EGFR promotes ALKBH5 nuclear retention to attenuate N6-methyladenosine and protect against ferroptosis in glioblastoma.

Growth factor receptors rank among the most important oncogenic pathways, but pharmacologic inhibitors often demonstrate limited benefit as monotherapy. Here, we show that epidermal growth factor receptor (EGFR) signaling repressed N6-methyladenosine (m6A) levels in glioblastoma stem cells (GSCs), whereas genetic or pharmacologic EGFR targeting elevated m6A levels. Activated EGFR induced non-receptor tyrosine kinase SRC to phosphorylate the m6A demethylase, AlkB homolog 5 (ALKBH5), thereby inhibiting chromosomal maintenance 1 (CRM1)-mediated nuclear export of ALKBH5 to permit sustained mRNA m6A demethylation in the nucleus. ALKBH5 critically regulated ferroptosis through m6A modulation and YTH N6-methyladenosine RNA binding protein (YTHDF2)-mediated decay of the glutamate-cysteine ligase modifier subunit (GCLM). Pharmacologic targeting of ALKBH5 augmented the anti-tumor efficacy of EGFR and GCLM inhibitors, supporting an EGFR-ALKBH5-GCLM oncogenic axis. Collectively, EGFR reprograms the epitranscriptomic landscape through nuclear retention of the ALKBH5 demethylase to protect against ferroptosis, offering therapeutic paradigms for the treatment of lethal cancers.

m6A-driven SF3B1 translation control steers splicing to direct genome integrity and leukemogenesis.

SF3B1 is the most mutated splicing factor (SF) in myelodysplastic syndromes (MDSs), which are clonal hematopoietic disorders with variable risk of leukemic transformation. Although tumorigenic SF3B1 mutations have been extensively characterized, the role of "non-mutated" wild-type SF3B1 in cancer remains largely unresolved. Here, we identify a conserved epitranscriptomic program that steers SF3B1 levels to counteract leukemogenesis. Our analysis of human and murine pre-leukemic MDS cells reveals dynamic regulation of SF3B1 protein abundance, which affects MDS-to-leukemia progression in vivo. Mechanistically, ALKBH5-driven 5' UTR m6A demethylation fine-tunes SF3B1 translation directing splicing of central DNA repair and epigenetic regulators during transformation. This impacts genome stability and leukemia progression in vivo, supporting an integrative analysis in humans that SF3B1 molecular signatures may predict mutational variability and poor prognosis. These findings highlight a post-transcriptional gene expression nexus that unveils unanticipated SF3B1-dependent cancer vulnerabilities.

RBM33 is a unique m6A RNA-binding protein that regulates ALKBH5 demethylase activity and substrate selectivity.

Regulation of RNA substrate selectivity of m6A demethylase ALKBH5 remains elusive. Here, we identify RNA-binding motif protein 33 (RBM33) as a previously unrecognized m6A-binding protein that plays a critical role in ALKBH5-mediated mRNA m6A demethylation of a subset of mRNA transcripts by forming a complex with ALKBH5. RBM33 recruits ALKBH5 to its m6A-marked substrate and activates ALKBH5 demethylase activity through the removal of its SUMOylation. We further demonstrate that RBM33 is critical for the tumorigenesis of head-neck squamous cell carcinoma (HNSCC). RBM33 promotes autophagy by recruiting ALKBH5 to demethylate and stabilize DDIT4 mRNA, which is responsible for the oncogenic function of RBM33 in HNSCC cells. Altogether, our study uncovers the mechanism of selectively demethylate m6A methylation of a subset of transcripts during tumorigenesis that may explain demethylation selectivity in other cellular processes, and we showed its importance in the maintenance of tumorigenesis of HNSCC.

Dynamic control of chromatin-associated m6A methylation regulates nascent RNA synthesis.

N6-methyladenosine (m6A) methylation is co-transcriptionally deposited on mRNA, but a possible role of m6A on transcription remains poorly understood. Here, we demonstrate that the METTL3/METTL14/WTAP m6A methyltransferase complex (MTC) is localized to many promoters and enhancers and deposits the m6A modification on nascent transcripts, including pre-mRNAs, promoter upstream transcripts (PROMPTs), and enhancer RNAs. PRO-seq analyses demonstrate that nascent RNAs originating from both promoters and enhancers are significantly decreased in the METTL3-depleted cells. Furthermore, genes targeted by the Integrator complex for premature termination are depleted of METTL3, suggesting a potential antagonistic relationship between METTL3 and Integrator. Consistently, we found the Integrator complex component INTS11 elevated at promoters and enhancers upon loss of MTC or nuclear m6A binders. Taken together, our findings suggest that MTC-mediated m6A modification protects nascent RNAs from Integrator-mediated termination and promotes productive transcription, thus unraveling an unexpected layer of gene regulation imposed by RNA m6A modification.

FOXD3-mediated transactivation of ALKBH5 promotes neuropathic pain via m6A-dependent stabilization of 5-HT3A mRNA in sensory neurons.

The N6-methyladenosine (m6A) modification of RNA is an emerging epigenetic regulatory mechanism that has been shown to participate in various pathophysiological processes. However, its involvement in modulating neuropathic pain is still poorly understood. In this study, we elucidate a functional role of the m6A demethylase alkylation repair homolog 5 (ALKBH5) in modulating trigeminal-mediated neuropathic pain. Peripheral nerve injury selectively upregulated the expression level of ALKBH5 in the injured trigeminal ganglion (TG) of rats. Blocking this upregulation in injured TGs alleviated trigeminal neuropathic pain, while mimicking the upregulation of ALKBH5 in intact TG neurons sufficiently induced pain-related behaviors. Mechanistically, histone deacetylase 11 downregulation induced by nerve injury increases histone H3 lysine 27 acetylation (H3K27ac), facilitating the binding of the transcription factor forkhead box protein D3 (FOXD3) to the Alkbh5 promoter and promoting Alkbh5 transcription. The increased ALKBH5 erases m6A sites in Htr3a messenger RNA (mRNA), resulting in an inability of YT521-B homology domain 2 (YTHDF2) to bind to Htr3a mRNA, thus causing an increase in 5-HT3A protein expression and 5-HT3 channel currents. Conversely, blocking the increased expression of ALKBH5 in the injured TG destabilizes nerve injury-induced 5-HT3A upregulation and reverses mechanical allodynia, and the effect can be blocked by 5-HT3A knockdown. Together, FOXD3-mediated transactivation of ALKBH5 promotes neuropathic pain through m6A-dependent stabilization of Htr3a mRNA in TG neurons. This mechanistic understanding may advance the discovery of new therapeutic targets for neuropathic pain management.

Identification of differential m6A RNA methylomes and ALKBH5 as a potential prevention target in the developmental neurotoxicity induced by multiple sevoflurane exposures.

Sevoflurane, as a commonly used inhaled anesthetic for pediatric patients, has been reported that multiple sevoflurane exposures are associated with a greater risk of developing neurocognitive disorder. N6-Methyladenosine (m6A), as the most common mRNA modification in eukaryotes, has emerged as a crucial regulator of brain function in processes involving synaptic plasticity, learning and memory, and neurodevelopment. Nevertheless, the relevance of m6A RNA methylation in the multiple sevoflurane exposure-induced developmental neurotoxicity remains mostly elusive. Herein, we evaluated the genome-wide m6A RNA modification and gene expression in hippocampus of mice that received with multiple sevoflurane exposures using m6A-sequencing (m6A-seq) and RNA-sequencing (RNA-seq). We discovered 19 genes with differences in the m6A methylated modification and differential expression in the hippocampus. Among these genes, we determined that a total of nine differential expressed genes may be closely associated with the occurrence of developmental neurotoxicity induced by multiple sevoflurane exposures. We further found that the alkB homolog 5 (ALKBH5), but not methyltransferase-like 3 (METTL3) and Wilms tumor 1-associated protein (WTAP), were increased in the hippocampus of mice that received with multiple sevoflurane exposures. And the IOX1, as an inhibitor of ALKBH5, significantly improved the learning and memory defects and reduced neuronal damage in the hippocampus of mice induced by multiple sevoflurane exposures. The current study revealed the role of m6A methylated modification and m6A-related regulators in sevoflurane-induced cognitive impairment, which might provide a novel insight into identifying biomarkers and therapeutic strategies for inhaled anesthetic-induced developmental neurotoxicity.

ALKBH5-mediated upregulation of CPT1A promotes macrophage fatty acid metabolism and M2 macrophage polarization, facilitating malignant progression of colorectal cancer.

m6A modification has been studied in tumors, but its role in host anti-tumor immune response and TAMs polarization remains unclear. The fatty acid oxidation (FAO) process of TAMs is also attracting attention. A co-culture model of colorectal cancer (CRC) cells and macrophages was used to simulate the tumor microenvironment. Expression changes of m6A demethylase genes FTO and ALKBH5 were screened. ALKBH5 was further investigated. Gain-of-function experiments were conducted to study ALKBH5's effects on macrophage M2 polarization, CRC cell viability, proliferation, migration, and more. Me-RIP and Actinomycin D assays were performed to study ALKBH5's influence on CPT1A, the FAO rate-limiting enzyme. AMP, ADP, and ATP content detection, OCR measurement, and ECAR measurement were used to explore ALKBH5's impact on macrophage FAO level. Rescue experiments validated ALKBH5's mechanistic role in macrophage M2 polarization and CRC malignant development. In co-culture, CRC cells enhance macrophage FAO and suppress m6A modification in M2 macrophages. ALKBH5 was selected as the gene for further investigation. ALKBH5 mediates CPT1A upregulation by removing m6A modification, promoting M2 macrophage polarization and facilitating CRC development. These findings indicate that ALKBH5 enhances fatty acid metabolism and M2 polarization of macrophages by upregulating CPT1A, thereby promoting CRC development.

RNA m6A demethylase ALKBH5 regulates the development of γδ T cells.

γδ T cells are an abundant T cell population at the mucosa and are important in providing immune surveillance as well as maintaining tissue homeostasis. However, despite γδ T cells' origin in the thymus, detailed mechanisms regulating γδ T cell development remain poorly understood. N6-methyladenosine (m6A) represents one of the most common posttranscriptional modifications of messenger RNA (mRNA) in mammalian cells, but whether it plays a role in γδ T cell biology is still unclear. Here, we show that depletion of the m6A demethylase ALKBH5 in lymphocytes specifically induces an expansion of γδ T cells, which confers enhanced protection against gastrointestinal Salmonella typhimurium infection. Mechanistically, loss of ALKBH5 favors the development of γδ T cell precursors by increasing the abundance of m6A RNA modification in thymocytes, which further reduces the expression of several target genes including Notch signaling components Jagged1 and Notch2. As a result, impairment of Jagged1/Notch2 signaling contributes to enhanced proliferation and differentiation of γδ T cell precursors, leading to an expanded mature γδ T cell repertoire. Taken together, our results indicate a checkpoint role of ALKBH5 and m6A modification in the regulation of γδ T cell early development.

Loss of m6A demethylase ALKBH5 alleviates hypoxia-induced pulmonary arterial hypertension via inhibiting Cyp1a1 mRNA decay.

BACKGROUND: Hypoxia-induced pulmonary artery hypertension (HPH) is a complication of chronic hypoxic lung disease and the third most common type of pulmonary artery hypertension (PAH). Epigenetic mechanisms play essential roles in the pathogenesis of HPH. N6-methyladenosine (m6A) is an important modified RNA nucleotide involved in a variety of biological processes and an important regulator of epigenetic processes. To date, the precise role of m6A and regulatory molecules in HPH remains unclear. METHODS: HPH model and pulmonary artery smooth muscle cells (PASMCs) were constructed from which m6A changes were observed and screened for AlkB homolog 5 (Alkbh5). Alkbh5 knock-in (KI) and knock-out (KO) mice were constructed to observe the effects on m6A and evaluate right ventricular systolic pressure (RVSP), left ventricular and septal weight [RV/(LV + S)], and pulmonary vascular remodeling in the context of HPH. Additionally, the effects of Alkbh5 knockdown using adenovirus were examined in vitro on m6A, specifically in PASMCs with regard to proliferation, migration and cytochrome P450 1A1 (Cyp1a1) mRNA stability. RESULTS: In both HPH mice lung tissues and hypoxic PASMCs, a decrease in m6A was observed, accompanied by a significant up-regulation of Alkbh5 expression. Loss of Alkbh5 attenuated the proliferation and migration of hypoxic PASMCs in vitro, with an associated increase in m6A modification. Furthermore, Alkbh5 KO mice exhibited reduced RVSP, RV/(LV + S), and attenuated vascular remodeling in HPH mice. Mechanistically, loss of Alkbh5 inhibited Cyp1a1 mRNA decay and increased its expression through an m6A-dependent post-transcriptional mechanism, which hindered the proliferation and migration of hypoxic PASMCs. CONCLUSION: The current study highlights the loss of Alkbh5 impedes the proliferation and migration of PASMCs by inhibiting post-transcriptional Cyp1a1 mRNA decay in an m6A-dependent manner.

ALKBH5 regulates ovarian cancer growth via demethylating long noncoding RNA PVT1 in ovarian cancer.

The long noncoding RNA PVT1 is reported to act as an oncogene in several kinds of cancers, especially ovarian cancer (OV). Abnormal levels of N6 -methyladenosine, a dynamic and reversible modification, are associated with tumorigenesis and malignancies. Our previous study reported that PVT1 plays critical roles in regulating OV. However, it is still largely unknown how m6 A modification affects OV via PVT1. In this study, we aimed to investigate the regulation of ALKBH5 by affecting PVT1 in OV. We first found that the PVT1 RNA level was higher in OV cells than in IOSE80 cells, and conversely, the m6 A modification level of PVT1 was lower in OV cells. By searching the HPA, ALKBH5, which is responsible for PVT1 demethylation, was found to be upregulated in OV tissues versus normal ovarian tissues. ALKBH5 binds to PVT1 RNA, and knockdown of ALKBH5 decreased PVT1 RNA levels. ALKBH5 also increased FOXM1 levels by upregulating PVT1, at least partially. Knockdown of ALKBH5 suppressed OV growth, colony formation, tumour formation and invasion, which were partially reversed by overexpression of PVT1. Moreover, ALKBH5 knockdown decreased FOXM1 levels by regulating PVT1 RNA expression, subsequently increasing the sensitivity to carboplatin, 5-FU and docetaxel chemotherapy. Taken together, these results indicate that ALKBH5 directly regulates the m6 A modification and stability of PVT1. Then, modified PVT1 further regulates FOXM1 and thus affects malignant behaviours and chemosensitivity in OV cells. All these results indicate that ALKBH5 regulates the malignant behaviour of OV by regulating PVT1/FOXM1.

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.

The disordered C terminus of ALKBH5 promotes phase separation and paraspeckles assembly.

Paraspeckles (PS) are nuclear structures scaffolded by the long noncoding RNA NEAT1 and protein components such as NONO and SFPQ. We previously found that the upregulation of RNA N6-methyl-adenosine (m6A) demethylase ALKBH5 facilitates hypoxia-induced paraspeckle assembly through erasing m6A marks on NEAT1, thus stabilizing it. However, it remains unclear how these processes are spatiotemporally coordinated. Here we discover that ALKBH5 specifically binds to proteins in PS and forms phase-separated droplets that are incorporated into PS through its C-terminal intrinsically disordered region (cIDR). Upon exposure to hypoxia, rapid ALKBH5 condensation in PS induces m6A demethylation of NEAT1, which further facilitates PS formation before the upregulation of ALKBH5 expression. In cells expressing ALKBH5 lacking cIDR, PS fail to be formed in response to hypoxia, accompanied with insufficient m6A demethylation of NEAT1 and its destabilization. We also demonstrate that ALKBH5-cIDR is indispensable for hypoxia-induced effects such as cancer cell invasion. Therefore, our study has identified the role of ALKBH5 in phase separation as the molecular basis of the positive feedback loop for PS formation between ALKBH5 incorporation into PS and NEAT1 stabilization.

ALKBH5 regulates chicken adipogenesis by mediating LCAT mRNA stability depending on m6A modification.

BACKGROUND: Previous studies have demonstrated the role of N6-methyladenosine (m6A) RNA methylation in various biological processes, our research is the first to elucidate its specific impact on LCAT mRNA stability and adipogenesis in poultry. RESULTS: The 6 100-day-old female chickens were categorized into high (n = 3) and low-fat chickens (n = 3) based on their abdominal fat ratios, and their abdominal fat tissues were processed for MeRIP-seq and RNA-seq. An integrated analysis of MeRIP-seq and RNA-seq omics data revealed 16 differentially expressed genes associated with to differential m6A modifications. Among them, ELOVL fatty acid elongase 2 (ELOVL2), pyruvate dehydrogenase kinase 4 (PDK4), fatty acid binding protein 9 (PMP2), fatty acid binding protein 1 (FABP1), lysosomal associated membrane protein 3 (LAMP3), lecithin-cholesterol acyltransferase (LCAT) and solute carrier family 2 member 1 (SLC2A1) have ever been reported to be associated with adipogenesis. Interestingly, LCAT was down-regulated and expressed along with decreased levels of mRNA methylation methylation in the low-fat group. Mechanistically, the highly expressed ALKBH5 gene regulates LCAT RNA demethylation and affects LCAT mRNA stability. In addition, LCAT inhibits preadipocyte proliferation and promotes preadipocyte differentiation, and plays a key role in adipogenesis. CONCLUSIONS: In conclusion, ALKBH5 mediates RNA stability of LCAT through demethylation and affects chicken adipogenesis. This study provides a theoretical basis for further understanding of RNA methylation regulation in chicken adipogenesis.

Hypoxia-induced ALKBH5 aggravates synovial aggression and inflammation in rheumatoid arthritis by regulating the m6A modification of CH25H.

Previous studies have shown that epigenetic factors are involved in the occurrence and development of rheumatoid arthritis (RA). However, the role of N6-methyladenosine (m6A) methylation in RA has not been determined. The aim of this study was to investigate the role and regulatory mechanisms of hypoxia-induced expression of the m6A demethylase alkB homolog 5 (ALKBH5) in RA fibroblast-like synoviocytes (FLSs). Synovial tissues were collected from RA and osteoarthritis (OA) patients, and RA FLSs were obtained. ALKBH5 expression in RA FLSs and collagen-induced arthritis (CIA) model rats was determined using quantitative reverse transcription-PCR (qRT-PCR), western blotting and immunohistochemistry (IHC). Using ALKBH5 overexpression and knockdown, we determined the role of ALKBH5 in RA FLS aggression and inflammation. The role of ALKBH5 in RA FLS regulation was explored using m6A-methylated RNA sequencing and methylated RNA immunoprecipitation coupled with quantitative real-time PCR. The expression of ALKBH5 was increased in RA synovial tissues, CIA model rats and RA FLSs, and a hypoxic environment increased the expression of ALKBH5 in FLSs. Increased expression of ALKBH5 promoted the proliferation and migration of RA-FLSs and inflammation. Conversely, decreased ALKBH5 expression inhibited the migration of RA-FLSs and inflammation. Mechanistically, hypoxia-induced ALKBH5 expression promoted FLS aggression and inflammation by regulating CH25H mRNA stability. Our study elucidated the functional roles of ALKBH5 and mRNA m6A methylation in RA and revealed that the HIF1α/2α-ALKBH5-CH25H pathway may be key for FLS aggression and inflammation. This study provides a novel approach for the treatment of RA by targeting the HIF1α/2α-ALKBH5-CH25H pathway.

ALKBH5 regulates somatic cell reprogramming in a phase-specific manner.

Establishment of the pluripotency regulatory network in somatic cells by introducing four transcription factors [octamer binding transcription factor 4 (OCT4; also known as POU5F1), sex determining region Y (SRY)-box 2 (SOX2), Kruppel-like factor 4 (KLF4) and cellular myelocytomatosis (c-MYC)] provides a promising tool for cell-based therapies in regenerative medicine. Nevertheless, the mechanisms at play when generating induced pluripotent stem cells from somatic cells are only partly understood. Here, we show that the RNA-specific N6-methyladenosine (m6A) demethylase ALKBH5 regulates somatic cell reprogramming in a stage-specific manner through its catalytic activity. Knockdown or knockout of Alkbh5 in the early reprogramming phase impairs reprogramming efficiency by reducing the proliferation rate through arresting the cells at G2/M phase and decreasing the upregulation of epithelial markers. On the other hand, ALKBH5 overexpression at the early reprogramming phase has no significant impact on reprogramming efficiency, whereas overexpression at the late phase enhances reprogramming by stabilizing Nanog transcripts, resulting in upregulated Nanog expression. Our study provides mechanistic insight into the crucial dynamic role of ALKBH5, mediated through its catalytic activity, in regulating somatic cell reprogramming at the post-transcriptional level. This article has an associated First Person interview with the first author of the paper.

The methyltransferase METTL3 regulates endothelial cell proliferation and inflammation via m6A RNA methylation-mediated TRAF1 expression.

The imbalance of vascular endothelial cell homeostasis is the key mechanism for the progression of many vascular diseases. RNA modification, particularly N6-Methyladenosine (m6A), plays important function in numerous biological processes. Nevertheless, the regulatory function of m6A RNA methylation in endothelial dysfunction remains insufficiently characterized. In this study, we established that the m6A methyltransferase METTL3 is critical for regulating endothelial function. Functionally, depletion of METTL3 results in decreased endothelial cells proliferation, survival and inflammatory response. Conversely, overexpression of METTL3 elicited the opposite effects. Mechanistically, MeRIP-seq identified that METTL3 catalyzed m6A modification of TRAF1 mRNA and enhanced TRAF1 translation, thereby up-regulation of TRAF1 protein. Over-expression of TRAF1 successfully rescued the inhibition of proliferation and adhesion of endothelial cells due to METTL3 knockdown. Additionally, m6A methylation-mediated TRAF1 expression can be reversed by the demethylase ALKBH5. Knockdown of ALKBH5 upregulated the level of m6A and protein level of TRAF1, and also increased endothelial cells adhesion and inflammatory response. Collectively, our findings suggest that METTL3 regulates vascular endothelium homeostasis through TRAF1 m6A modification, suggesting that targeting the METTL3-m6A-TRAF1 axis may hold therapeutic potential for patients with vascular diseases.

ALKBH5 regulates paclitaxel resistance in NSCLC via inhibiting CEMIP-mediated EMT.

N6-methyladenosine (m6A) is the most prevalent mRNA modification, and it is verified to be closely correlated with cancer occurrence and progression. The m6A demethylase ALKBH5 (alkB homolog 5) is dysregulated in various cancers. However, the role and underlying mechanism of ALKBH5 in the pathogenesis and especially the chemo-resistance of non-small cell lung cancer (NSCLC) is poorly elucidated. The current study shows that ALKBH5 expression is reduced in paclitaxel (PTX) resistant NSCLC cells and down-regulation of ALKBH5 usually implies poor prognosis of NSCLC patients. Over-expression of ALKBH5 in PTX-resistant cells can suppress cell proliferation and enhance chemo-sensitivity, while knockdown of ALKBH5 exerts the opposite effect, which further supports the tumor suppressive role of ALKBH5. Over-expression of ALKBH5 can also reverse the epithelial-mesenchymal transition (EMT) process in PTX-resistant cancer cells. Mechanistically, data from RNA-seq, real-time PCR and western blotting indicate that CEMIP (cell migration inducing hyaluronidase 1), also known as KIAA1199, may be the downstream target of ALKBH5. Furthermore, ALKBH5 negatively regulates the CEMIP level by reducing the stability of CEMIP mRNA. Collectively, the current data demonstrate that the ALKBH5/CEMIP axis modulates the EMT process in NSCLC, which in turn regulates the chemo-sensitivity of cancer cells to PTX.

Complicated role of ALKBH5 in gastrointestinal cancer: an updated review.

Gastrointestinal cancer is the most common malignancy in humans, often accompanied by poor prognosis. N6-methyladenosine (m6A) modification is widely present in eukaryotic cells as the most abundant RNA modification. It plays a crucial role in RNA splicing and processing, nuclear export, translation, and stability. Human AlkB homolog 5 (ALKBH5) is a type of RNA demethylase exhibiting abnormal expression in various gastrointestinal cancers.It is closely related to the tumorigenesis, proliferation, migration, and other biological functions of gastrointestinal cancer. However, recent studies indicated that the role and mechanism of ALKBH5 in gastrointestinal cancer are complicated and even controversial. Thus, this review summarizes recent advances in elucidating the role of ALKBH5 as a tumor suppressor or promoter in gastrointestinal cancer. It examines the biological functions of ALKBH5 and its potential as a therapeutic target, providing new perspectives and insights for gastrointestinal cancer research.

Downregulation of ALKBH5 rejuvenates aged human mesenchymal stem cells and enhances their therapeutic efficacy in myocardial infarction.

Despite promising results in myocardial infarction (MI), mesenchymal stem cell (MSC)-based therapy is limited by cell senescence. N6-methyladenosine (m6A) messenger RNA methylation has been reported to be closely associated with cell senescence. Nonetheless, its role in the regulation of MSC senescence remains unclear. We examined the role of ALKB homolog 5 (ALKBH5) in regulating MSC senescence and determined whether ALKBH5 downregulation could rejuvenate aged MSCs (AMSCs) to improve their therapeutic efficacy for MI. RNA methylation was determined by m6A dot blotting assay. MSC senescence was evaluated by senescence-associated ß-galactosidase (SA-ß-gal) staining. A mouse model of acute MI was established by ligation of the left anterior decedent coronary artery (LAD). Compared with young MSCs (YMSCs), m6A level was significantly reduced but ALKBH5 was greatly increased in AMSCs. Overexpression of ALKBH5 reduced m6A modification and accelerated YMSC senescence. Conversely, ALKBH5 knockdown increased m6A modifications and alleviated AMSC senescence. Mechanistically, ALKBH5 regulated the m6A modification and stability of CDKN1C mRNA, which further upregulated CDKN1C expression, leading to MSC senescence. CDKN1C overexpression ameliorated the inhibition of cellular senescence of ALKBH5 siRNA-treated AMSCs. More importantly, compared with AMSCs, shALKBH5-AMSCs transplantation provided a superior cardioprotective effect against MI in mice by improving MSC survival and angiogenesis. We determined that ALKBH5 accelerated MSC senescence through m6A modification-dependent stabilization of the CDKN1C transcript, providing a potential target for MSC rejuvenation. ALKBH5 knockdown rejuvenated AMSCs and enhanced cardiac function when transplanted into the mouse heart following infarction.

ALKBH5 facilitates the progression of infantile hemangioma by increasing FOXF1 expression in a m6A-YTHDF2 dependent manner to activate HK-2 signaling.

Alkylation repair homolog protein 5 (ALKBH5) is reported to participate in infantile hemangioma (IH) progression. However, the underlying mechanism of ALKBH5 in IH remains unclear. Using qRT-PCR and Western blotting, ALKBH5, forkhead box F1 (FOXF1) and hexokinase 2 (HK-2) expressions in IH tissues and IH-derived endothelial cells XPTS-1 were assessed. The Me-RIP assay was used to analyze FOXF1 m6A level. CCK8, colony formation, flow cytometry and transwell assays were employed to determine IH cell viability, proliferation, apoptosis, migration and invasion. The interactions between YTH (YT521-B homology) domain 2 (YTHDF2), FOXF1 and HK-2 were analyzed by RIP, dual luciferase reporter gene assay and/or ChIP assay. The in vivo IH growth was evaluated in immunocompromised mice. FOXF1 was overexpressed in IH tissues, and its silencing inhibited IH cell proliferation, migration and invasion whereas promoting cell apoptosis in vitro. ALKBH5 upregulation facilitated FOXF1 mRNA stability and expression in IH cells in a m6A-YTHDF2-dependent manner. FOXF1 downregulation reversed the impact of ALKBH5 upregulation on IH cellular phenotypes. It also turned out that FOXF1 positively regulated HK-2 expression in IH cells through interacting with the HK-2 promoter. HK-2 upregulation abolished FOXF1 knockdown's inhibition on IH cell aggressive behaviors. ALKBH5 or FOXF1 silencing suppressed IH tumor development via HK-2 signaling in immunocompromised mice. ALKBH5 promoted FOXF1 expression m6A-YTHDF2 dependently, which in turn elevated HK-2 expression, thereby accelerating IH development.