 1, 5, 6, and 9 showed moderate inhibition against the same cell line with IC50 values of 8.0, 3.6, 3.1, and 5.6 µM. Compound 2 displayed a notable inhibitory effect against Rho-associated protein kinase (ROCK2) with an IC50 value of 5.7 nM, which was similar to the positive control staurosporine (IC50 7.8 nM).

Cyclizidine-Type Alkaloids from Streptomyces sp. HNA39.

Eight new cyclizidine-type alkaloids (1-8) and one known alkaloid (9) were identified from the chemical investigations of a marine-derived actinomycete, Streptomyces sp. HNA39. Among these alkaloids, compounds 3, 7, and 8 contain a chlorine atom, and the known alkaloid, (+)-ent-cyclizidine (9), is now first reported as a natural product. Their structures were elucidated by extensive NMR-spectroscopic analysis and HRESIMS data. The absolute configurations of all of the compounds were established by ECD calculations. Cytotoxicity evaluations of all of the compounds showed that compound 2 exhibited significant activity against the PC3 and HCT116 human-cancer-cell lines with IC50 values of 0.52 ± 0.03 and 8.3 ± 0.1 µM, respectively. Interestingly, compounds 2, 5, 7, and 8 exhibited moderate inhibition against the ROCK2 protein kinase with IC50 values from 7.0 ± 0.8 to 42 ± 3 µM.

Medermycin-Type Naphthoquinones from the Marine-Derived Streptomyces sp. XMA39.

Four new medermycin-type naphthoquinones, strepoxepinmycins A-D (1-4), and one known compound, medermycin (5), were identified from Streptomyces sp. XMA39. Their structures were elucidated by analysis of HRESIMS, 1D and 2D NMR spectroscopic data, and ECD calculations. Among these compounds, strepoxepinmycin A (1) represents a rare 5,10-oxepindione ring system typically formed by a Baeyer-Villiger oxidation, and strepoxepinmycin B (2) is an isolation artifact derived from 1. Bioactivity evaluations of these compounds showed that compounds 3 and 4 exhibited cytotoxicity against HCT-116 and PC-3 cancer cell lines and 4 exhibited moderate inhibition of ROCK 2 protein kinase. In addition, all of the new compounds showed antibacterial activity against Escherichia coli and methicillin-resistant Staphylococcus aureus and antifungal activity against Candida albicans.

GABP transcription factor is required for development of chronic myelogenous leukemia via its control of PRKD2.

Hematopoietic stem cells (HSCs) are the source of all blood lineages, and HSCs must balance quiescence, self-renewal, and differentiation to meet lifelong needs for blood cell development. Transformation of HSCs by the breakpoint cluster region-ABL tyrosine kinase (BCR-ABL) oncogene causes chronic myelogenous leukemia (CML). The E-twenty six (ets) transcription factor GA binding protein (GABP) is a tetrameric transcription factor complex that contains GABPα and GABPß proteins. Deletion in bone marrow of Gabpa, the gene that encodes the DNA-binding component, caused cell cycle arrest in HSCs and profound loss of hematopoietic progenitor cells. Loss of Gabpα prevented development of CML, although mice continued to generate BCR-ABL-expressing Gabpα-null cells for months that were serially transplantable and contributed to all lineages in secondary recipients. A bioinformatic screen identified the serine-threonine kinase protein kinase D2 (PRKD2) as a potential effector of GABP in HSCs. Prkd2 expression was markedly reduced in Gabpα-null HSCs and progenitor cells. Reduced expression of PRKD2 or pharmacologic inhibition decreased cell cycling, and PRKD2 rescued growth of Gabpα-null BCR-ABL-expressing cells. Thus, GABP is required for HSC cell cycle entry and CML development through its control of PRKD2. This offers a potential therapeutic target in leukemia.