m6A transferase METTL3 regulates endothelial-mesenchymal transition in diabetic retinopathy via lncRNA SNHG7/KHSRP/MKL1 axis

Diabetic retinopathy is one of the microvascular complications in diabetic patients and the leading cause of blindness worldwide. The levels of METTL3, lncRNA SNHG7, KHSRP, MKL1, endothelial and mesenchymal markers were determined by RT-qPCR or western blot assays in vitro and in vivo. H&E staining was used to observe the retinal structure in a mouse model of DR. The expression levels of METTL3 and SNHG7 were significantly downregulated in DR patients, DR mice and high glucose-induced HRMECs cells. Notably, METTL3 installed the m6A modification and enhanced the stability of SNHG7. Besides, METTL3 inhibited HRMECs EndoMT by promoting the expression of SNHG7. Additionally, SNHG7 was found to weaken MKL1 mRNA stability by binding to the RNA-binding protein KHSRP. Furthermore, we verified that METTL3 regulated EndoMT in DR through the SNHG7/MKL1 axis. We conclude that METTL3 regulates endothelial-mesenchymal transition in DR via the SNHG7/KHSRP/MKL1 axis, providing a new target for DR treatment.     

Involvement of the MAPK and PI3K pathways in chitinase 3-like 1-regulated hyperoxia-induced airway epithelial cell death

BackgroundExposure to 100% oxygen causes hyperoxic acute lung injury characterized by cell death and injury of alveolar epithelial cells. Recently, the role of chitinase 3-like 1 (CHI3L1), a member of the glycosyl hydrolase 18 family that lacks chitinase activity, in oxidative stress was demonstrated in murine models. High levels of serum CHI3L1 have been associated with various diseases of the lung, such as asthma, chronic obstructive pulmonary disease, and cancer. However, the role of CHI3L1 in human airway epithelial cells undergoing oxidative stress remains unknown. In addition, the signaling pathways associated with CHI3L1 in this process are poorly understood.PurposeIn this study, we demonstrate the role of CHI3L1, along with the MAPK and PI3K signaling pathways, in hyperoxia-exposed airway epithelial cells.MethodThe human airway epithelial cell line, BEAS-2B, was exposed to >95% oxygen (hyperoxia) for up to 72 h. Hyperoxia-induced cell death was determined by assessing cell viability, Annexin-V FITC staining, caspase-3 and -7 expression, and electron microscopy. CHI3L1 knockdown and overexpression studies were conducted in BEAS-2B cells to examine the role of CHI3L1 in hyperoxia-induced apoptosis. Activation of the MAPK and PI3K pathways was also investigated to determine the role of these signaling cascades in this process.ResultsHyperoxia exposure increased CHI3L1 expression and apoptosis in a time-dependent manner. CHI3L1 knockdown protected cells from hyperoxia-induced apoptosis. In contrast, CHI3L1 overexpression promoted cell death after hyperoxia exposure. Finally, phosphorylation of ERK1/2, p38, and Akt were affected by CHI3L1 knockdown.ConclusionThis study indicates that CHI3L1 is involved in hyperoxia-induced cell death, suggesting that CHI3L1 may be one of several cell death regulators influencing the MAPK and PI3K pathways during oxidative stress in human airway epithelial cells.     

LncRNA DGUOK-AS1 facilitates non-small cell lung cancer growth and metastasis through increasing TRPM7 stability via m6A modification

BackgroundN6-methyladenosine (m6A) modification plays key roles in tumor progression. LncRNA deoxyguanosine kinase antisense RNA 1 (DGUOK-AS1) has been reported as a promoter in tumors, but its role and mechanism in non-small cell lung cancer (NSCLC) development remain uncertain.MethodsCell proliferation, migration, invasion and angiogenesis were investigated via CCK-8, colony formation, transwell, and tube formation assays, respectively. The location of DGUOK-AS1 was detected via FISH assay. The interaction relationship among DGUOK-AS1, IGF2BP2 and TRPM7 was confirmed by RIP and MeRIP assays. The effects of DGUOK-AS1 on NSCLC growth and metastasis in vivo were investigated using xenograft and pulmonary metastatic models.ResultsDGUOK-AS1 was upregulated in NSCLC. DGUOK-AS1 silencing inhibited NSCLC cell proliferation, migration, invasion and angiogenesis. DGUOK-AS1 was mostly expressed in cytoplasm, and positively regulated IGF2BP2. METTL3/IGF2BP2 axis could increase TRPM7 mRNA stability in m6A-dependent manner. TRPM7 overexpression reversed the inhibitive function of DGUOK-AS1 silencing on NSCLC development. DGUOK-AS1 knockdown suppressed NSCLC cell growth and metastasis in nude mice.ConclusionDGUOK-AS1 silencing restrains NSCLC cell growth and metastasis through decreasing TRPM7 stability via regulation of the METTL3/IGF2BP2-mediated m6A modification.     

METTL14-regulated PI3K/Akt signaling pathway via PTEN affects HDAC5-mediated epithelial–mesenchymal transition of renal tubular cells in diabetic kidney disease

Histone deacetylase 5 (HDAC5) belongs to class II HDAC subfamily and is reported to be increased in the kidneys of diabetic patients and animals. However, little is known about its function and the exact mechanism in diabetic kidney disease (DKD). Here, we found that HDAC5 was located in renal glomeruli and tubular cells, and significantly upregulated in diabetic mice and UUO mice, especially in renal tubular cells and interstitium. Knockdown of HDAC5 ameliorated high glucose-induced epithelial–mesenchymal transition (EMT) of HK2 cells, indicated in the increased E-cadherin and decreased α-SMA, via the downregulation of TGF-β1. Furthermore, HDAC5 expression was regulated by PI3K/Akt signaling pathway and inhibition of PI3K/Akt pathway by LY294002 treatment or Akt phosphorylation mutation reduced HDAC5 and TGF-β1 expression in vitro high glucose-cultured HK2 cells. Again, high glucose stimulation downregulated total m6A RNA methylation level of HK2 cells. Then, m6A demethylase inhibitor MA2 treatment decreased Akt phosphorylation, HDAC5, and TGF-β1 expression in high glucose-cultured HK2 cells. In addition, m6A modification-associated methylase METTL3 and METTL14 were decreased by high glucose at the levels of mRNA and protein. METTL14 not METTL3 overexpression led to PI3K/Akt pathway inactivation in high glucose-treated HK2 cells by enhancing PTEN, followed by HDAC5 and TGF-β1 expression downregulation. Finally, in vivo HDACs inhibitor TSA treatment alleviated extracellular matrix accumulation in kidneys of diabetic mice, accompanied with HDAC5, TGF-β1, and α-SMA expression downregulation. These above data suggest that METTL14-regulated PI3K/Akt signaling pathway via PTEN affected HDAC5-mediated EMT of renal tubular cells in diabetic kidney disease.Subject terms: Epithelial-mesenchymal transition, Insulin signalling, Diabetes complications     

METTL3对猪干细胞多能基因表达的调控作用

m~6A是真核生物m RNA中重要的转录后修饰,METTL3作为m~6A甲基转移酶复合物中的重要组分,在细胞重编程、胚胎干细胞和诱导多能干细胞的干性维持、胚胎发育等过程中发挥重要作用。为了揭示猪METTL3的表达模式,对不同物种METTL3蛋白序列进行了比对,用RT-PCR检测了METTL3基因在不同猪组织和细胞中的表达情况,并确认了METTL3的细胞核定位。为了研究METTL3对猪干细胞多能基因表达的调控作用,克隆了猪METTL3编码区序列,设计了METTL3干扰片段,并构建了相应的过表达和沉默载体。发现干扰METTL3的表达后,猪多能干细胞出现类似na?ve状态的细胞克隆,NANOG、OCT4和LIN28A表达水平显著升高。在猪多能干细胞培养基中添加m~6A甲基化抑制剂环亮氨酸培养细胞48 h后,试验结果与干扰METTL3表达的结果一致。本研究为优化猪多能干细胞的培养体系提供了新的方向和依据。     

Mesenchymal stem cells alleviate experimental immune-mediated liver injury via chitinase 3-like protein 1-mediated T cell suppression

Liver diseases with different pathogenesis share common pathways of immune-mediated injury. Chitinase-3-like protein 1 (CHI3L1) was induced in both acute and chronic liver injuries, and recent studies reported that it possesses an immunosuppressive ability. CHI3L1 was also expressed in mesenchymal stem cells (MSCs), thus we investigates the role of CHI3L1 in MSC-based therapy for immune-mediated liver injury here. We found that CHI3L1 was highly expressed in human umbilical cord MSCs (hUC-MSCs). Downregulating CHI3L1 mitigated the ability of hUC-MSCs to inhibit T cell activation, proliferation and inflammatory cytokine secretion in vitro. Using Concanavalin A (Con A)-induced liver injury mouse model, we found that silencing CHI3L1 significantly abrogated the hUC-MSCs-mediated alleviation of liver injury, accompanying by weakened suppressive effects on infiltration and activation of hepatic T cells, and secretion of pro-inflammatory cytokines. In addition, recombinant CHI3L1 (rCHI3L1) administration inhibited the proliferation and function of activated T cells, and alleviated the Con A-induced liver injury in mice. Mechanistically, gene set enrichment analysis showed that JAK/STAT signalling pathway was one of the most significantly enriched gene pathways in T cells co-cultured with hUC-MSCs with CHI3L1 knockdown, and further study revealed that CHI3L1 secreted by hUC-MSCs inhibited the STAT1/3 signalling in T cells by upregulating peroxisome proliferator-activated receptor δ (PPARδ). Collectively, our data showed that CHI3L1 was a novel MSC-secreted immunosuppressive factor and provided new insights into therapeutic treatment of immune-mediated liver injury.Subject terms: T cells, Mesenchymal stem cells, Autoimmune hepatitis     

MiR‐103‐3p targets the m6A methyltransferase METTL14 to inhibit osteoblastic bone formation

Impaired osteoblast function is involved in osteoporosis, and microRNA (miRNA) dysregulation may cause abnormal osteoblast osteogenic activity. However, the influence of miRNA on osteoblast activity and the underlying mechanisms remain elusive. In this study, miR‐103‐3p was found to be negatively correlated with bone formation in bone specimens from elderly women with fractures and ovariectomized (OVX) mice. Additionally, miR‐103‐3p directly targeted Mettl14 to inhibit osteoblast activity, and METTL14‐dependent N⁶‐methyladenosine (m⁶A) methylation inhibited miR‐103‐3p processing by the microprocessor protein DGCR8 and promoted osteoblast activity. Moreover, miR‐103‐3p inhibited bone formation in vivo, and therapeutic inhibition of miR‐103‐3p counteracted the decreased bone formation in OVX mice. Further, METTL14 was negatively correlated with miR‐103‐3p but positively correlated with bone formation in bone specimens from elderly women with fractures and OVX mice. Collectively, our results highlight the critical roles of the miR‐103‐3p/METTL14/m⁶A signaling axis in osteoblast activity, identifying this axis as a potential target for ameliorating osteoporosis.     

CircMETTL3, upregulated in a m6A-dependent manner,promotes breast cancer progression

Growing evidence indicates N6-methyladenosine (m6A) has biological function in oncogenesis. METTL3, the catalytic component, is the most important part of methyltransferase complex and plays a crucial role in cancers. However, the biological function of circRNAs derived from METTL3 in breast cancer and the underlying molecular mechanism remains unclear. Herein, we report circMETTL3, which has not been explored in breast cancer, and it is markedly upregulated in breast cancer. Moreover, we uncovered that circMETTL3 could facilitate cell proliferation, migration and invasion in breast cancer. Mechanism investigation showed that circMETTL3 might act as a competing endogenous RNA (ceRNA) of miR-31-5p and upregulate its target cyclin-dependent kinases (CDK1). Moreover, m6A modification of circMETTL3 might affect its expression. Taken together, our results elucidate that circMETTL3 promotes breast cancer progression through circMETTL3/miR-31-5p/CDK1 axis. Moreover, METTL3, the host gene of circMETTL3, may regulate circMETTL3 expression in an m6A-dependent manner, while circMETTL3 has no effect on METTL3 expression, providing a new relationship between the circRNA and the corresponding host gene. Thus, it may serve as a new therapeutic target for breast cancer.     

METTL3-mediated m6A modification of STEAP2 mRNA inhibits papillary thyroid cancer progress by blocking the Hedgehog signaling pathway and epithelial-to-mesenchymal transition

Papillary thyroid cancer (PTC) is a common endocrine system malignancy all over the world. Aberrant expression of six transmembrane epithelial antigen of the prostate 2 (STEAP2) has been functionally associated with cancer progression in many cancers. Nevertheless, its biological function in PTC is still unclear. Here, we found that PTC tissues had preferentially downregulated STEAP2 as compared with noncancerous tissues. Low STEAP2 expression correlated with aggressive clinicopathological characteristics and dismal prognosis in patients with PTC. We performed gain- and loss-of-function experiments, including cell proliferation assay (Cell Counting Kit-8 assay), EdU (5-ethynyl-2′-deoxyuridine) and colony formation assays, transwell migration, and invasion assays, and constructed a nude mouse xenograft tumor model. The results demonstrated that STEAP2 overexpression inhibited PTC cell proliferation, migration, and invasion in vitro and inhibited lung metastasis and tumorigenicity in vivo. Conversely, silencing STEAP2 yielded the opposite results in vitro. Mechanistically, bioinformatics analysis combined with validation experiments identified STEAP2 as the downstream target of methyltransferase-like 3 (METTL3)-mediated N6-methyladenosine (m6A) modification. METTL3 stabilized STEAP2 mRNA and regulated STEAP2 expression positively in an m6A-dependent manner. We also showed that m6A-mediated STEAP2 mRNA translation initiation relied on a pathway dependent on the m6A reader protein YTHDF1. Rescue experiments revealed that silencing STEAP2 partially rescued the tumor-suppressive phenotype induced by METTL3 overexpression. Lastly, we verified that the METTL3–STEAP2 axis functions as an inhibitor in PTC by suppressing epithelial–mesenchymal transition and the Hedgehog signaling pathway. Taken together, these findings strongly suggest that METTL3-mediated STEAP2 m6A modification plays a critical tumor-suppressive role in PTC progression. The METTL3–STEAP2 axis may be a potential therapeutic molecular target against PTC.Subject terms: Metastasis, Prognostic markers     

CircRNA VIM silence synergizes with sevoflurane to inhibit immune escape and multiple oncogenic activities of esophageal cancer by simultaneously regulating miR-124/PD-L1 axis

Background

Circular RNA of vimentin (circ-VIM) is a predictor for poor prognosis of acute myeloid leukemia, but we had little information on its function in esophageal cancer (EC). Here we examined the effects of circ-VIM together with sevoflurane on immune escape and multiple oncogenic activities of EC.

Methods

Bioinformatic tools, luciferase assay, and RNA immunoprecipitation were used to examine regulations between circ-VIM, miR-124-3p (miR-124), and PD-L1. CCK-8, wound healing, and Transwell assays were used to measure cell proliferation, migration, and invasion, respectively. The impacts of EC cells on cytotoxicity, proliferation, and apoptosis of CD8⁺ T cells were examined using LDH assay, CFSE staining, and Annexin V/PI staining, respectively. The in vivo tumorigenesis and lung metastases were assessed using xenograft model and tail vein injection of EC cells.

Results

Significant upregulation of circ-VIM and PD-L1 and downregulation of miR-124 were detected in EC tissues or cells. Circ-VIM sponged miR-124 and released its suppression on the downstream target PD-L1. Sevoflurane, independent of circ-VIM, also upregulated miR-124 to lower PD-L1 expression. By modulating miR-124/PD-L1 axis, silencing circ-VIM and applying sevoflurane both inhibited immune escape and multiple oncogenic activities of EC in vitro, and suppressed xenograft growth and lung metastases in vivo. The inactivation of Ras/ERK signaling pathway was involved in suppression of malignant phenotypes by silencing circ-VIM and sevoflurane treatment.

Conclusions

Silencing circ-VIM and applying sevoflurane, by separately regulating miR-124/PD-L1 axis, presented synergistic effects in inhibiting immune escape and multiple malignant phenotypes of EC cells.

Graphical abstract

     

METTL3 promotes prostatic hyperplasia by regulating PTEN expression in an m6A-YTHDF2-dependent manner

Uncontrolled epithelial cell proliferation in the prostate transition zone and the hyper-accumulation of mesenchymal-like cells derived from the epithelial-mesenchymal transition (EMT) of prostatic epithelium are two key processes in benign prostatic hyperplasia (BPH). m⁶A RNA modification affects multiple cellular processes, including cell proliferation, apoptosis, and differentiation. In this study, the aberrant up-regulation of methylase METTL3 in BPH samples suggests its potential role in BPH development. Elevated m⁶A modification in the prostate of the BPH rat was partially reduced by METTL3 knockdown. METTL3 knockdown also partially reduced the prostatic epithelial thickness and prostate weight, significantly improved the histological features of the prostate, inhibited epithelial proliferation and EMT, and promoted apoptosis. In vitro, METTL3 knockdown decreased TGF-β-stimulated BPH-1 cell proliferation, m⁶A modification, and EMT, whereas promoted cell apoptosis. METTL3 increased the m⁶A modification of PTEN and inhibited its expression through the reading protein YTHDF2. PTEN knockdown aggravated the molecular, cellular, and pathological alterations in the prostate of BPH rats and amplified TGF-β-induced changes in BPH-1 cells. More importantly, PTEN knockdown partially abolished the improving effects of METTL3 knockdown both in vivo and in vitro. In conclusion, the level of m⁶A modification is elevated in BPH; the METTL3/YTHDF2/PTEN axis disturbs the balance between epithelial proliferation and apoptosis, promotes EMT, and accelerates BPH development in an m⁶A modification-related manner.Subject terms: Cell biology, Molecular biology     

Pseudogene PTENP1 sponges miR-214 to regulate the expression of PTEN to modulate osteoclast differentiation and attenuate osteoporosis

Background aimsOsteoporosis (OP) is a common bone metabolic disease with a high incidence. Our study aimed to explore the pseudogene PTENP1/miR-214/PTEN axis to modulate the osteoclast differentiation in osteoporosis.MethodsPatients with osteoporosis were recruited in our study, and RANKL-induced osteoclast differentiation and ovariectomy-induced osteoporosis mouse model were established in vitro and in vivo, respectively.ResultsPseudogene PTENP1 and PTEN were significantly down-regulated and miR-214 was up-regulated in osteoporosis patients. In addition, overexpression of PTENP1 or silence of miR-214 inhibited the expression levels of osteoclast specific markers and osteoclast differentiation induced by RANKL. Overexpression of PTENP1 or silence of miR-214 also inhibited the levels of phosphorylation of PI3K and AKT, p65 nuclear translocation, IκBα degradation and the expression level of NFATc1. AlsoSilence of PTENP1 or overexpression of miR-214 induced the osteoclast differentiation under normal physiological condition. Pseudogene PTENP1 sponged miR-214 to regulate the expression of PTEN.ConclusionsIn an ovariectomy-induced osteoporosis mouse model, obvious pathological changes in bone tissues were found, and bone marrow mononuclear cells in this group were more likely to differentiate into osteoclasts. Therefore, pseudogene PTENP1 sponged miR-214 to regulate the expression of PTEN to inhibit osteoclast differentiation and attenuate osteoporosis by suppressing the PI3K/AKT/NF-κB signaling pathway.     

Chitinase 3-Like 1 Protein Levels Are Elevated in Schistosoma haematobium Infected Children

Background

Currently there are few studies characterising the nature and aetiology of human schistosome-related inflammatory processes. The aim of this study was to determine the relationship between Chitinase 3-like 1 (CHI3L1), also known as YKL-40, a molecule associated with inflammatory processes, and schistosome infection, morbidity and systemic cytokine levels.

Methods

Serological levels of CHI3L1 and a panel of cytokines (IFN-y, IL-4/5/6/9/10/13 and 17) were measured in two Zimbabwean populations resident in a high and low schistosome infection area. CHI3L1 levels were related to schistosome infection, haematuria status and cytokine levels after allowing for confounding variables. The effect of antihelminthic treatment with praziquantel on CHI3L1 levels was determined in 246 participants 6 weeks post-treatment.

Results

CHI3L1 levels increased with age in both areas but were significantly higher in the high infection areas compared to the low infection area. CHI3L1 levels were also higher in infected compared to uninfected individuals with this difference being significant in the youngest age group. Curative antihelminthic treatment resulted in a significant decrease in CHI3L1 levels. Of the cytokines, only IL-10 and IL-17 had a significant association with CHI3L1 levels, and this association was negative.

Conclusions

Serum CHI3L1 levels differ between infected and uninfected people before and after antihelminthic treatment. The greatest difference occurs in the youngest age group, in keeping with the period when schistosome-related pathological processes are initiated. Following from previous studies in non-infectious diseases showing that CHI3L1 is a biomarker for the inflammatory process, this study suggests that the potential for CHI3L1 as a biomarker for schistosome-related pathology should be explored further.     

Methyltransferase-like 3-mediated N6-methyladenosine modification of miR-7212-5p drives osteoblast differentiation and fracture healing

N6-methyladenosine (m6A) modification has been reported in various diseases and implicated in increasing numbers of biological processes. However, previous studies have not focused on the role of m6A modification in fracture healing. Here, we demonstrated that m6A modifications are decreased during fracture healing and that methyltransferase-like 3 (METTL3) is the main factor involved in the abnormal changes in m6A modifications. Down-regulation of METTL3 promotes osteogenic processes both in vitro and in vivo, and this effect is recapitulated by the suppression of miR-7212-5p maturation. Further studies have shown that miR-7212-5p inhibits osteoblast differentiation in MC3T3-E1 cells by targeting FGFR3. The present study demonstrated an important role of the METTL3/miR-7212-5p/FGFR3 axis and provided new insights on m6A modification in fracture healing.     

METTL14 promotes tumorigenesis by regulating lncRNA OIP5-AS1/miR-98/ADAMTS8 signaling in papillary thyroid cancer

BackgroundPapillary thyroid cancer (PTC) is the most common type of cancer of the endocrine system. Long noncoding RNAs (lncRNAs) are emerging as a novel class of gene expression regulators associated with tumorigenesis. Through preexisting databases available for differentially expressed lncRNAs in PTC, we uncovered that lncRNA OIP5-AS1 was significantly upregulated in PTC tissues. However, the function and the underlying mechanism of OIP5-AS1 in PTC are poorly understood.MethodsExpression of lncRNA OIP5-AS1 and miR-98 in PTC tissue and cells were measured by quantitative real-time PCR (qRT-PCR). And expression of METTL14 and ADAMTS8 in PTC tissue and cells were measured by qRT-PCR and western blot. The biological functions of METTL14, OIP5-AS1, and ADAMTS8 were examined using MTT, colony formation, transwell, and wound healing assays in PTC cells. The relationship between METTL14 and OIP5-AS1 were evaluated using RNA immunoprecipitation (RIP) and RNA pull down assay. And the relationship between miR-98 and ADAMTS8 were examined by luciferase reporter assay. For in vivo experiments, a xenograft model was used to investigate the effects of OIP5-AS1 and ADAMTS8 in PTC.ResultsFunctional validation revealed that OIP5-AS1 overexpression promotes PTC cell proliferation, migration/invasion in vitro and in vivo, while OIP5-AS1 knockdown shows an opposite effect. Mechanistically, OIP5-AS1 acts as a target of miR-98, which activates ADAMTS8. OIP5-AS1 promotes PTC cell progression through miR-98/ADAMTS8 and EGFR, MEK/ERK pathways. Furthermore, RIP and RNA pull down assays identified OIP5-AS1 as the downstream target of METTL14. Overexpression of METTL14 suppresses PTC cell proliferation and migration/invasion through inhibiting OIP5-AS1 expression and regulating EGFR, MEK/ERK pathways.ConclusionsCollectively, our findings demonstrate that OIP5-AS1 is a METTL14-regulated lncRNA that plays an important role in PTC progression and offers new insights into the regulatory mechanisms underlying PTC development.Subject terms: Tumour biomarkers, Oncogenes     

The HNF1A-AS1/miR-92a-3p axis affects the radiosensitivity of non-small cell lung cancer by competitively regulating the JNK pathway

Background

It has been widely reported that long non-coding RNAs (lncRNAs) could affect the varieties of tumor response to radiotherapy. LncRNA HNF1A-AS1 is transcribed from HNF1A gene cluster’s antisense strand. This work focused on the mechanism of how HNF1A-AS1 participated in the radiosensitivity of non-small cell lung cancer (NSCLC).

Methods

The mRNA or protein expression of HNF1A-AS1, miR-92a-3p MAP2K4, and JNK in NSCLC cells and tissues was detected by qRT-PCR or western blotting. RNA immunoprecipitation (RIP) detection and luciferase reporting system were used to evaluate the relationship between HNFA-AS1 and miR-92a-3p or between miR-92a-3p and MAP2K4. Flow cytometry assays, colony formation, and MTT were performed to analyze the function changes in A549 and Calu-1 cells. The rescue experiment was also conducted to explore the underlying mechanisms.

Results

HNF1A-AS1 was investigated in NSCLC cells and tissues and highly related to the advanced pathological stage. HNF1A-AS1 bound with miR-92a-3p, which was downregulated in NSCLC. It showed that miR-92a-3p was negatively related to HNF1A-AS1. Knockdown of HNF1A-AS1 impacted most cell biological behaviors in NSCLC cells, including restricting the proliferation and aggravating apoptosis. Furthermore, knockdown of HNF1A-AS1 dramatically enhanced radiotherapy sensitivity of NSCLC. Moreover, miR-92a-3p was found to target MAP2K4 and could reduce MAP2K4 expression. Inhibition of HNF1A-AS1 elevated radiotherapy sensitivity and retarded the progression of NSCLC cells, followed by decreasing expression levels of MAP2K4. Besides, MAP2K4 mimic rescued the si-HNF1A-AS1 effects on the biological behavior of NSCLC cells.

Conclusion

HNF1A-AS1 is highly expressed in NSCLC. MiR-92a-3p is the target gene of HNF1A-AS1 and involved in tumor progression by regulating the MAP2K4/JNK pathway. HNF1AS1/miR-92a-3p/MAP2K4 axis plays important roles in radiotherapy resistance of NSCLC.

Graphical abstract