ALYREF promotes malignant behaviors and inhibits ferroptosis in colon cancer cells by stabilizing PCSK9 mRNA

BACKGROUNDThe incidence of colon cancer (CC) is currently high, and is mainly treated with chemotherapy. Oxaliplatin (L-OHP) is a commonly used drug in chemotherapy; however, long-term use can induce drug resistance and seriously affect the prognosis of patients. Therefore, this study investigated the mechanism of Opa-interacting protein 5 antisense RNA 1 (OIP5-AS1) on L-OHP resistance by determining the expression of OIP5-AS1 and microRNA-137 (miR-137) in CC cells and the effects on L-OHP resistance, with the goal of identifying new targets for the treatment of CC.AIMTo study the effects of long non-coding RNA OIP5-AS1 on L-OHP resistance in CC cell lines and its regulation of miR-137.METHODSA total of 114 CC patients admitted to China-Japan Union Hospital of Jilin University were enrolled, and the expression of miR-137 and OIP5-AS1 in tumor tissues and corresponding normal tumor-adjacent tissues was determined. The influence of OIP5-AS1 and miR-137 on the biological behavior of CC cells was evaluated. Resistance to L-OHP was induced in CC cells, and their activity was determined and evaluated using cell counting kit-8. Flow cytometry was used to analyze the apoptosis rate, Western blot to determine the levels of apoptosis-related proteins, and dual luciferase reporter assay combined with RNA-binding protein immunoprecipitation to analyze the relationship between OIP5-AS1 and miR-137.RESULTSOIP5-AS1 was up-regulated in CC tissues and cells, while miR-137 was down-regulated in CC tissues and cells. OIP5-AS1 was inversely correlated with miR-137 (P < 0.001). Silencing OIP5-AS1 expression significantly hindered the proliferation, invasion and migration abilities of CC cells and markedly increased the apoptosis rate. Up-regulation of miR-137 expression also suppressed these abilities in CC cells and increased the apoptosis rate. Moreover, silencing OIP5-AS1 and up-regulating miR-137 expression significantly intensified growth inhibition of drug-resistant CC cells and improved the sensitivity of CC cells to L-OHP. OIP5-AS1 targetedly inhibited miR-137 expression, and silencing OIP5-AS1 reversed the resistance of CC cells to L-OHP by promoting the expression of miR-137.CONCLUSIONHighly expressed in CC, OIP5-AS1 can affect the biological behavior of CC cells, and can also regulate the resistance of CC cells to L-OHP by mediating miR-137 expression.

With a high mortality rate, colon cancer (CC) is the third most common malignant tumor worldwide. The primary causes are thought to be the high invasiveness and migration of CC cells. The functions of Golgi phosphoprotein 3 (GOLPH3), stress-induced phosphoprotein 1 (STIP1), and the signal transducer and activator of transcription 3 (STAT3) signaling pathway in the invasion and migration of CC cells were examined in this study. We collected the exosomes by high-speed centrifugation. The expressions of GOLPH3, STIP1, and epithelial-mesenchymal transition (EMT)-related proteins in CC tissues, cells, and exosomes were analyzed using Western blotting (WB) experiments. The abilities of CC cell invasion and migration were evaluated by the Transwell assay. The binding relationship between GOLPH3 and STIP1 was validated through Co-immunoprecipitation (Co-IP), and their sublocalization in CC cells was determined by immunofluorescence detection under laser confocal microscopy. Immunohistochemistry (IHC) experiments detected the expression levels of each protein in the transplanted tumor mass. Animal experiments confirmed the impact of the GOLPH3/STIP1/STAT3 regulatory axis on CC growth. We found that in CC tissues and cells, GOLPH3 was highly expressed, and silencing GOLPH3 not only greatly reduced CC cell invasion and migration but also prevented EMT. Furthermore, GOLPH3 and STIP1 interacted in CC cells, and the GOLPH3-STIP1 complex affected the capacity for cell invasion and migration by triggering the STAT3 signaling pathway. Noteworthily, GOLPH3, and STIP1 could also be detected in CC cell exosomes, and the exosomes carried the GOLPH3-ST1P1 complex to act on CC cells to activate intracellular STAT3 signaling, ultimately affecting the cancer cell migration and invasion. The above molecular regulatory mechanisms have also been validated in mice. In conclusion, the GOLPH3-STIP1 complex acted on surrounding CC cells through exosomes and activated the STAT3 signaling pathway to stimulate CC cell invasion and migration.

Alkaline phosphatase and dipeptidyl peptidase are markers of differentiation in colon cancer cells. In colon cancer cells differentiation can often be induced by treatment with inhibitors of histone deacetylase (HDAC) activity. In some colon cancer cells butyrate causes a several-fold induction of alkaline phosphatase but only modest effects on dipeptidyl peptidase. There is relatively little information on changes in activity of cell surface hydrolases in bladder cancer. However, there is some evidence for decreased alkaline phosphatase activity in bladder cancer. We tested the hypothesis that there may be retention of activity in more slowly growing cancer cells and the activity will be regulated by HDAC inhibitors in a manner similar to that in colon cancer. In our initial studies we screened seven human bladder cancer cell lines for alkaline phosphatase activity. We identified three cell lines (5637, HT1197 and HT1376) with activity comparable to that in more differentiated colon cancer cell lines. The response to incubation with the HDAC inhibitors butyrate and valproate was compared with effects in three colon cancer cell lines Caco-2, HT29 and SW1116. In all six cell lines there were growth inhibitory effects that tended to be a little greater with butyrate than with valproate. Induction of alkaline phosphatase activity was generally greater with butyrate than with valproate and was apparent even without normalizing the activity on a protein basis. After determining the activity per unit protein there were increases in dipeptidyl peptidase activity after treatment with the HDAC inhibitors that were smaller than for alkaline phosphatase. Growth inhibition was observed with all the HDAC inhibitors examined including LMK235, nicotinamide, RGFP966, romidepsin and tubacin but increased activity of alkaline phosphatase was not always seen. Activities of the enzymes were very low in CRL1790 human colonocytes of fetal origin and there was little or no induction with HDAC inhibitors. Although pyruvate and carnitine have been reported to be HDAC inhibitors, incubation with colon and bladder cancer cells did not cause induction of alkaline phosphatase activity. It was concluded that regulation of alkaline phosphatase and dipeptidyl peptidase by HDAC inhibitors can be similar in bladder and colon cancer cells. Citation Format: Michael A. Lea, Lauren Cué, Elizabeth Batista, Erik Lew, Charles desBordes. Effects of histone deacetylase inhibitors on differentiation markers and growth of colon and bladder cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1386. doi:10.1158/1538-7445.AM2017-1386

Ferroptosis, a genetically and biochemically distinct form of programmed cell death, is characterized by iron-dependent accumulation of lipid peroxides. Ferroptosis is induced in cancer cells by inhibition of lipid peroxide quencher GPx4. System xc - imports cystine into cytosol for the biosynthesis of glutathione. Tumor cells that are resistant to chemotherapeutic drugs called ‘drug tolerant’ or ‘Persister’ cells which have distinct vulnerability towards iron mediated cell death or ferroptosis. Unfolded Protein Response (UPR) plays critical role for cancer cells to become drug tolerant. Tweaking the balance of UPR to make drug tolerant cells susceptible to Ferroptotic cell death could be an attractive therapeutic strategy. To decipher the emerging contribution of ER-stress in ferroptosis, we investigated the status of UPR following treatment of potent ferroptosis inducer RSL3 (Ras Selective Lethal) in colon cancer cells. We observed an overall up-regulation of UPR activators (ATF6, IRE1α and PERK) and their downstream effectors along with a marked overexpression of cystine-glutamate transporter (System xc -). To further delineate the contribution of particular UPR arm in modulating System xc - expression and subsequent ferroptosis, we made stable knock down cells of each UPR arm and discovered that PERK is selective and critical in inducing ferroptosis in colon cancer cells. Loss of PERK function not only promotes ferroptosis via increasing lipid peroxidation but also limits in vivo tumor growth in colon xenograft model. Further, we find that low PERK expression is associated with higher patient survival as per TCGA COLON CANCER (COAD) database. Overall, our experimental data indicate that PERK is a negative regulator of ferroptosis and genetic silencing of PERK sensitizes colon cancer cells to selective Ferroptotic cell death. Therefore, small molecule PERK inhibitors hold huge promise as novel therapeutics that can sensitize apoptosis resistant cancer cells towards Ferroptotic cell death. Citation Format: Krishan K. Saini, Priyank Chaturvedi, Ayushi Verma, Mushtaq A. Nengroo, Abhipsa Sinha, Akhilesh Singh, Sanjeev Meena, Muqtada A. Khan, Manish P. Singh, Dipak Datta. PERK arm of UPR selectively regulates ferroptosis in colon cancer cells by modulating the expression of system xc - (SLC7A11) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6249.

Colon cancer undergoes a traditional pathway from colon polyps to colon cancer. It is of great significance to investigate the key molecules involved in carcinogenesis from polyps to malignancies. Circular RNAs (circRNAs) are stably expressed in human body fluids such as plasma. Here, we demonstrated a differential expression pattern of plasma circRNAs in healthy individuals, colon polyp patients and colon cancer patients using circRNA Arraystar microarray. We explored that circRNA HADHA (circHADHA) was upregulated in plasma from polyp patients, whereas it was downregulated in plasma from colon cancer patients. Overexpression of circHADHA promoted autophagy in colon epithelial cells. Moreover, in colon cancer cells, overexpression of circHADHA promoted autophagy, whereas it inhibited cell proliferation and colony formation. CircHADHA increased the expression of ATG13 via miR-361 in both colon epithelial and cancer cells. ATG13 knockdown reduced autophagy even in the presence of circHADHA in colon cancer cells. Furthermore, the growth of circHADHA-overexpressing colon cancer cell-derived xenograft tumors was significantly decreased compared with control tumors in nude mice. In conclusion, circHADHA was differentially expressed in the plasma of healthy individuals, colon polyp patients and colon cancer patients. CircHADHA promoted autophagy by regulating ATG13 via miR-361 in both colon epithelial and cancer cells. CircHADHA suppressed tumor growth by inducing cell autophagy in colon cancer cells. CircHADHA potentially serves as a biomarker for screening of precursor colon cancer and a therapeutic target for colon cancer treatment.

Rotenone, a natural hydrophobic pesticide, has been reported to display anticancer activity in a variety of cancer cells. However, the mechanism of rotenone on colon cancer (CC) cell migration, invasion and metastasis is still unknown. In the present study, the cytotoxicity of rotenone on CC cells were detected by the Cell Counting Kit-8 assay and confirmed by clone formation assay. The effects of rotenone on CC cell invasion and migration activity were determined in vitro by Transwell invasion and wound healing assays, respectively. In addition, to reveal whether rotenone affected the epithelial-mesenchymal-transition (EMT) process, reverse transcription-quantitative PCR, western blotting and immunofluorescence assays were used to detect the expression of EMT markers. The expression levels of the key markers of the PI3K/AKT pathway after rotenone treatment alone or in combination with a PI3K/AKT signaling activator in CC were also detected by western blotting. Finally, the in vivo anti-tumor effects of rotenone were evaluated in a subcutaneous xenotransplant tumor model treated with an intraperitoneal injection of rotenone. The results of the present study demonstrated that rotenone treatment induced CC cell cytotoxicity and greater effects were observed with increasing concentrations and inhibited cell proliferation compared with untreated cells. In vitro cell function assays revealed that rotenone inhibited CC cell migration, invasion and EMT compared with untreated cells. Mechanically, the phosphorylation levels of AKT and mTOR were downregulated in rotenone-treated CC cells compared with untreated cells. Additionally, AKT and mTOR phosphorylation levels were increased by the PI3K/AKT signaling activator insulin-like growth factor 1 (IGF-1), which was reversed by rotenone treatment. The cell function assays confirmed that the IGF-1-activated cell proliferation, migration and invasion were decreased by rotenone treatment. These results indicated that rotenone affected CC cell proliferation and metastatic capabilities by inhibiting the PI3K/AKT/mTOR signaling pathway. In addition, rotenone inhibited tumor growth and metastatic capability of CC, which was confirmed in a xenograft mouse model. In conclusion, the present study revealed that rotenone inhibited CC cell viability, motility, EMT and metastasis in vitro and in vivo by inhibiting the PI3K/AKT/mTOR signaling pathway.

The application of intestinal microflora is involved in various cancers; however, researches reporting the potential of metabolites of intestinal microflora (MIM) on biological activities of colon cancer (CC) cells are unavailable. This study was designed to testify the functions of MIM on CC cells and its mechanism. qRT-PCR/Western blot were applied to test the expression levels of miR-192-5p and BMPR2 in human colonic epithelial cells and CC cells (HCT116, SW480). The effects of MIM, mimics-miR-192-5p or inhibitors-miR-192-5p on mRNA and protein expressions of miR-192-5p and BMPR2 were verified by qRT-PCR and Western blot. MTT assay for CC cell viability, flow cytometry for CC cells apoptosis rate, and cell scratch and cell chamber served for the analysis of invasion and migration ability of CC cells. The relationship between miR-192-5p and BMPR2 was validated employing Luciferase reporter gene assay. Compared with human normal colonic epithelial cells, HCT116 and SW480 cells had lower expression of miR-192-5p and higher expression of BMPR2 (p < 0.01). MIM and mimics-miR-192-5p could enhance cell apoptosis and suppress the migration and proliferation of CC cells. MIM were also found to up-regulate miR-192-5p and down-regulate the expression levels of BMPR2 and p-LIMK2 (p < 0.01). Transfection of inhibitors-miR-192-5p reversed the inhibitory effect of MIM on CC cells. MIM could up-regulate miR-192-5p to inhibit CC cell growth via down-regulating BMPR2 and inhibiting the activity of RhoA-ROCK-LIMK2 pathway.

β1-Integrin is highly expressed and is a negative prognostic factor for colon and pancreatic cancer patients and the gene plays a functional role in cell migration and invasion. In this study, we demonstrate that β1-integrin expression is regulated in pancreatic and colon cancer cells by the pro-oncogenic orphan nuclear receptor 4A1 (NR4A1, Nur77, TR3) and knockdown of this receptor by RNA interference decreases β1-integrin protein and mRNA expression, α5-integrin, and also expression of β1-integrin-dependent phosphorylation of FAK (pFak). Knockdown of NR4A1 also decreased migration and fibronectin-induced adhesion in pancreatic (Panc1, L3.6 pL, and MiaPaCa2) and colon (RKO and SW480) cancer cells. 1,1-Bis(3'-indolyl)-1-(p-substituted phenyl)methane (C-DIM) compounds containing p-hydroxy (DIM-C-pPhOH) and p-carbomethoxy (DIM-C-pPhCO2 Me) groups are NR4A1 ligands that act as antagonists for this receptor. Treatment of pancreatic and colon cancer cells with DIM-C-pPhOH or DIM-C-pPhCO2 Me mimics the effects of NR4A1 knockdown and decreases β1-integrin expression, β1-integrin regulated genes and responses including migration and adhesion. The results demonstrate a novel method for targeting β1-integrin in colon and pancreatic cancer cells and indicate possible clinical applications for C-DIM/NR4A1 antagonists for pancreatic and colon cancer therapy.

ObjectiveSerum response factor (SRF), a sequence-specific transcription factor, is closely related to metastasis of gastric cancer, a digestive tract cancer. Herein, we probed the effect of SRF on metastasis and progression of colon cancer (CC), another digestive tract disorder, and the detailed mechanism.MethodsMicroarray analysis was conducted on tumor and adjacent tissues to filter differentially expressed miRNA, followed by RT-qPCR validation in CC cell lines. The transcription factor and the target gene of microRNA-214 (miR-214) were predicted, and their binding relationships were tested by luciferase reporter assays and ChIP assays. Subsequently, SRF and protein tyrosine kinase 6 (PTK6) expression in CC patients and cells was evaluated by RT-qPCR, while JAK2 and STAT3 expression in cells by Western blot analysis. To further explore functions of miR-214, PTK6 and SRF on CC, CC cells were delivered with si-PTK6, miR-214 mimic and/or SRF overexpression.ResultsmiR-214 expressed poorly in CC tissues and cell lines, which related to advanced TNM staging and survival. miR-214 mimic inhibited proliferation, migration, invasion, xenograft tumor growth and metastasis of CC cells. SRF, overexpressed in CC samples and cells, suppressed the transcription of miR-214. Meanwhile, SRF upregulation counteracted the inhibitory role of miR-214 mimic in CC cell growth. miR-214 negatively regulated PTK6 expression to impair the JAK2/STAT3 pathway activation, thereby halting CC cell proliferation, migration, invasion, xenograft tumor growth and metastasis.ConclusionAltogether, miR-214 may perform as a tumor suppressor in CC, and the SRF/miR-214/PTK6/JAK2/STAT3 axis could be applied as a biomarker and potential therapeutic target.

Colon cancer (CC) is a tumor of the large intestine. miR-92b-3p is often deregulated in the tumorigensis. Here, the role of miR-92b-3p in the development of CC was investigated. miR-92b-3p and Kruppel-like factor 3 (KLF3) expression was examined in CC tissues and cells. miR-92b-3p inhibitor or KLF3 overexpression vector was transfected into CC cells, respectively to observe its role in CC cell proliferation, invasion, migration, and apoptosis. The targeting relationship between miR-92b-3p and KLF3 was validated. Meanwhile, rescue experiments were performed by co-transfection of miR-92b-3p inhibitor and KLF3 siRNA, followed by determining CC cell proliferation, invasion, migration, and apoptosis. Higher miR-92b-3p and lower KLF3 expression levels were observed in CC tissues and cells. miR-92b-3p inhibition or KLF3 overexpression reduced proliferation, invasion, and migration whereas induced apoptosis of CC cells. KLF3 was validated to be the target gene of miR-92b-3p. Depletion of KLF3 could reverse the antitumor role of miR-92b-3p inhibition in CC cells. miR-92b-3p augments CC development through inhibiting KLF3, which may confers a novel way to develop future treatment target.

BackgroundHuman mitochondrial peptide deformylase (PDF) has been proposed as a novel cancer therapeutic target. However, very little is known about its expression and regulation in human tissues. The purpose of this study was to characterize the expression pattern of PDF in cancerous tissues and to identify mechanisms that regulate its expression.MethodsThe mRNA expression levels of PDF and methionine aminopeptidase 1D (MAP1D), an enzyme involved in a related pathway with PDF, were determined using tissue panels containing cDNA from patients with various types of cancer (breast, colon, kidney, liver, lung, ovarian, prostate, or thyroid) and human cell lines. Protein levels of PDF were also determined in 2 colon cancer patients via western blotting. Colon cancer cells were treated with inhibitors of ERK, Akt, and mTOR signaling pathways and the resulting effects on PDF and MAP1D mRNA levels were determined by qPCR for colon and lung cancer cell lines. Finally, the effects of a PDF inhibitor, actinonin, on the proliferation of breast, colon, and prostate cell lines were determined using the CyQUANT assay.ResultsPDF and MAP1D mRNA levels were elevated in cancer cell lines compared to non-cancer lines. PDF mRNA levels were significantly increased in breast, colon, and lung cancer samples while MAP1D mRNA levels were increased in just colon cancers. The expression of PDF and MAP1D varied with stage in these cancers. Further, PDF protein expression was elevated in colon cancer tissue samples. Inhibition of the MEK/ERK, but not PI3K or mTOR, pathway reduced the expression of PDF and MAP1D in both colon and lung cancer cell lines. Further, inhibition of PDF with actinonin resulted in greater reduction of breast, colon, and prostate cancer cell proliferation than non-cancer cell lines.ConclusionsThis is the first report showing that PDF is over-expressed in breast, colon, and lung cancers, and the first evidence that the MEK/ERK pathway plays a role in regulating the expression of PDF and MAP1D. The over-expression of PDF in several cancers and the inhibition of cancer cell growth by a PDF inhibitor suggest this enzyme may act as an oncogene to promote cancer cell proliferation.

BackgroundIn an effort to achieve better cancer therapies, we elucidated the combination cancer therapy of STI571 (an inhibitor of Bcr-Abl and clinically used for chronic myelogenous leukemia) and TNF-related apoptosis-inducing ligand (TRAIL, a developing antitumor agent) in leukemia, colon, and prostate cancer cells.MethodsColon cancer (HCT116, SW480), prostate cancer (PC3, LNCaP) and leukemia (K562) cells were treated with STI571 and TRAIL. Cell viability was determined by MTT assay and sub-G1 appearance. Protein expression and kinase phosphorylation were determined by Western blotting. c-Abl and p73 activities were inhibited by target-specific small interfering (si)RNA. In vitro kinase assay of c-Abl was conducted using CRK as a substrate.ResultsWe found that STI571 exerts opposite effects on the antitumor activity of TRAIL. It enhanced cytotoxicity in TRAIL-treated K562 leukemia cells and reduced TRAIL-induced apoptosis in HCT116 and SW480 colon cancer cells, while having no effect on PC3 and LNCaP cells. In colon and prostate cancer cells, TRAIL caused c-Abl cleavage to the active form via a caspase pathway. Interestingly, JNK and p38 MAPK inhibitors effectively blocked TRAIL-induced toxicity in the colon, but not in prostate cancer cells. Next, we found that STI571 could attenuate TRAIL-induced c-Abl, JNK and p38 activation in HCT116 cells. In addition, siRNA targeting knockdown of c-Abl and p73 also reduced TRAIL-induced cytotoxicity, rendering HCT116 cells less responsive to stress kinase activation, and masking the cytoprotective effect of STI571.ConclusionsAll together we demonstrate a novel mediator role of p73 in activating the stress kinases p38 and JNK in the classical apoptotic pathway of TRAIL. TRAIL via caspase-dependent action can sequentially activate c-Abl, p73, and stress kinases, which contribute to apoptosis in colon cancer cells. Through the inhibition of c-Abl-mediated apoptotic p73 signaling, STI571 reduces the antitumor activity of TRAIL in colon cancer cells. Our results raise additional concerns when developing combination cancer therapy with TRAIL and STI571 in the future.

Colon cancer is one of the most prevalent malignancies that lead to high occurrence of cancer-related deaths. Currently, chemotherapies and radiotherapies remain the primary treatments for advanced colon cancer. Despite the initial effectiveness, a fraction of colon cancer patients developed cisplatin resistance, resulting in therapeutic failure. The long non-coding RNA differentiation antagonizing non-coding RNA (DANCR) has been shown to be upregulated in multiple cancers, indicating an oncogenic role of DANCR. This study aims to elucidate the roles of DANCR in regulating cisplatin (CDDP) resistance of colon cancer. We found DANCR was significantly upregulated in colon cancer tissues and cells compared with normal colon tissues and cells. DANCR was upregulated in cisplatin-resistant colon cancer cells. Moreover, overexpression of DANCR significantly desensitized colon cancer cells to cisplatin. On the other way, silencing DANCR dramatically overrode CDDP resistance of colon cancer cells. Bioinformatics prediction revealed DANCR could bind to seeding region of miR-125b-5p as a competitive endogenous RNA. This interference was further validated by luciferase assay. Moreover, we detected a negative correlation between DANCR and miR-125b-5p in colon cancer patient tissues: miR-125b-5p was clearly downregulated in colon cancer tissues and cells. Overexpression of miR-125b-5p significantly sensitized cisplatin-resistant cells. Interestingly, we observed the cisplatin-resistant cells were associated with a significantly increased glycolysis rate. We further identified glycolysis enzyme, hexokinase 2 (HK2), as a direct target of miR-125b-5p in colon cancer cells. Rescue experiments showed overexpression of miR-125b-5p suppressed cellular glycolysis rate and increased cisplatin sensitivity through direct targeting the 3′ UTR of HK2. Importantly, silencing endogenous DANCR significantly induced the miR-125b-5p/HK2 axis, resulting in suppression of the glycolysis rate and increase in cisplatin sensitivity of colon cancer cell. Expectedly, these processes could be further rescued by inhibiting miR-125b-5p in the DANCR-silenced cells. Finally, we validated the DANCR-promoted cisplatin resistance via the miR-125b-5p/HK2 axis from an in vivo xenograft mice model. In summary, our study reveals a new mechanism of the DANCR-promoted cisplatin resistance, presenting the lncRNA-DANCR–miR-125b-5p/HK2 axis as a potential target for treating chemoresistant colon cancer.

Epigenetic alterations, especially histone modification, play vital roles in the pathogenesis of colon cancer. Upregulation of the enhancer of zeste homolog 2 (EZH2) has been reported to contribute to the initiation and progression of colon cancer. This study analyzed the association between EZH2 and phosphorylation of H2B at tyrosine 37 (H2BY37ph ) in colon cancer tissues and cells, along with the influences of the EZH2-H2BY37ph axis on colon cancer cell autophagy. Immunohistochemistry was utilized to assess EZH2 and H2BY37ph expressions in clinical samples of colon cancer. Cell transfection was carried out to alter EZH2 and H2BY37ph expressions in colon cancer cells. Co-immunoprecipitation analysis and glutathione-S-transferase (GST) pull down assay were conducted to analyze the association between EZH2 and H2BY37ph . Western blotting was utilized to measure proteins expressions related to cell autophagy. We found that there was a positive association between EZH2 and H2BY37ph in colon cancer tissues and cells. EZH2 directly interacted with H2B and promoted H2BY37ph in colon cancer cells using ATP as a phosphate donor. Moreover, EZH2 levated colon cancer cell autophagy in starvation condition. H2BY37ph was required for EZH2-elevated colon cancer cell autophagy under starvation condition. The EZH2-H2BY37ph axis elevated colon cancer cell autophagy possibly via activating transcriptional regulation of ATG genes. In conclusion, EZH2-elevated colon cancer initiation and progression at least in part via inducing colon cancer cell autophagy. EZH2 could phosphorylate H2BY37 and then induce transcription activation of ATG genes in colon cancer cells under starvation condition.

PurposeWe conducted the study to elucidate how LncRNA LINC00941 affects colon cancer progression and its possible regulatory mechanism.MethodsThe expression level of LINC00941 in colon cancer tissues and cells was detected by qRT-PCR. The function of LINC00941 on colon cancer cell proliferation, migration, and invasion was detected by CCK-8 and Transwell assay respectively. The target interactions among LINC00941, miR-205-5p, and MYC were further confirmed by dual-luciferase reporter gene assays and RNA pull-down experiments. Meanwhile, in vivo experiments were carried out to study the role of LINC00941 in the xenotransplantation model.ResultsLINC00941 expression level was elevated in colon cancer tissues and cells. LINC00941 overexpression accelerated proliferation, migration, and invasion of colon cancer cells, while the LINC00941 knockdown showed the opposite results. In addition, LINC00941 regulated the expression of MYC by sponging miR-205-5p as a competitive endogenous RNA, and miR-205-5p knockdown reversed the tumor inhibition of LINC00941 knockdown on colon cancer cells. Xenograft model assay confirmed that LINC00941 silencing could inhibit colon cancer cell growth and metastasis.ConclusionLINC00941 may markedly promote colon cancer progression by acting on the miR-205-5p/MYC axis as a ceRNA, which offers novel clues for lncRNA to guide the treatment and prognosis of colon cancer.