MicroRNA-143 (miR-143) regulates cancer glycolysis via targeting hexokinase 2 gene

High glycolysis, well known as "Warburg effect," is frequently observed in a variety of cancers. Whether the deregulation of miRNAs contributes to the Warburg effect remains largely unknown. Because miRNA regulates gene expression at both mRNA and protein levels, we constructed a gene functional association network, which allows us to detect the gene activity instead of gene expression, to integratively analyze the microarray data for gene expression and miRNA expression profiling and identify glycolysis-related gene-miRNA pairs deregulated in cancer. Hexokinase 2 (HK2), coding for the first rate-limiting enzyme of glycolysis, is among the top list of genes predicted and potentially regulated by multiple miRNAs including miR-143. Interestingly, miR-143 expression was inversely associated with HK2 protein level but not mRNA level in human lung cancer samples. miR-143, down-regulated by mammalian target of rapamycin activation, reduces glucose metabolism and inhibits cancer cell proliferation and tumor formation through targeting HK2. Collectively, we have not only established a novel methodology for gene-miRNA pair prediction but also identified miR-143 as an essential regulator of cancer glycolysis via targeting HK2.     

miR-19a-3p在前列腺癌细胞侵袭和迁移中的作用

miR-19a-3p通过多种机制调节癌细胞的增殖和转移,然而,miR-19a-3p在前列腺癌转移中的生物学作用和机制尚不清楚。本研究旨在考察mir-19a-3p在前列腺癌中的表达情况,及其对细胞侵袭和迁移的影响。本研究发现,miR-19a-3p在骨转移性前列腺癌组织和前列腺癌细胞中明显下调。上调miR-19a-3p可显著抑制前列腺癌细胞的侵袭和迁移。然而,下调miR-19a-3p则会逆转上述变化。此外,本研究还发现miR-19a-3p通过靶向TGF-β信号传导的下游效应物SMAD2来抑制前列腺癌细胞中TGF-β信号传导的活性,从而抑制前列腺癌细胞的侵袭和迁移。因此,本研究表明mir-19a-3p与前列腺癌的发生发展密切相关,mir-19a-3p有望成为前列腺癌的新治疗靶点及生物标志物。     

Targeting Notch signalling by the conserved miR-8/200 microRNA family in development and cancer cells

Notch signalling is crucial for the correct development and growth of numerous organs and tissues, and when subverted it can cause cancer. Loss of miR-8/200 microRNAs (miRNAs) is commonly observed in advanced tumours and correlates with their invasion and acquisition of stem-like properties. Here, we show that this miRNA family controls Notch signalling activation in Drosophila and human cells. In an overexpression screen, we identified the Drosophila miR-8 as a potent inhibitor of Notch-induced overgrowth and tumour metastasis. Gain and loss of mir-8 provoked developmental defects reminiscent of impaired Notch signalling and we demonstrated that miR-8 directly inhibits Notch ligand Serrate. Likewise, miR-200c and miR-141 directly inhibited JAGGED1, impeding proliferation of human metastatic prostate cancer cells. It has been suggested that JAGGED1 may also be important for metastases. Although in metastatic cancer cells, JAGGED1 modestly regulated ZEB1, the miR-200c's target in invasion, studies in Drosophila revealed that only concurrent overexpression of Notch and Zfh1/ZEB1 induced tumour metastases. Together, these data define a new way to attenuate or boost Notch signalling that may have clinical interest.     

miR-625-5p/PKM2 negatively regulates melanoma glycolysis state

PKM2 plays an important role in cancer glycolysis, however, the link of PKM2 and microRNAs (miRNAs) in melanoma is still unclear. The study will investigate the role of miRNAs in regulating PKM2 mediated melanoma cell glycolysis. We found that high PKM2 expression in melanoma tissues and cell lines was positively associated with glycolysis. Further study indicated that miR-625-5p regulated PKM2 expression on mRNA and protein levels in melanoma cells. There was a negative relationship between miR-625-5p and PKM2 expression in the clinical melanoma samples. These findings provide an evidence that miR-625-5p/PKM2 plays a role in melanoma cell glucose metabolism.     

Inhibition of miR-155 in MCF-7 breast cancer cell line by gold nanoparticles functionalized with antagomir and AS1411 aptamer

MicroRNAs are key factors for many biological functions. These regulatory molecules affect various gene networks and involve the subsequent signaling pathways. Therefore, disrupting the expression of these molecules is associated with multiple anomalies in the cells and body. One of the most important related abnormalities is the incidence of cancer. Thus, targeting microRNAs (miRNAs) is an effective approach for cancer gene therapy. Various factors are used for this purpose, including the antagomir nucleotide structure. There are some obstacles in the delivery of nucleotide therapeutics to the target cells, however, the use of nanoparticles could partly overcome these defeciencies. On the other hand, targeted delivery of antagomirs using aptamers, reduces nonspecific effects on nontarget cells. Considering the above, in this study, we designed and fabricated a nanocarrier composed of gold nanoparticles (GNPs), antagomir-155, and nucleolin specific aptamer for breast cancer study and therapy. Here, GNPs were synthesized using citrate reduction and were modified by polyA sequences, AS1411 aptamer, and antagomir-155. Attachment of molecules were confirmed using gel electrophoresis, atomic force microscopy imaging and electrochemical test. The specific entry of modified nanoparticles was investigated by fluorescence microscopy. The efficacy of modified nanoparticles was evaluated using a quantitative polymerase chain reaction (q-PCR) for miR-155 and its target gene. Efficient and specific delivery of AuNP–Apt–anti-miR-155 to target cells was confirmed in comparison with the control cell. The q-PCR analysis showed not only a significant decrease in mir-155 levels but also an elevated TP53INP1 mRNA, direct target of miR-155. The proposed structure inhibits proliferation and stimulates apoptosis by increasing the expression of TP53INP1. Our results suggest that AuNP–Apt–anti-miR-155 could be a promising nano constructor for breast cancer treatment.     

The negative feedback between miR-143 and DNMT3A regulates cisplatin resistance in ovarian cancer

Emerging evidence suggests that miR-143 plays an important role in the regulation of tumor sensitivity to chemotherapeutic agents. The study explores the underlying mechanism of miR-143 in reversing cisplatin resistance in ovarian cancer. The cisplatin-resistant ovarian cancer cell line A2780/CDDP was induced and established via treating A2780 cells by gradually increasing cisplatin concentrations. The IC₅₀ values of A2780/CDDP and A2780 to cisplatin were 218.10 ± 1.12 and 21.99 ± 1.12 μM, respectively. Quantitative real-time polymerase chain reaction (qRT-PCR) results showed that miR-143 was significantly decreased in A2780/CDDP cells compared with A2780 cells. miR-143 overexpression decreased cisplatin resistance in A2780/CDDP, and miR-143 inhibition decreased A2780 sensitivity to cisplatin. Results of qRT-PCR, Western blot analysis, and luciferase reporter assay indicated that the direct target of miR-143 was DNMT3A, which, in turn, was upregulated in A2780/CDDP. DNMT3A overexpression antagonized the sensitizing effect of miR-143 on A2780/CDDP to cisplatin. Knocking down of DNMT3A reduced cisplatin resistance in A2780/CDDP, while overexpression of DNMT3A increased cisplatin resistance in A2780. Methylation-specific polymerase chain reaction results showed that the methylation level in the promoter region of the miR-143 precursor gene was higher in A2780/CDDP cells than in A2780 cells. DNMT3A mediated the hypermethylation of the miR-143 precursor gene, resulting in miR-143 downregulation in A2780/CDDP. miR-143 inhibited cell growth of A2780/CDDP cell in nude mice. Our findings indicated the negative feedback between miR-143 and DNMT3A as a crucial epigenetic modifier of cisplatin resistance in ovarian cancer.     

miR-34基因家族的分子进化

根据miRNA基因在进化中高度保守的特点,利用生物信息学方法在目前已测序的动物物种中搜寻参与哺乳动物早期发育调控的mir-34基因的同源序列,在33个不同的动物物种中获得了miR-34基因的54条同源序列,其中18条为新发现的序列。表明miR-34是高度保守的,广泛存在于后生动物中。目前发现的mir-34基因80%位于基因间隔区,少数位于蛋白编码基因的内含子区和3′UTR上。不同动物中,mir-34基因成熟序列的同源性为68%,前体序列为38.89%。在无脊椎动物中只有一个mir-34,而在几乎所有的脊椎动物中都有mir-34a,mir-34b,mir-34c,形成miR-34基因家族。系统进化分析表明,脊椎动物中miR-34基因家族是通过基因的串联和局部重复形成的,这个过程中伴随着个别碱基的变异。     

MIR-122 慢病毒表达载体构建及稳定转染HepG2 细胞系

目的:构建人mir-122慢病毒表达载体,感染肝癌细胞HepG2,建立稳定表达mir-122的HepG2细胞系。方法:以人has-mir-122成熟序列,设计并合成引物,采用PCR的方法扩增目的基因,并连接到慢病毒表达质粒pGCSIL-GFP(含绿色荧光蛋白GFP基因)中。对重组质粒进行双酶切鉴定后,进行mir-122基因慢病毒(pGCSIL-GFP-miR-122)的包装及病毒滴度测定,用构建好的慢病毒表达载体感染HepG2细胞,qPCR检测感染后细胞中MIR-122的变化。通过流式细胞仪检测荧光蛋白GFP,westernblot检测mir-122靶分子CAT-1,验证pGCSIL-GFP-miR-122在HepG2细胞中的表达效果。结果:pGCSIL-GFP-miR-122经双酶切分析及测序,插入序列正确。qPCR检测显示转入病毒后mir-122在细胞中的表达显著提高。表明mir-122慢病毒表达载体构建成功。流式细胞仪根据GFP荧光筛选纯化感染后细胞,感染率达90%以上。Western blot显示mir-122明显抑制其靶分子表达。进一步验证pGCSIL-GFP-miR-122在细胞中的稳定表达。结论:成功构建mir-122慢病毒表达载体,并建立稳定表达的细胞系,为研究mir-122在人体所起的作用及功能机制打下基础。