LncRNA TUG1 promotes pulmonary fibrosis progression via up-regulating CDC27 and activating PI3K/Akt/mTOR pathway.

Idiopathic pulmonary fibrosis (IPF) is a fatal interstitial lung disease with an unclear pathogenesis. This study aimed to elucidate the function and potential mechanisms of TUG1 in IPF progression. Cell viability and migration were detected by CCK-8 and transwell assays. Autophagy, fibrosis, or EMT-related proteins were measured by Western blotting. Pro-inflammatory cytokine levels were assessed by ELISA kits. The subcellular localization of TUG1 was observed by FISH assay. RIP assay detected the interaction between TUG1 and CDC27. TUG1 and CDC27 was up-regulated in TGF-ß1-induced RLE-6TN cells. TUG1 depletion suppressed pulmonary fibrosis via attenuating inflammation, EMT, inducing autophagy and inactivating PI3K/Akt/mTOR pathway in vitro and in vivo. TUG1 knockdown prevented CDC27 expression. TUG1 silencing ameliorated pulmonary fibrosis by reducing CDC27 expression and inhibiting PI3K/Akt/mTOR pathway.

Circ_0000775 promotes the migration, invasion and EMT of hepatic carcinoma cells by recruiting IGF2BP2 to stabilize CDC27.

BACKGROUND: Hepatic carcinoma (HC) is one of the leading causes of cancer-related death, and the incidence keeps high in the world. The vital role of circular RNAs (circRNAs) in HC development has been revealed to some extent. Circ_0000775, a novel circRNA, has never been thoroughly studied regarding HC. METHODS: Online datasets were utilized to obtain expression pattern of genes in tumor tissues. RT-qPCR and western blot examined the RNA and protein levels of indicated genes. ChIP, DNA pull down, RNA pull down, RIP and luciferase reporter assays were carried out to verify correlation between different factors. Supported by RT-qPCR and western blot analyses, transwell and wound healing assay were implemented for detecting cell migration and invasion and EMT. Additionally, cell EMT was also evaluated via cell morphology observation for calculation of spindle cell number. RESULTS: High expression of circ_0000775 in HC cells was induced by transcriptionally stimulation by TCF7L2. Circ_0000775 in cytoplasm recruited IGF2BP2 to enhance the mRNA stability of CDC27, thus positively modulating CDC27 expression. Circ_0000775 exacerbated HC cell migration, invasion and EMT through CDC27. CONCLUSION: TCF7L2 promoted the transcription of circ_0000775, and circ_0000775 recruited IGF2BP2 to maintain CDC27 mRNA stability, thereby facilitating HC cell migration, invasion and EMT.

Circular RNA circSLC8A1 inhibits the proliferation and invasion of glioma cells through targeting the miR-214-5p/CDC27 axis.

Circular RNA circSLC8A1 is one of the cancer-related circRNAs that is implicated in various cancers. However, studies focusing on the role of circSLC8A1 in glioma is rare. Here we attempted to evaluate the biological function of circSLC8A1 in glioma and explore the potential mechanism. The relative expression of circSLC8A1, miR-214-5p and CDC27 in tissues and cell lines was determined by qRT-PCR. Cell proliferation and invasion were respectively measured by CCK-8 and transwell assays. Protein level of CDC27 was analyzed by western blot. Luciferase reporter assay was performed to confirm the regulatory interaction of cirRNA-miRNA-mRNA. Lowly expressed circSLC8A1 was observed in both glioma tissues and cell lines. Further biological analyses showed that circSLC8A1 inhibits the cell proliferation and invasion of glioma cells. CircSLC8A1 directly sponged miR-214-5p and inhibited miR-214-5p expression in glioma cells. CDC27 was a direct target of miR-214-5p and could be regulated by miR-214-5p. Moreover, miR-214-5p mimics and CDC27 knockdown reversed the inhibitory effects of circSLC8A1 on cell proliferation and invasion. Taken together, our results demonstrated a tumor suppressive role of circSLC8A1 in glioma through regulation of glioma cells proliferation and invasion. The effects of circSLC8A1 were mediated by miR-214-5p/CDC27 axis. Our study provided a new understanding of the occurrence and development of glioma.

A novel long non-coding RNA RP11-286H15.1 represses hepatocellular carcinoma progression by promoting ubiquitination of PABPC4.

Hepatocellular carcinoma (HCC) is a malignancy found at high frequency around the world. Unfortunately, the scarcity of effective early diagnostic methods invariably results in poor outcomes. Long noncoding RNAs (lncRNAs) are known to regulate the progression of hepatocellular carcinoma (HCC). A novel lncRNA RP11-286H15.1(OTTHUMG00000186042) has been identified and associated with HCC; however, the potential role of RP11-286H15.1 in HCC remains undefined. The transcript abundance of RP11-286H15.1 in 80 pairs of HCC samples and cell lines was evaluated by qRT-PCR analysis. The functional role of RP11-286H15.1 in HCC was tested in vivo and in vitro. The mechanisms underlying the role of RP11-286H15.1 in HCC were explored by RNA pulldown, transcriptome sequencing, and RNA immunoprecipitation (RIP), ubiquitination and fluorescence in situ hybridization (FISH) assays as well as Western blot analysis. The qRT-PCR and FISH assays revealed that RP11-286H15.1 was significantly decreased in HCC, and implied a shorter survival time. RP11-286H15.1 overexpression inhibited HCC cell proliferation and metastasis in vitro and in vivo, whereas RP11-286H15.1 knockdown produced the opposite results. Furthermore, we confirmed that RP11-286H15.1 (620-750 nucleotides) binds to poly(A) binding protein 4 (PABPC4) and promotes its ubiquitination, thus, reducing the stability of TRIM37 and CDC27 mRNAs. Our study demonstrates that a novel lncRNA, RP11-286H15.1, represses HCC progression by promoting PABPC4 ubiquitination. These findings highlight potential therapeutic targets for HCC.

CDC27 Facilitates Gastric Cancer Cell Proliferation, Invasion and Metastasis via Twist-Induced Epithelial-Mesenchymal Transition.

BACKGROUND/AIMS: Lymph node metastasis is the primary cause of cancer-related death among patients with gastric cancer (GC), and cell division cycle 27 (CDC27) promotes the metastasis and epithelial-mesenchymal transition in many cancers. Till now, the mechanisms underlying CDC27-induced the epithelial-mesenchymal transition (EMT) of GC are still unclear. METHODS: We analyzed the expression levels of CDC27 and EMT-related biomarkers using immunohistochemistry and Western blot in 60 cases of GC tissues, and then GC cells with CDC27 shRNAs or plasmids were subjected to in vitro and in vivo assays, including CCK-8, wound healing and transwell assays. RESULTS: The CDC27 expression was obviously increased in GC tissues, and significantly correlates with EMT-related biomarkers, lymph node metastasis and poor 5-year overall survival. Additionally, in vitro and in vivo assays demonstrated that silencing of CDC27 expression effectively inhibited GC cell proliferation, invasion and metastasis. Conversely, CDC27 overexpression led to the opposite results. Finally, we demonstrated that Twist shRNA inhibited CDC27-meditated invasion and EMT of GC cells. CONCLUSION: CDC27 facilitates gastric cancer cell proliferation, invasion and metastasis via Twist-induced EMT; thus, this study offered a new therapy method for GC patients.

Germline and somatic variations influence the somatic mutational signatures of esophageal squamous cell carcinomas in a Chinese population.

BACKGROUND: Esophageal squamous cell carcinomas (ESCC) is the fourth most lethal cancer in China. Previous studies reveal several highly conserved mutational processes in ESCC. However, it remains unclear what are the true regulators of the mutational processes. RESULTS: We analyzed the somatic mutational signatures in 302 paired whole-exome sequencing data of ESCC in a Chinese population for potential regulators of the mutational processes. We identified three conserved subtypes based on the mutational signatures with significantly different clinical outcomes. Our results show that patients of different subpopulations of Chinese differ significantly in the activity of the "NpCpG" signature (FDR = 0.00188). In addition, we report ZNF750 and CDC27, of which the somatic statuses and the genetic burdens consistently influence the activities of specific mutational signatures in ESCC: the somatic ZNF750 status is associated with the AID/APOBEC-related mutational process (FDR = 0.0637); the somatic CDC27 copy-number is associated with the "NpCpG" (FDR = 0.00615) and the AID/APOBEC-related mutational processes (FDR = 8.69 × 10- 4). The burdens of germline variants in the two genes also significantly influence the activities of the same somatic mutational signatures (FDR < 0.1). CONCLUSIONS: We report multiple factors that influence the mutational processes in ESCC including: the subpopulations of Chinese; the germline and somatic statuses of ZNF750 and CDC27 and exposure to alcohol and tobacco. Our findings based on the evidences from both germline and somatic levels reveal potential genetic regulators of the somatic mutational processes and provide insights into the biology of esophageal carcinogenesis.

UBE2S associated with OSCC proliferation by promotion of P21 degradation via the ubiquitin-proteasome system.

Ubiquitin-conjugating enzyme E2S (UBE2S), a family of E2 protein in the ubiquitin-proteasome system, is highly expressed in several types of cancers; however, its roles in oral squamous cell carcinoma (OSCC) have not yet been well elucidated. The purpose of this study was to clarify the functional activities of UBE2S in OSCCs. We analyzed the expression levels of UBE2S in nine OSCC cell lines and primary OSCC tissues by quantitative reverse transcriptase-polymerase chain reaction, Western blotting, and immunohistochemistry (IHC). The correlations between UBE2S expression and clinical classifications of OSCCs were analyzed using the IHC scoring system. We also used UBE2S knockdown OSCC cells for functional assays (proliferation assay, flow cytometry, and Western blotting). UBE2S was overexpressed in OSCCs in vitro and in vivo and was correlated significantly (P < 0.05) with the primary tumoral size. The cellular growth was decreased and the cell-cycle was arrested in the G2/M phase in the UBE2S knockdown (shUBE2S) cells. The expression level of P21, a target of the ubiquitin-proteasome system, was increased in the shUBE2S cells because of lower anaphase activity that promotes complex subunit 3 (APC3), an E3 ubiquitin ligase, compared with shMock cells. These findings might promote the understanding of the relationship between UBE2S overexpression and oral cancer proliferation, indicating that UBE2S would be a potential biomarker of and therapeutic target in OSCCs.

APC/C Dysfunction Limits Excessive Cancer Chromosomal Instability.

Intercellular heterogeneity, exacerbated by chromosomal instability (CIN), fosters tumor heterogeneity and drug resistance. However, extreme CIN correlates with improved cancer outcome, suggesting that karyotypic diversity required to adapt to selection pressures might be balanced in tumors against the risk of excessive instability. Here, we used a functional genomics screen, genome editing, and pharmacologic approaches to identify CIN-survival factors in diploid cells. We find partial anaphase-promoting complex/cyclosome (APC/C) dysfunction lengthens mitosis, suppresses pharmacologically induced chromosome segregation errors, and reduces naturally occurring lagging chromosomes in cancer cell lines or following tetraploidization. APC/C impairment caused adaptation to MPS1 inhibitors, revealing a likely resistance mechanism to therapies targeting the spindle assembly checkpoint. Finally, CRISPR-mediated introduction of cancer somatic mutations in the APC/C subunit cancer driver gene CDC27 reduces chromosome segregation errors, whereas reversal of an APC/C subunit nonsense mutation increases CIN. Subtle variations in mitotic duration, determined by APC/C activity, influence the extent of CIN, allowing cancer cells to dynamically optimize fitness during tumor evolution. SIGNIFICANCE: We report a mechanism whereby cancers balance the evolutionary advantages associated with CIN against the fitness costs caused by excessive genome instability, providing insight into the consequence of CDC27 APC/C subunit driver mutations in cancer. Lengthening of mitosis through APC/C modulation may be a common mechanism of resistance to cancer therapeutics that increase chromosome segregation errors. Cancer Discov; 7(2); 218-33. ©2017 AACR.See related commentary by Burkard and Weaver, p. 134This article is highlighted in the In This Issue feature, p. 115.

mir-218-2 promotes glioblastomas growth, invasion and drug resistance by targeting CDC27.

Glioma has become a significant global health problem with substantial morbidity and mortality, underscoring the importance of elucidating its underlying molecular mechanisms. Recent studies have identified mir-218 as an anti-oncogene; however, the specific functions of mir-218-1 and mir-218-2 remain unknown, especially the latter. The objective of this study was to further investigate the role of mir-218-2 in glioma. Our results indicated that mir-218-2 is highly overexpressed in glioma. Furthermore, we showed that mir-218-2 is positively correlated with the growth, invasion, migration, and drug susceptibility (to ß-lapachone) of glioma cells. In vitro, the overexpression of mir-218-2 promoted glioma cell proliferation, invasion, and migration. In addition, the overexpression of mir-218-2 in vivo was found to increase glioma tumor growth. Accordingly, the inhibition of mir-218-2 resulted in the opposite effects. Cell division cycle 27 (CDC27), the downstream target of mir-218-2, is involved in the regulation of glioma cells. Our results indicate that the overexpression of CDC27 counteracted the function of mir-218-2 in glioma cells. These novel findings provide new insight in the application of mir-218-2 as a potential glioma treatment.

Inhibition of Anaphase-Promoting Complex by Silence APC/CCdh1 to Enhance Radiosensitivity of Nasopharyngeal Carcinoma Cells.

The aim of this study was to investigate the possibility of APC/CCdh1 as a potential therapeutic target in the radiosensitivity of nasopharyngeal carcinoma (NPC) cell CNE-1, and explain the role of APC subunits after silence of Cdh1 combined with radiotherapy. Transfection with Cdh1 shRNA significantly increased the radiosensitivity of CNE-1 cells and the radiation enhancement ratio (RER) of sh-Cdh1 cells was 1.76. Knockdown of Cdh1 in CNE-1 cells increased irradiation induced apoptosis and G2/M phase cell cycle arrest. The levels of CDC20 and CylinB1 increased and the levels of Ku70 and APC3 decreased after irradiation. APC/CCdh1 is involved in regulation of radiosensitivity in human NPC CNE-1 cells. Our study may provide a promising therapeutic strategy for NPC by targeting Cdh1. J. Cell. Biochem. 118: 3150-3157, 2017. © 2016 Wiley Periodicals, Inc.

PCBP1/HNRNP E1 Protects Chromosomal Integrity by Translational Regulation of CDC27.

UNLABELLED: CDC27 is a core component of the anaphase-promoting complex/cyclosome (APC/C), a multisubunit E3 ubiquitin ligase, whose oscillatory activity is responsible for the metaphase-to-anaphase transition and mitotic exit. Here, in normal murine mammary gland epithelial cells (NMuMG), CDC27 expression is controlled posttranscriptionally through the RNA binding protein poly(rC) binding protein 1 (PCBP1)/heterogeneous nuclear ribonucleoprotein E1 (HNRNP E1). shRNA-mediated knockdown of HNRNP E1 abrogates translational silencing of the Cdc27 transcript, resulting in constitutive expression of CDC27. Dysregulated expression of CDC27 leads to premature activation of the G2-M-APC/C-CDC20 complex, resulting in the aberrant degradation of FZR1/CDH1, a cofactor of the G1 and late G2-M-APC/C and a substrate normally reserved for the SCF-ßTRCP ligase. Loss of CDH1 expression and of APC/C-CDH1 activity, upon constitutive expression of CDC27, results in mitotic aberrations and aneuploidy in NMuMG cells. Furthermore, tissue microarray of breast cancer patient tumor samples reveals high CDC27 levels compared with nonneoplastic breast tissue and a significant correlation between disease recurrence and CDC27 expression. These results suggest that dysregulation of HNRNP E1-mediated translational regulation of Cdc27 leads to chromosomal instability and aneuploidy and that CDC27 expression represents a significant predictor of breast cancer recurrence. IMPLICATIONS: The RNA-binding protein HNRNP E1 mediates translational regulation of the cell-cycle regulator CDC27 and that dysregulation of CDC27 leads to aneuploidy. In addition, high CDC27 expression in breast cancer patient tumor specimens significantly predicts disease recurrence, suggesting a novel role for CDC27 as a predictor of relapse. Mol Cancer Res; 14(7); 634-46. ©2016 AACR.

Downregulation of CDC27 inhibits the proliferation of colorectal cancer cells via the accumulation of p21Cip1/Waf1.

Dysregulated cell cycle progression has a critical role in tumorigenesis. Cell division cycle 27 (CDC27) is a core subunit of the anaphase-promoting complex/cyclosome, although the specific role of CDC27 in cancer remains unknown. In our study, we explored the biological and clinical significance of CDC27 in colorectal cancer (CRC) growth and progression and investigated the underlying molecular mechanisms. Results showed that CDC27 expression is significantly correlated with tumor progression and poor patient survival. Functional assays demonstrated that overexpression of CDC27 promoted proliferation in DLD1 cells, whereas knockdown of CDC27 in HCT116 cells inhibited proliferation both in vitro and in vivo. Further mechanistic investigation showed that CDC27 downregulation resulted in G1/S phase transition arrest via the significant accumulation of p21 in HCT116 cells, and the upregulation of CDC27 promoted G1/S phase transition via the attenuation of p21 in DLD1 cells. Furthermore, we also demonstrated that CDC27 regulated inhibitor of DNA binding 1 (ID1) protein expression in DLD1 and HCT116 cells, and rescue assays revealed that CDC27 regulated p21 expression through modulating ID1 expression. Taken together, our results indicate that CDC27 contributes to CRC cell proliferation via the modulation of ID1-mediated p21 regulation, which offers a novel approach to the inhibition of tumor growth. Indeed, these findings provide new perspectives for the future study of CDC27 as a target for CRC treatment.

MiR-27a modulates radiosensitivity of triple-negative breast cancer (TNBC) cells by targeting CDC27.

BACKGROUND: MiR-27a is significantly overexpressed in triple-negative breast cancer (TNBC). However, the exact biological function of MiR-27a in TNBC is not fully understood. In this study, we verified miR-27a expression in TNBC cells and explored how its overexpression modulates radiosensitivity of the cells. MATERIAL/METHODS: qRT-PCR analysis was performed to study miR-27a expression in TNBC lines MDA-MB-435 and MDA-MB-231 and in normal human breast epithelial cell line MCF10A. Dual luciferase assay was performed to verify a putative downstream target of miR-27a, CDC27. CCK-8 assay was used to assess the influence of miR-27a-CDC27 axis on cell proliferation under irradiation (IR) treatment. RESULTS: We confirmed significantly higher miR-27a expression in 2 TNBC cell lines--MDA-MB-435 and MDA-MB-231--than in human breast epithelial cell line MCF10A. miR-27a could modulate proliferation and radiosensitivity of TNBC cells. CDC-27 is a direct target of miR-27a and its downregulation conferred increased radioresistance of the cells. CONCLUSIONS: The miR-27a-CDC27 axis might play an important role in modulating response to radiotherapy in TNBC cells. Testing miR-27a expression might be a useful way to identify a subgroup of patients who will benefit from an IR-based therapeutic approach.

Association between polymorphisms in cdc27 and breast cancer in a Chinese population.

Cdc27, as a core component of anaphase-promoting complex (APC), is a cell cycle regulator, which participates in control of mitotic checkpoint and surveys the mitotic spindle to maintain chromosomal integrity. It was hypothesized that polymorphisms in cdc27 gene might contribute to the susceptibility of breast cancer (BC) through influencing the mitotic progression of cells. Therefore, a hospital-based case-control study with 463 BC patients and 536 controls was implemented to investigate the association of six single-nucleotide polymorphisms (SNPs) in cdc27 and BC risk in a Chinese Han population. Among the six SNPs, two SNPs of rs11570443 and rs12601027 were positively correlated with BC risk. Individuals carrying rs11570443-CT or CC genotypes showed a higher BC risk with the OR of 1.75 (95 % confidence interval (CI) = 1.13-1.69), compared with those carrying rs11570443-TT genotype. For rs12601027, an increased BC risk was significantly associated with homozygote TT genotype (odds ratio (OR) = 1.49, 95 % CI = 1.12-1.98) compared with homozygote CC and heterozygote CT genotypes. In addition, a significant interaction effect of these two SNPs was found. The rs12601027-TT in combination with rs11570443-CT/CC genotypes showed a strongly elevated risk of BC compared with rs12601027-CC/CT and rs11570443-TT genotype (OR = 1.95, 95 % CI = 1.06-3.59). These findings suggested that polymorphisms in cdc27 may contribute to the susceptibility of BC though functional studies are needed to further elucidate the underling mechanisms of the associations.

Whole exome sequencing of a single osteosarcoma case--integrative analysis with whole transcriptome RNA-seq data.

BACKGROUND: Osteosarcoma (OS) is a prevalent primary malignant bone tumour with unknown etiology. These highly metastasizing tumours are among the most frequent causes of cancer-related deaths. Thus, there is an urgent need for different markers, and with our study, we were aiming towards finding novel biomarkers for OS. METHODS: For that, we analysed the whole exome of the tumorous and non-tumour bone tissue from the same patient with OS applying next-generation sequencing. For data analysis, we used several softwares and combined the exome data with RNA-seq data from our previous study. RESULTS: In the tumour exome, we found wide genomic rearrangements, which should qualify as chromotripsis-we detected almost 3,000 somatic single nucleotide variants (SNVs) and small indels and more than 2,000 copy number variants (CNVs) in different chromosomes. Furthermore, the somatic changes seem to be associated to bone tumours, whereas germline mutations to cancer in general. We confirmed the previous findings that the most significant pathway involved in OS pathogenesis is probably the WNT/ß-catenin signalling pathway. Also, the IGF1/IGF2 and IGF1R homodimer signalling and TP53 (including downstream tumour suppressor gene EI24) pathways may have a role. Additionally, the mucin family genes, especially MUC4 and cell cycle controlling gene CDC27 may be considered as potential biomarkers for OS. CONCLUSIONS: The genes, in which the mutations were detected, may be considered as targets for finding biomarkers for OS. As the study is based on a single case and only DNA and RNA analysis, further confirmative studies are required.

Discovery of biclonal origin and a novel oncogene SLC12A5 in colon cancer by single-cell sequencing.

Single-cell sequencing is a powerful tool for delineating clonal relationship and identifying key driver genes for personalized cancer management. Here we performed single-cell sequencing analysis of a case of colon cancer. Population genetics analyses identified two independent clones in tumor cell population. The major tumor clone harbored APC and TP53 mutations as early oncogenic events, whereas the minor clone contained preponderant CDC27 and PABPC1 mutations. The absence of APC and TP53 mutations in the minor clone supports that these two clones were derived from two cellular origins. Examination of somatic mutation allele frequency spectra of additional 21 whole-tissue exome-sequenced cases revealed the heterogeneity of clonal origins in colon cancer. Next, we identified a mutated gene SLC12A5 that showed a high frequency of mutation at the single-cell level but exhibited low prevalence at the population level. Functional characterization of mutant SLC12A5 revealed its potential oncogenic effect in colon cancer. Our study provides the first exome-wide evidence at single-cell level supporting that colon cancer could be of a biclonal origin, and suggests that low-prevalence mutations in a cohort may also play important protumorigenic roles at the individual level.