Mitochondrial tRNA pseudouridylation governs erythropoiesis.

Pseudouridine is the most prevalent RNA modification, and its aberrant function is implicated in various human diseases. However, the specific impact of pseudouridylation on hematopoiesis remains poorly understood. In this study, we investigated the role of tRNA pseudouridylation in erythropoiesis and its association with mitochondrial myopathy, lactic acidosis, and sideroblastic anemia syndrome (MLASA) pathogenesis. By utilizing patient-specific induced pluripotent stem cells (iPSCs) carrying a genetic PUS1 mutation and a corresponding mutant mouse model, we demonstrated impaired erythropoiesis in MLASA iPSCs and anemia in the MLASA mouse model. Both MLASA iPSCs and mouse erythroblasts exhibited compromised mitochondrial function and impaired protein synthesis. Mechanistically, we revealed that PUS1 deficiency resulted in reduced mitochondrial tRNA levels due to pseudouridylation loss, leading to aberrant mitochondrial translation. Screening of mitochondrial supplements aimed at enhancing respiration or heme synthesis showed limited effect in promoting erythroid differentiation. Interestingly, the mTOR inhibitor rapamycin facilitated erythroid differentiation in MLASA-iPSCs by suppressing mTOR signaling and protein synthesis, and consistent results were observed in the MLASA mouse model. Importantly, rapamycin treatment effectively ameliorated anemia phenotypes in the MLASA patient. Our findings provide novel insights into the crucial role of mitochondrial tRNA pseudouridylation in governing erythropoiesis and present potential therapeutic strategies for anemia patients facing challenges related to protein translation.

Distinguish Esophageal Cancer Cells through VOCs Induced by Methionine Regulation.

Detection of exhaled volatile organic compounds (VOCs) is promising for noninvasive screening of esophageal cancer (EC). Cellular VOC analysis can be used to investigate potential biomarkers. Considering the crucial role of methionine (Met) during cancer development, exploring associated abnormal metabolic phenotypes becomes imperative. In this work, we employed headspace solid-phase microextraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS) to investigate the volatile metabolic profiles of EC cells (KYSE150) and normal esophageal epithelial cells (HEECs) under a Met regulation strategy. Using untargeted approaches, we analyzed the metabolic VOCs of the two cell types and explored the differential VOCs between them. Subsequently, we utilized targeted approaches to analyze the differential VOCs in both cell types under gradient Met culture conditions. The results revealed that there were five/six differential VOCs between cells under Met-containing/Met-free culture conditions. And the difference in levels of two characteristic VOCs (1-butanol and ethyl 2-methylbutyrate) between the two cell types intensified with the increase of the Met concentration. Notably, this is the first report on VOC analysis of EC cells and the first to consider the effect of Met on volatile metabolic profiles. The present work indicates that EC cells can be distinguished through VOCs induced by Met regulation, which holds promise for providing novel insights into diagnostic strategies.

HS-SPME-GC-MS Untargeted Analysis of Normal Rat Organs Ex Vivo: Differential VOC Discrimination and Fingerprint VOC Identification.

The investigation of volatile organic compounds (VOCs) in human metabolites has been a topic of interest as it holds the potential for the development of non-invasive technologies to screen for organ lesions in vivo. However, it remains unclear whether VOCs differ among healthy organs. Consequently, a study was conducted to analyze VOCs in ex vivo organ tissues obtained from 16 Wistar rats, comprising 12 different organs. The VOCs released from each organ tissue were detected by the headspace-solid phase microextraction-gas chromatography-mass spectrometry technique. In the untargeted analysis of 147 chromatographic peaks, the differential volatiles of rat organs were explored based on the Mann-Whitney U test and fold change (FC > 2.0) compared with other organs. It was found that there were differential VOCs in seven organs. A discussion on the possible metabolic pathways and related biomarkers of organ differential VOCs was conducted. Based on the orthogonal partial least squares discriminant analysis and receiver operating characteristic curve, we found that differential VOCs in the liver, cecum, spleen, and kidney can be used as the unique identification of the corresponding organ. In this study, differential VOCs of organs in rats were systematically reported for the first time. Profiles of VOCs produced by healthy organs can serve as a reference or baseline that may indicate the presence of disease or abnormalities in the organ's function. Differential VOCs can be used as the fingerprint of organs, and future integration with metabolic research may contribute to the development of healthcare.

Mass cytometry analysis identifies T cell immune signature of aplastic anemia and predicts the response to cyclosporine.

Aplastic anemia (AA) is an auto-activated T cell-mediated bone marrow failure. Cyclosporine is often used to treat non-severe AA, which demonstrates a more heterogeneous condition than severe AA. The response rate to cyclosporine is only around 50% in non-severe AA. To better predict response to cyclosporine and pinpoint who is the appropriate candidate for cyclosporine, we performed phenotypic and functional T cell immune signature at single cell level by mass cytometry from 30 patients with non-severe AA. Unexpectedly, non-significant differences of T cell subsets were observed between AA and healthy control or cyclosporine-responder and non-responders. Interestingly, when screening the expression of co-inhibitory molecules, T cell trafficking mediators, and cytokines, we found an increase of cytotoxic T lymphocyte antigen 4 (CTLA-4) on T cells in response to cyclosporine and a lower level of CTLA-4 on CD8+ T cells was correlated to hematologic response. Moreover, a decreased expression of sphingosine-1-phosphate receptor 1 (S1P1) on naive T cells and a lower level of interleukin-9 (IL-9) on T helpers also predicted a better response to cyclosporine, respectively. Therefore, the T cell immune signature, especially in CTAL-4, S1P1, and IL-9, has a predictive value for response to cyclosporine. Collectively, our study implies that immune signature analysis of T cell by mass cytometry is a useful tool to make a strategic decision on cyclosporine treatment of AA.

Osteopontin is required for the maintenance of leukemia stem cells in acute myeloid leukemia.

Acute myeloid leukemia (AML) is a heterogeneous hematopoietic disorder with a poor prognosis. The clinical significance of Leukemia stem cells (LSCs) plays an important role in the generation of AML and is the main cause of the recurrence after remission. Osteopontin (OPN), an extracellular matrix protein, has been implicated in hematopoietic malignancies. However, the specific role and the underlying mechanism of AML cell autocrined OPN in leukemia maintenance remain unknown. Here, we showed that knockdown of Opn expression significantly prolonged the survival of mice with MLL-AF9 cell-induced AML and markedly reduced the tumor burden. The LSCs from the Opn-knockdown groups exhibited decreased numbers and impaired function as determined by immunophenotype, colony-forming and limiting dilution assays. Further analysis revealed that Opn prevents LSCs from undergoing apoptosis and cell cycle arrest. Repression of OPN in human AML cell lines in vitro mimics the phenotypes observed in the mouse model. Overall, our data indicated that OPN is a potent therapeutic target for eradicating LSCs in AML.

Distinguish oral-source VOCs and control their potential impact on breath biomarkers.

By means of glass bottle sampling followed by solid-phase microextraction gas chromatography-mass spectrometry (SPME-GC-MS) technique, the change characteristics of volatile organic compounds (VOCs) in breaths, between before gargling and after gargling, were investigated, respectively, in 41 healthy subjects and 50 esophageal cancer patients. Using an untargeted strategy, 143 VOC chromatographic peaks were enrolled in the statistical analysis. Based on the orthogonal partial least squares discriminant analysis (OPLS-DA), the VOC variations after gargling for each breath test group were obtained according to the combined criteria of variable importance in projection (VIP > 1.5), Wilcoxon signed-rank test (P < 0.05), and fold change (FC > 2.0). When gargled, the levels of indole, phenol, 1-propanol, and p-cresol in the breath of healthy people decreased; meanwhile, for esophageal cancer patients, the declined VOCs in breath were indole, phenol, dimethyl disulfide, and p-cresol. Particularly, these substances were previously reported as breath biomarkers in some diseases such as esophageal, gastric, thyroid, breast, oral, and lung cancers, as well as certain non-cancer disorders. The present work indicates that expiratory VOCs involve the prominent oral cavity source, and in the breath biomarkers study, the potential impact that originates from oral volatiles should be considered. In view of the present results, it is also proposed that gargle pretreatment could eliminate possible interference from the oral cavity VOCs that might benefit breath biomarker investigation. Gargle pretreatment helps to distinguish oral-source VOCs and control their potential impact on breath biomarkers.

Impact and outcomes of postoperative anaemia in colorectal cancer patients: a systematic review.

AIM: Preoperative anaemia is common in colorectal cancer patients. Little attention has been given to the prevalence and consequences of postoperative anaemia. The aim of this study was to systematically review the published literature and determine the knowledge of the prevalence and impact of postoperative anaemia in colorectal cancer patients. METHODS: The databases Cumulative Index of Nursing and Allied Health Literature (CINAHL) and Medline, via EBSCOhost, were systematically searched to identify suitable articles published between 2004 and 2020. After an initial search, articles were screened and all eligible articles reporting on the prevalence of postoperative anaemia and clinical and long-term outcome data in colorectal cancer patients undergoing surgery were included. The Risk of Bias 2.0 tool for the assessment of randomized controlled trials and the Risk of Bias 1.0 tool for non-randomized studies were used for the assessment of bias in the studies selected in our review. RESULTS: Six studies, one randomized control trial and five cohort studies, were included with a total population size of 1714. The prevalence of anaemia at discharge of 76.6% was reported as the primary end-point in only one study. The rate of red blood cell transfusion and length of hospital stay were found to be significantly increased in anaemic patients, while postoperative infection rate results were variable. Quality of life scores and overall survival at 5 years were significantly affected among anaemic patients as reported in two papers. CONCLUSION: The available limited evidence on postoperative anaemia indicates its high prevalence with negative impact on clinical and long-term outcomes. Further research is required to standardize the measurement and address the true impact of correcting postoperative anaemia on functional and oncological outcomes.

Consequences of Lipid Remodeling of Adipocyte Membranes Being Functionally Distinct from Lipid Storage in Obesity.

Obesity is a complex disorder where the genome interacts with diet and environmental factors to ultimately influence body mass, composition, and shape. Numerous studies have investigated how bulk lipid metabolism of adipose tissue changes with obesity and, in particular, how the composition of triglycerides (TGs) changes with increased adipocyte expansion. However, reflecting the analytical challenge posed by examining non-TG lipids in extracts dominated by TGs, the glycerophospholipid composition of cell membranes has been seldom investigated. Phospholipids (PLs) contribute to a variety of cellular processes including maintaining organelle functionality, providing an optimized environment for membrane-associated proteins, and acting as pools for metabolites (e.g. choline for one-carbon metabolism and for methylation of DNA). We have conducted a comprehensive lipidomic study of white adipose tissue in mice which become obese either through genetic modification (ob/ob), diet (high fat diet), or a combination of the two, using both solid phase extraction and ion mobility to increase coverage of the lipidome. Composition changes in seven classes of lipids (free fatty acids, diglycerides, TGs, phosphatidylcholines, lyso-phosphatidylcholines, phosphatidylethanolamines, and phosphatidylserines) correlated with perturbations in one-carbon metabolism and transcriptional changes in adipose tissue. We demonstrate that changes in TGs that dominate the overall lipid composition of white adipose tissue are distinct from diet-induced alterations of PLs, the predominant components of the cell membranes. PLs correlate better with transcriptional and one-carbon metabolism changes within the cell, suggesting that the compositional changes that occur in cell membranes during adipocyte expansion have far-reaching functional consequences. Data are available at MetaboLights under the submission number: MTBLS1775.

Interleukin-1ß inhibits normal hematopoietic expansion and promotes acute myeloid leukemia progression via the bone marrow niche.

Enhanced interleukin-1ß (IL-1ß) signaling is a common event in patients with acute myeloid leukemia (AML). It was previously demonstrated that chronic IL-1ß exposure severely impaired hematopoietic stem cell (HSC) self-renewal capability in mice and promoted leukemia cell growth in primary AML cells. However, the role of IL-1ß in the murine bone marrow (BM) niche remains unclear. Here, we explored the role of IL-1ß in the BM niche in Il-1r1-/- mice, chronic IL-1ß exposure mice and mixed lineage leukemia-AF9 fusion gene (MLL-AF9)-induced AML mice models. We demonstrated that IL-1R1 deficiency did not affect the function of HSCs or niche cells under steady-state conditions or during transplantation. Chronic exposure to IL-1ß decreased the expansion of Il-1r1-/- hematopoietic cells in Il-1r1+/+ recipient mice. These results indicated that IL-1ß exposure impaired the ability of niche cells to support hematopoietic cells. Furthermore, we revealed that IL-1R1 deficiency in niche cells prolonged the survival of MLL-AF9-induced AML mice. The results of our study suggest that inhibition of the IL-1ß/IL-1R1 signaling pathway in the niche might be a non-cell-autonomous therapy strategy for AML.

Variable VOCs in plastic culture flasks and their potential impact on cell volatile biomarkers.

In order to find out cancer markers in human breath, in vitro cell culture is often used to study the characteristic volatile organic compounds (VOCs). In the cell culture process, disposable vessels are frequently adopted. However, these vessels are normally made of plastic, and they have the possibility to release some VOCs, which may interfere with the cell-specific volatiles and even can result in an incorrect conclusion. In this study, by using glass cell culture flasks as control, the headspace solid-phase microextraction gas chromatography mass spectrometry (HS-SPME-GC-MS) analyses of the VOCs in plastic cell culture flasks were systematically carried out for the first time. A total of 35 VOCs were detected in five brands of flasks. In each flask, there were between 13 and 25 volatile compounds. Furthermore, the components and packaging bag of each flask were also sampled and analyzed by HS-SPME-GC-MS. The results show that the flask cap, septum, flask body, and packaging bag exhibit respectively different volatile behaviors. The former two parts release the most volatiles which have obvious contributions to the headspace gases in the flasks, while the flask body mainly liberates styrene. For different flasks packed within the same bag, the headspace analyses show that their residual VOCs are inconsistent with each other. Moreover, the residual VOCs in the same flask are variable in three consecutive days. These results indicate that the multiple flasks in parallel cell culture experiments, or the same flask with different cell culture durations, will produce an indelible disturbance to the cell-specific VOCs. In addition, among the 35 VOCs detectable in five brands of empty plastic flasks, 15 VOCs were previously reported as characteristic VOCs from lung cancer, melanoma, cervical cancer cells, or normal cells. This is an alert that, when using plastic flasks, it must be careful to treat the possible interference from the background VOCs in the flasks. This study demonstrates that the cell culture tool needs to be standardized, and the clean glass or metal vessels are strongly recommended for usage when studying cell volatile biomarkers. Graphical abstract.

Six1 regulates leukemia stem cell maintenance in acute myeloid leukemia.

Molecular genetic changes in acute myeloid leukemia (AML) play crucial roles in leukemogenesis, including recurrent chromosome translocations, epigenetic/spliceosome mutations and transcription factor aberrations. Six1, a transcription factor of the Sine oculis homeobox (Six) family, has been shown to transform normal hematopoietic progenitors into leukemia in cooperation with Eya. However, the specific role and the underlying mechanism of Six1 in leukemia maintenance remain unexplored. Here, we showed increased expression of SIX1 in AML patients and murine leukemia stem cells (c-Kit+ cells, LSCs). Importantly, we also observed that a higher level of Six1 in human patients predicts a worse prognosis. Notably, knockdown of Six1 significantly prolonged the survival of MLL-AF9-induced AML mice with reduced peripheral infiltration and tumor burden. AML cells from Six1-knockdown (KD) mice displayed a significantly decreased number and function of LSC, as assessed by the immunophenotype, colony-forming ability and limiting dilution assay. Further analysis revealed the augmented apoptosis of LSC and decreased expression of glycolytic genes in Six1 KD mice. Overall, our data showed that Six1 is essential for the progression of MLL-AF9-induced AML via maintaining the pool of LSC.

Rheb1 loss leads to increased hematopoietic stem cell proliferation and myeloid-biased differentiation in vivo.

Hematopoietic stem cells constitute a unique subpopulation of blood cells that can give rise to all types of mature cells in response to physiological demands. However, the intrinsic molecular machinery that regulates this transformative property remains elusive. In this paper, we demonstrate that small GTPase Rheb1 is a critical regulator of proliferation and differentiation of hematopoietic stem cells in vivo Rheb1 deletion led to increased phenotypic hematopoietic stem cell/hematopoietic progenitor cell proliferation under a steady state condition. Over-proliferating Rheb1-deficient hematopoietic stem cells were severely impaired in functional repopulation assays, and they failed to regenerate the blood system when challenged with hematopoietic ablation by sublethal irradiation. In addition, it was discovered that Rheb1 loss resulted in a lack of maturation of neutrophils / caused neutrophil immaturation by reducing mTORC1 activity, and that activation of the mTORC1 signaling pathway by mTOR activator 3BDO partially restored the maturation of Rheb1-deficient neutrophils. Rheb1 deficiency led to a progressive enlargement of the hematopoietic stem cell population and an eventual excessive myeloproliferation in vivo, including an overproduction of peripheral neutrophils and an excessive expansion of extramedullary hematopoiesis. Moreover, low RHEB expression was correlated with poor survival in acute myeloid leukemia patients with normal karyotype. Our results, therefore, demonstrate a critical and unique role for Rheb1 in maintaining proper hematopoiesis and myeloid differentiation.

Acetylation targets the M2 isoform of pyruvate kinase for degradation through chaperone-mediated autophagy and promotes tumor growth.

Most tumor cells take up more glucose than normal cells but metabolize glucose via glycolysis even in the presence of normal levels of oxygen, a phenomenon known as the Warburg effect. Tumor cells commonly express the embryonic M2 isoform of pyruvate kinase (PKM2) that may contribute to the metabolism shift from oxidative phosphorylation to aerobic glycolysis and tumorigenesis. Here we show that PKM2 is acetylated on lysine 305 and that this acetylation is stimulated by high glucose concentration. PKM2 K305 acetylation decreases PKM2 enzyme activity and promotes its lysosomal-dependent degradation via chaperone-mediated autophagy (CMA). Acetylation increases PKM2 interaction with HSC70, a chaperone for CMA, and association with lysosomes. Ectopic expression of an acetylation mimetic K305Q mutant accumulates glycolytic intermediates and promotes cell proliferation and tumor growth. These results reveal an acetylation regulation of pyruvate kinase and the link between lysine acetylation and CMA.

Hematopoietic IKBKE limits the chronicity of inflammasome priming and metaflammation.

Obesity increases the risk of developing life-threatening metabolic diseases including cardiovascular disease, fatty liver disease, diabetes, and cancer. Efforts to curb the global obesity epidemic and its impact have proven unsuccessful in part by a limited understanding of these chronic progressive diseases. It is clear that low-grade chronic inflammation, or metaflammation, underlies the pathogenesis of obesity-associated type 2 diabetes and atherosclerosis. However, the mechanisms that maintain chronicity and prevent inflammatory resolution are poorly understood. Here, we show that inhibitor of κB kinase epsilon (IKBKE) is a novel regulator that limits chronic inflammation during metabolic disease and atherosclerosis. The pathogenic relevance of IKBKE was indicated by the colocalization with macrophages in human and murine tissues and in atherosclerotic plaques. Genetic ablation of IKBKE resulted in enhanced and prolonged priming of the NLRP3 inflammasome in cultured macrophages, in hypertrophic adipose tissue, and in livers of hypercholesterolemic mice. This altered profile associated with enhanced acute phase response, deregulated cholesterol metabolism, and steatoheptatitis. Restoring IKBKE only in hematopoietic cells was sufficient to reverse elevated inflammasome priming and these metabolic features. In advanced atherosclerotic plaques, loss of IKBKE and hematopoietic cell restoration altered plaque composition. These studies reveal a new role for hematopoietic IKBKE: to limit inflammasome priming and metaflammation.

PBX3 is essential for leukemia stem cell maintenance in MLL-rearranged leukemia.

Interaction of HOXA9/MEIS1/PBX3 is responsible for hematopoietic system transformation in MLL-rearranged (MLL-r) leukemia. Of these genes, HOXA9 has been shown to be critical for leukemia cell survival, while MEIS1 has been identified as an essential regulator for leukemia stem cell (LSC) maintenance. Although significantly high expression of PBX3 was observed in clinical acute myeloid leukemia (AML) samples, the individual role of PBX3 in leukemia development is still largely unknown. In this study, we explored the specific role of PBX3 and its associated regulatory network in leukemia progression. By analyzing the clinical database, we found that the high expression of PBX3 is significantly correlated with a poor prognosis in AML patients. ChIP-Seq/qPCR analysis in MLL-r mouse models revealed aberrant epigenetic modifications with increased H3K79me2, and decreased H3K9me3 and H3K27me3 levels in LSCs, which may account for the high expression levels of Pbx3. To further examine the role of Pbx3 in AML maintenance and progression, we used the CRISPR/Cas9 system to delete Pbx3 in leukemic cells in the MLL-AF9 induced AML mouse model. We found that Pbx3 deletion significantly prolonged the survival of leukemic mice and decreased the leukemia burden by decreasing the capacity of LSCs and promoting LSC apoptosis. In conclusion, we found that PBX3 is epigenetically aberrant in the LSCs of MLL-r AML and is essential for leukemia development. Significantly, the differential expression of PBX3 in normal and malignant hematopoietic cells suggests PBX3 as a potential prognostic marker and therapeutic target for MLL-r leukemia.

Liquid Extraction Surface Analysis Mass Spectrometry Method for Identifying the Presence and Severity of Nonalcoholic Fatty Liver Disease.

The early stages of nonalcoholic fatty liver disease (NAFLD) are characterized by the accumulation of fat in the liver (steatosis). This can lead to cell injury and inflammation resulting in nonalcoholic steatohepatitis (NASH). To determine whether lipid profiling of liver tissue can identify metabolic signatures associated with disease presence and severity, we explored liquid extraction surface analysis mass spectrometry (LESA-MS) as a novel sampling tool. Using LESA-MS, lipids were extracted directly from the surface of ultrathin slices of liver tissue prior to detection by high-resolution mass spectrometry (MS). An isotopically labeled internal standard mix was incorporated into the extraction solvent to attain semiquantitative data. Data mining and multivariate statistics were employed to evaluate the generated lipid profiles and abundances. With this approach, we were able to differentiate healthy and NAFLD liver in mouse and human tissue samples, finding several triacylglyceride (TAG) and free fatty acid (FFA) species to be significantly increased. Furthermore, LESA-MS was able to successfully differentiate between simple steatosis and more severe NASH, based on a set of short-chain TAGs and FFAs. We compared the data obtained by LESA-MS to that from liquid chromatography (LC)-MS and matrix-assisted laser desorption ionization MS. Advantages of LESA-MS include rapid analysis, minimal sample preparation, and high lipid coverage. Furthermore, since tissue slices are routinely used for diagnostics in clinical settings, LESA-MS is ideally placed to complement traditional histology. Overall LESA-MS is found to be a robust, fast, and discriminating approach for determining NAFLD presence and severity in clinical samples.

Phosphoinositide-dependent kinase 1 regulates leukemia stem cell maintenance in MLL-AF9-induced murine acute myeloid leukemia.

Although great efforts have been made to improve available therapies, the mortality rate of acute myeloid leukemia (AML) remains high due to poor treatment response and frequent relapse after chemotherapy. Leukemia stem cells (LSCs) are thought to account for this poor prognosis and relapse. Phosphoinositide-dependent kinase 1 (PDK1) is a critical regulator of the PI3K/Akt pathway and has been shown to be frequently activated in leukemia. However, the role of PDK1 in the regulation of LSCs in AML is still not clear. Using a PDK1 conditional deletion MLL-AF9 murine AML model, we revealed that the deletion of PDK1 prolonged the survival of AML mice by inducing LSC apoptosis. This was accompanied by the increased expression of the pro-apoptotic genes Bax and p53 and the reduced expression of Stat5, which has been shown to be constitutively activated in leukemia. Thus, our findings suggest that PDK1 plays an essential role in maintaining LSCs. Further delineating the function of PDK1 in LSCs may provide a new strategy for the improved treatment of AML relapse.

Pseudouridine synthase 1 regulates erythropoiesis via transfer RNAs pseudouridylation and cytoplasmic translation.

Pseudouridylation plays a regulatory role in various physiological and pathological processes. A prime example is the mitochondrial myopathy, lactic acidosis, and sideroblastic anemia syndrome (MLASA), characterized by defective pseudouridylation resulting from genetic mutations in pseudouridine synthase 1 (PUS1). However, the roles and mechanisms of pseudouridylation in normal erythropoiesis and MLASA-related anemia remain elusive. We established a mouse model carrying a point mutation (R110W) in the enzymatic domain of PUS1, mimicking the common mutation in human MLASA. Pus1-mutant mice exhibited anemia at 4 weeks old. Impaired mitochondrial oxidative phosphorylation was also observed in mutant erythroblasts. Mechanistically, mutant erythroblasts showed defective pseudouridylation of targeted tRNAs, altered tRNA profiles, decreased translation efficiency of ribosomal protein genes, and reduced globin synthesis, culminating in ineffective erythropoiesis. Our study thus provided direct evidence that pseudouridylation participates in erythropoiesis in vivo. We demonstrated the critical role of pseudouridylation in regulating tRNA homeostasis, cytoplasmic translation, and erythropoiesis.

Loss of RNA-binding protein CELF2 promotes acute leukemia development via FAT10-mTORC1.

RNA-binding proteins (RBPs) are critical regulators for RNA transcription and translation. As a key member of RBPs, ELAV-like family protein 2 (CELF2) has been shown to regulate RNA splicing and embryonic hematopoietic development and was frequently seen dysregulated in acute myeloid leukemia (AML). However, the functional role(s) of CELF2 in hematopoiesis and leukemogenesis has not been fully elucidated. In the current study, we showed that Celf2 deficiency in hematopoietic system led to enhanced HSCs self-renewal and differentiation toward myeloid cells in mice. Loss of Celf2 accelerated myeloid cell transformation and AML development in MLL-AF9-induced AML murine models. Gene expression profiling integrated with RNA immunoprecipitation sequencing (RIP-Seq), together with biochemical experiments revealed that CELF2 deficiency stabilizes FAT10 mRNA, promotes FAT10 translation, thereby increases AKT phosphorylation and mTORC1 signaling pathway activation. Notably, combination therapy with a mTORC1 inhibitor (Rapamycin) and a MA9/DOTL1 inhibitor (EPZ-5676) reduced the leukemia burden in MLL-AF9 mice lacking Celf2 in vivo. Our study elucidated a novel mechanism by which the CELF2/FAT10-AKT/mTORC1 axis regulates the proliferation of normal blood cells and the development of AML, thus providing potential therapeutic targets for myeloid leukemia suppression.