Search

MicroRNAs are important coordinators of circadian regulation that mediate the fine-tuning of gene expression. Although many studies have shown the effects of individual miRNAs on the circadian clock, the global functional miRNA-mRNA interaction network involved in the circadian system remains poorly understood. Here, we used CLEAR (Covalent Ligation of Endogenous Argonaute-bound RNAs)-CLIP (Cross-Linking and Immuno-Precipitation) to explore the regulatory functions of miRNAs in the circadian system by comparing the miRNA-mRNA interactions between Drosophila wild-type strain W(1118) and a mutant of the key circadian transcriptional regulator Clock (Clk(jrk)). This experimental approach unambiguously identified tens of thousands of miRNA-mRNA interactions in both the head and body. The miRNA-mRNA interactome showed dramatic changes in the Clk(jrk) flies. Particularly, among ~300 miRNA-mRNA circadian relevant interactions, multiple interactions involving core clock genes pdp1, tim and vri displayed distinct changes as a result of the Clk mutation. Based on the CLEAR-CLIP analysis, we found a novel regulation of the circadian rhythm and sleep by the miR-375-timeless interaction. The results indicated that Clk disruption abolished normal rhythmic expression of miR-375 and the functional regulation occurred in the l-LNv neurons, where miR-375 modulated circadian rhythm and sleep via targeting timeless. This work provides the first global view of miRNA regulation in the circadian rhythm.

Krüppel homologue 1 (Kr-h1) is a zinc finger transcription factor that is upregulated in insects by juvenile hormone (JH) in metamorphosis and adult reproduction. The molecular function of Kr-h1 in reproduction remains largely unknown. Here we report that AaKr-h1 functions as an important transcription regulator in adult female Aedes aegypti mosquitoes. The amount of AaKr-h1 protein increases with rising JH levels after adult emergence, reaches its peak at 48 h after eclosion, then decreases gradually and disappears after blood feeding. RNA interference (RNAi)-mediated depletion of AaKr-h1 substantially reduced egg production after blood feeding. Using a chromatin immunoprecipitation cloning approach, we identified in vivo AaKr-h1 binding sites in previtellogenic female mosquitoes. Binding of AaKr-h1 to the target genes correlated with its protein abundance. Interestingly, RNAi experiments indicated that AaKr-h1 played distinct roles when it bound to individual target genes. For example, depletion of AaKr-h1 led to substantial upregulation of AAEL005545 and AAEL004444, but also significantly decreased the expression of AAEL005957 and AAEL013177 when compared with the control mosquitoes. In summary, AaKr-h1 directly binds to the regulatory regions of its target genes and acts as a transcriptional activator or a repressor in a promoter-specific manner.

In this study, we found that let-7c-5p expression was clearly downregulated in breast cancer tissues compared with that of corresponding adjacent tissues. Furthermore, overexpression of let-7c-5p in MCF-7 breast cancer cells could significantly inhibit cell proliferation and induce cell apoptosis. The target genes of let-7c-5p were predicted by the way of bioinformatics, and validated by dual luciferase reporter assay and western blotting demonstrating that excision repair cross complementing 6 (ERCC6) gene was a direct target. Collectively, the present study suggested that let-7c-5p acted as a tumor suppressor in breast cancer possibly by negatively regulating ERCC6, which took an important part in nucleotide excision repair and it may provide a new potential strategy for breast cancer therapy.

Juvenile hormone (JH) regulates many aspects of insect development and reproduction. In some processes, JH plays a critical role in defining the action of the steroid hormone 20-hydroxyecdysone (20E). In Aedes aegypti mosquitoes, JH prepares newly emerged female adults to become competent to synthesize vitellogenin in response to 20E after blood ingestion. The molecular basis of this competence is still not well understood. Here, we report that JH regulates pre-mRNA splicing of the taiman gene, which encodes a key transcriptional regulator required for both JH- and 20E-controlled gene expression. JH stimulated the production of the Taiman isoforms A/B, while reducing the levels of the isoforms C/D, in the fat body after adult eclosion. The appearance of the A/B isoforms in maturing mosquitoes was accompanied by acquisition of the competence to respond to 20E. Depletion of the A/B isoforms, by inhibiting the alternative splicing or by isoform-specific RNA interference, considerably diminished the 20E-induced gene expression after a blood meal and substantially impaired oocyte development. In accordance with this observation, further studies indicated that in the presence of 20E, the Taiman A/B isoforms showed much stronger interactions with the 20E receptor complex than the Taiman C/D isoforms. In contrast, all four isoforms displayed similar capabilities of forming active JH receptor complexes with the methoprene-tolerant protein (Met). This study suggested that JH confers the competence to newly emerged female mosquitoes by regulating mRNA splicing to generate the Taiman isoforms that are essential for the vitellogenic 20E response. Published version

The vital role of microRNAs (miRNAs) involved in gene expression regulation has been confirmed in many biological processes. With the growing power and reducing cost of next-generation sequencing, more and more researchers turn to apply this high-throughput method to solve their biological problems. For miRNAs with known sequences, their expression profiles can be generated from the sequencing data. It also allows us to identify some novel miRNAs and explore the sequence variations under different conditions. Currently, there are a handful of tools available to analyze the miRNA sequencing data with separated or combined features, such as reads preprocessing, mapping and differential expression analysis. However, to our knowledge, a hands-on guideline for miRNA sequencing data analysis covering all steps is not available. Here we will utilize a set of published tools to perform the miRNA analysis with detailed explanation. Particularly, the miRNA target prediction and annotation may provide useful information for further experimental verification.

Drastic changes in gene expression occur after adult female mosquitoes take a blood meal and use the nutrients for egg maturation. A growing body of evidence indicates that microRNAs (miRNAs) contribute to this tightly controlled tissue- and stage-specific gene expression. To investigate the role of miRNAs, we monitored miRNA expression in the mosquito Anopheles gambiae during the 72-h period immediately after blood feeding. We also measured the association of miRNAs with Argonaute 1 (Ago1) and Argonaute 2 (Ago2) to assess the functional status of individual miRNA species. Overall, 173 mature miRNAs were precipitated with Ago1 and Ago2, including 12 new miRNAs, the orthologs of which are found thus far only in other Anopheles species. Ago1 is the predominant carrier of miRNAs in Anopheles gambiae. The abundance and Ago loading of most of the mature miRNAs were relatively stable after blood ingestion. However, miRNAs of the miR-309/286/2944 cluster were considerably upregulated after blood feeding. Injection of the specific antagomir for miR-309 resulted in smaller developing oocytes and ultimately fewer eggs. In addition, the Ago association of some miRNAs was not proportional to their cellular abundance, suggesting that integration of miRNAs into the Ago complexes is regulated by additional mechanisms.

Juvenile hormone (JH) controls many biological events in insects by triggering dramatic changes in gene expression in target cells. The Methoprene-tolerant (MET) protein, an intracellular JH receptor, acts as a transcriptional regulator and binds to the promoters of tissue- and stage-specific JH target genes when JH is present. Our recent study has demonstrated that the transcriptional activation by MET is modulated by a membrane- initiated JH signaling pathway, involving phospholipase C (PLC) and calcium/calmodulin-dependent protein kinase II (CaMKII). Here we report that protein kinase C (PKC) is another essential intermediate of this pathway. PKC was activated by JH and this action was PLC-dependent. Inhibition of the PKC activity substantially weakened the JH-induced gene expression in mosquito cells. RNAi experiments indicated that several PKC isoforms were involved in the JH action during the post- emergence development of adult female mosquitoes. JH treatment considerably increased the binding of MET to the promoters of JH response genes in cultured mosquito abdomens that were collected from newly emerged female adults. The JH-induced DNA binding of MET was hindered when the abdomens were treated with a PKC inhibitor and JH. Therefore, the results suggest that PKC modulates the transactivation activity of MET by enhancing the binding of MET to JH response elements in the JH target genes. This mechanism may allow for variable and stage- and tissue-specific genomic responses to JH.

Background Recent studies have shown that C-reactive protein (CRP) may be associated with breast cancer. The purpose of this study is to summarize the predictive role of CRP for survival in breast cancer as shown in all available studies worldwide. Methods Related studies were identified and evaluated for quality through multiple search strategies. Data were collected from studies comparing overall, cancer-specific, and disease-free survival (OS, CSS, and DFS) in patients with elevated CRP levels and those having lower levels. Studies were pooled, and combined hazard ratios (HRs) of CRP for survival were calculated. Results A total of 10 studies (n=4,502) were included for this meta-analysis (9 for OS, 3 for CSS, and 3 for DFS). For overall and disease-free survival, the pooled HRs of CRP were significant at 1.62 (95% confidence interval [95% CI], 1.20-2.18) and 1.81 (95% CI, 1.44-2.26), respectively. For cancer-specific survival, the pooled HR in higher CRP expression in breast cancer was 2.08 (95% CI, 1.48-2.94), which could strongly predict poorer survival in breast cancer. Conclusions CRP has a critical prognostic value in patients with breast cancer as an inflammation biomarker.