AP4 (TFAP4) encodes a basic helix-loop-helix leucine zipper (bHLH-LZ) transcription factor and is a direct target gene of the oncogenic transcription factor c-MYC. Here, we set out to determine the ...relevance of AP4 in human colorectal cancer (CRC) cells.
A CRISPR/Cas9 approach was employed to generate AP4-deficient CRC cell lines with inducible expression of c-MYC. Colony formation, β-gal staining, immunofluorescence, comet and homologous recombination (HR) assays and RNA-Seq analysis were used to determine the effects of AP4 inactivation. qPCR and qChIP analyses was performed to validate differentially expressed AP4 targets. Expression data from CRC cohorts was subjected to bioinformatics analyses. Immunohistochemistry was used to evaluate AP4 targets in vivo. Ap4-deficient APC
mice were analyzed to determine conservation. Immunofluorescence, chromosome and micronuclei enumeration, MTT and colony formation assays were used to determine the effects of AP4 inactivation and target gene regulation on chromosomal instability (CIN) and drug sensitivity.
Inactivation of AP4 in CRC cell lines resulted in increased spontaneous and c-MYC-induced DNA damage, chromosomal instability (CIN) and cellular senescence. AP4-deficient cells displayed increased expression of the long non-coding RNA MIR22HG, which encodes miR-22-3p and was directly repressed by AP4. Furthermore, Mediator of DNA damage Checkpoint 1 (MDC1), a central component of the DNA damage response and a known target of miR-22-3p, displayed decreased expression in AP4-deficient cells. Accordingly, MDC1 was directly induced by AP4 and indirectly by AP4-mediated repression of miR-22-3p. Adenomas and organoids from Ap4-deficient APC
mice displayed conservation of these regulations. Inhibition of miR-22-3p or ectopic MDC1 expression reversed the increased senescence, DNA damage, CIN and defective HR observed in AP4-deficient CRC cells. AP4-deficiency also sensitized CRC cells to 5-FU treatment, whereas ectopic AP4 conferred resistance to 5-FU in a miR-22-3p and MDC1-dependent manner.
In summary, AP4, miR-22-3p and MDC1 form a conserved and coherent, regulatory feed-forward loop to promote DNA repair, which suppresses DNA damage, senescence and CIN, and contributes to 5-FU resistance. These findings explain how elevated AP4 expression contributes to development and chemo-resistance of colorectal cancer after c-MYC activation.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Patients with estrogen receptor α (ERα)-positive breast cancer can be treated with endocrine therapy using anti-estrogens such as tamoxifen; nonetheless, patients often develop resistance limiting ...the success of breast cancer treatment. The potential mechanisms remain elusive. In detail, many miRNAs have been associated with breast cancer tamoxifen resistance, but no studies have addressed the role of miRNA-mediated competitive endogenous RNAs network (ceRNET) in tamoxifen resistance. The ceRNET between CYP4Z1 and pseudogene CYP4Z2P has been revealed to promote breast cancer angiogenesis. However, its function in tamoxifen resistance remains unclear. Here we report CYP4Z1 and CYP4Z2P were downregulated in MCF-7 cells compared with tamoxifen-resistant MCF-7-TamR cells. Enforced upregulation of CYP4Z1- or CYP4Z2P-3′UTR level renders MCF-7 Cells resistant to tamoxifen. We find that overexpression of CYP4Z1- or CYP4Z2P-3′UTR enhances the transcriptional activity of ERα through the activation of ERα phosphorylation. Furthermore, we find that CYP4Z1- and CYP4Z2P-3′UTRs increase ERα activity dependent on cyclin-dependent kinase 3 (CDK3). Reporter gene and western blot assays revealed that CYP4Z1- and CYP4Z2P-3′UTRs act as CDK3 ceRNAs. More importantly, the blocking of CYP4Z1- and CYP4Z2P-3′UTRs reversed tamoxifen resistance in MCF-7-TamR cells. Our data demonstrates that the ceRNET between CYP4Z1 and pseudogene CYP4Z2P acts as a sub-ceRNET to promote CDK3 expression in ER-positive breast cancer and is a potential therapeutic target for treatment of tamoxifen-resistant breast cancer.
•A new critical role of ceRNAs in ER + breast cancer is proposed.•We examine the relationship of CYP4Z2P, CYP4Z1 and CDK3 in breast cancer.•The roles of CYP4Z2P and CYP4Z1 in TAM resistance were further studied.•The novel mechanism provides new insights for TAM resistance.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
Competing endogenous RNAs (ceRNAs) network has been correlated with the initiation and development of cancer. Here, we identify CDH5, HOXD1, and HOXD10 as putative STARD13 ceRNAs and they display ...concordant patterns with STARD13 in different metastatic potential breast cancer cell lines and tissues. Notably, 3'UTRs of these genes suppress breast cancer metastasis via inhibiting epithelial-mesenchymal transition (EMT) in vitro and in vivo, which are activated through the crosstalk between STARD13 and its ceRNAs in 3'UTR- and miRNA-dependent manners. In addition, Kaplan-Meier survival analysis reveals that mRNA level of STARD13 and its ceRNAs is remarkably associated with survival of breast cancer patients. These results suggest that 3'UTRs of CDH5, HOXD1, and HOXD10 inhibit breast cancer metastasis via serving as STARD13 ceRNAs.
List of abbreviations 3’-UTR 3’-untranslated region AP-1 activator protein-1 Cas9 clustered regularly interspaced short palindromic repeats-associated protein 9 CDH1 E-cadherin 1 MYC c-MYC ...proto-oncogene CRC colorectal cancer CRISPR clustered regularly interspaced short palindromic repeats DOX doxycycline EMT epithelial-mesenchymal transition FOSL1/FRA1 FOS-like 1/FOS-related antigen 1 JNK1 c-Jun N-terminal protein kinase 1 JUN c-Jun proto-oncogene JUNB c-JunB proto-oncogene KO knock-out MAP2K7 mitogen-activated protein kinase kinase 7 MAP3K13 mitogen-activated protein kinase kinase kinase 13 MAPK mitogen-activated protein kinase mCRC metastatic colorectal cancer MDC1 mediator of DNA damage checkpoint 1 MIR22HG MIR22 host gene MUT mutant NOD/SCID non-obese diabetic/severe combined immunodeficiency qChIP quantitative real-time polymerase chain reaction-chromatin immunoprecipitation qPCR quantitative real-time polymerase chain reaction SMS seed-matching sequence t1/2 half-life time TCGA-COAD The Cancer Genome Atlas Colorectal Adenocarcinoma TF transcription factor TFAP4/AP4 transcription factor AP4/activating enhancer binding protein 4 VIM vimentin WT wild-type ΔSMS seed-matching sequence deletion Dear Editor, Colorectal cancer (CRC) is the third most deadly cancer worldwide 1. Furthermore, AP4 occupancy at the MAP3K13 and FOSL1 promoters was confirmed by quantitative real-time polymerase chain reaction-chromatin immunoprecipitation (qChIP) analysis (Figure 1C). For Western blot analysis, β-actin served as a loading control. (K) qPCR analysis of the FOSL1 mRNA decay rate in MIR22-deficient DLD-1 cells and FOSL1 ΔSMS DLD-1 cells after treatment with 10 μg/mL of Actinomycin D for the indicated durations to block gene transcription.
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FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Competitive endogenous messenger RNAs (ceRNAs) affect other RNAs transcription through competitively binding common microRNAs (miRNAs). In this study we identified long non-coding RNA (lncRNA) MALAT1 ...can function as a ceRNA of cell division cycle 42 (cdc42) 3′UTR in inducing migration and invasion of breast cancer cells via miR-1. We found that miR-1 bound both MALAT1 and cdc42 3′UTR directly. Further study showed that MALAT1 induced migration and invasion of breast cancer cells while reduced the level of cdc42. Our results suggest that MALAT1 regulated migration and invasion of breast cancer cells via affecting cdc42 through binding miR-1 competitively.
•Verifying MALAT1 does promote migration and invasion of breast cancer cells.•MiR-1 plays an important role in MALAT1 and cdc42 regulating network.•MALAT1 competitively binds miR-1 with cdc42 to promote migration and invasion of breast cancer cells.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
To investigate the effects of CYP4Z1 3′UTR in migration of breast cancer cells, a series of assays were used to confirm that overexpression of CYP4Z1 3′UTR could suppress the capacity of migration ...and adhesion of MCF-7 and MDA-MB-231 cells. EMT (Epithelial-mesenchymal transition)-related proteins were regulated by CYP4Z1 3′UTR. Mesenchyma markers like Vimentin, MMP-2, and MMP-9 were down-regulated, while the expression of E-cadherin was up-regulated with CYP4Z1 3′UTR overexpression. Notably, luciferase reporter and qRT-PCR assays were applied to verify that CYP4Z1 3′UTR was the potential target of miR-9. In addition, our results showed that CYP4Z1 3′UTR repressed the expression of E-cadherin in a miRNA-dependent manner. Combining with our previous study, we have discovered the underlying link between CYP4Z1 and E-cadherin. Therefore, those preliminary data suggest that CYP4Z1 3′UTR could inhibit the migration and EMT of breast cancer cells via acting as a ceRNA for E-cadherin.
•A new critical role of CYP4Z1-3′UTR in breast cancer is proposed.•We examine the relationship of CYP4Z1, E-cadherin and miR-9 in breast cancer.•The mechanism which CYP4Z1 3′UTR influenced EMT were further studied.•We firstly showed that CYP4Z1 3′UTR repressed the expression of E-cadherin in miRNA-dependent manner.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
Competitive endogenous messenger RNA (ceRNA) affects transcription of other RNA molecules by competitively binding common microRNAs. Previous studies have shown that TP53INP1 functions as a ...suppressor in tumor metastasis. Our study elucidated StarD13 messenger RNA as a ceRNA in regulating migration and invasion of breast cancer cells. MicroRNA-125b was identified to induce metastasis of MCF-7 cells and bind with both StarD13 3′UTR and TP53INP1 3′UTR. Therefore, a ceRNA interaction between StarD13 and TP53INP1 mediated by competitively binding to miR-125b was indicated. Importantly, a microRNA-125b binding site at 4546–4560 nt on StarD13 was verified more vital for this ceRNA interaction. Indirectly regulation of SPARC in inducing metastasis of breast cancer cells by StarD13 via competitively binding with TP53INP1 was further confirmed. In conclusion, our findings demonstrate a ceRNA regulatory network which could give a better understanding of metastatic mechanisms of breast cancer.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
CXCR4 is the most common chemokine receptor expressed on tumor cells, and it is closely correlated with cancer cell stemness. This study was carried out to explore whether CXCR4 could function as a ...competitive endogenous RNA to promote metastasis, proliferation and survival in MCF-7 breast cancer cells. We validated that CXCR4, together with TRAF6 and EGFR, was directly targeted by miR-146a in MCF-7 cells. Overexpression of CXCR4 3′UTR inhibited the activity of miR-146a, thus elevating the expression of CXCR4, TRAF6 and EGFR. These oncoproteins further activated NF-κB pathway and promoted the proliferation, migration, invasion and anti-apoptotic activity of MCF-7 cells. Collectively, our study provided new insights into the function of CXCR4 in breast cancer: it promotes tumor progression as both a protein-coding gene and a non-coding RNA, complicating the mechanism by which oncogenes promote tumor progression.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
CXCR4 is the most common chemokine receptor expressed on tumor cells, and it is closely correlated with cancer cell stemness. This study was carried out to explore whether CXCR4 could function as a ...competitive endogenous RNA to promote metastasis, proliferation and survival in MCF-7 breast cancer cells. We validated that CXCR4, together with TRAF6 and EGFR, was directly targeted by miR-146a in MCF-7 cells. Overexpression of CXCR4 3'UTR inhibited the activity of miR-146a, thus elevating the expression of CXCR4, TRAF6 and EGFR. These oncoproteins further activated NF-κB pathway and promoted the proliferation, migration, invasion and anti-apoptotic activity of MCF-7 cells. Collectively, our study provided new insights into the function of CXCR4 in breast cancer: it promotes tumor progression as both a protein-coding gene and a non-coding RNA, complicating the mechanism by which oncogenes promote tumor progression.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
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