FMRP and MOV10 regulate Dicer1 expression and dendrite development

Fragile X syndrome results from the loss of expression of the Fragile X Mental Retardation Protein (FMRP). FMRP and RNA helicase Moloney Leukemia virus 10 (MOV10) are important Argonaute (AGO) cofactors for miRNA-mediated translation regulation. We previously showed that MOV10 functionally associates with FMRP. Here we quantify the effect of reduced MOV10 and FMRP expression on dendritic morphology. Murine neurons with reduced MOV10 and FMRP phenocopied Dicer1 KO neurons which exhibit impaired dendritic maturation Hong J (2013), leading us to hypothesize that MOV10 and FMRP regulate DICER expression. In cells and tissues expressing reduced MOV10 or no FMRP, DICER expression was significantly reduced. Moreover, the Dicer1 mRNA is a Cross-Linking Immunoprecipitation (CLIP) target of FMRP Darnell JC (2011), MOV10 Skariah G (2017) and AGO2 Kenny PJ (2020). MOV10 and FMRP modulate expression of DICER1 mRNA through its 3’untranslated region (UTR) and introduction of a DICER1 transgene restores normal neurite outgrowth in the Mov10 KO neuroblastoma Neuro2A cell line and branching in MOV10 heterozygote neurons. Moreover, we observe a global reduction in AGO2-associated microRNAs isolated from Fmr1 KO brain. We conclude that the MOV10-FMRP-AGO2 complex regulates DICER expression, revealing a novel mechanism for regulation of miRNA production required for normal neuronal morphology.

Evaluating genomic biomarkers associated with resistance or sensitivity to chemotherapy in patients with advanced breast and colorectal cancer

Introduction Carcinogenesis is driven by an array of complex genomic patterns; these patterns can render an individual resistant or sensitive to certain chemotherapy agents. The Personalized Oncogenomics (POG) project at BC Cancer has performed integrative genomic analysis of whole tumour genomes and transcriptomes for over 700 patients with advanced cancers, with an aim to predict therapeutic sensitivities. The aim of this study was to utilize the POG genomic data to evaluate a discrete set of biomarkers associated with chemo-sensitivity or-resistance in advanced stage breast and colorectal cancer POG patients. Methods This was a retrospective multi-centre analysis across all BC CANCER sites. All breast and colorectal cancer patients enrolled in the POG program between July 1, 2012 and November 30, 2016 were eligible for inclusion. Within the breast cancer population, those treated with capecitabine, paclitaxel, and everolimus were analyzed, and for the colorectal cancer patients, those treated with capecitabine, bevacizumab, irinotecan, and oxaliplatin were analyzed. The expression levels of the selected biomarkers of interest ( EPHB4, FIGF, CD133, DICER1, DPYD, TYMP, TYMS, TAP1, TOP1, CKDN1A, ERCC1, GSTP1, BRCA1, PTEN, ABCB1, TLE3, and TXNDC17) were reported as mRNA percentiles. Results For the breast cancer population, there were 32 patients in the capecitabine cohort, 15 in the everolimus cohort, and 12 in the paclitaxel cohort. For the colorectal cancer population, there were 29 patients in the bevacizumab cohort, 12 in the oxaliplatin cohort, 29 in the irinotecan cohort, and 6 in the capecitabine cohort. Of the biomarkers evaluated, the strongest associations were found between Bevacizumab-based therapy and DICER1 (P = 0.0445); and between capecitabine therapy and TYMP (P = 0.0553). Conclusions Among breast cancer patients, higher TYMP expression was associated with sensitivity to capecitabine. Among colorectal cancer patients, higher DICER1 expression was associated with sensitivity to bevacizumab-based therapy. This study supports further assessment of the potential predictive value of mRNA expression of these genomic biomarkers.

MicroRNA-Independent Modulation of DICER1 Expression by hAgo2

ABSTRACT Many proteins, including DICER1 and hAgo2, are involved in the biogenesis of microRNAs (miRNAs). Whether hAgo2 regulates DICER1 expression is unknown. Exogenously overexpressed hAgo2 suppressed DICER1 expression at the levels of both protein and mRNA, and the reduction in hAgo2 expression enhanced DICER1 expression. Precursor miRNA processing mediated by DICER1 was also modulated by hAgo2. However, hAgo2 protein did not suppress DICER1 promoter activity. Therefore, hAgo2 protein probably regulates DICER1 expression at the posttranscriptional level. Indeed, hAgo2 protein inhibited the reporter assay of the DICER1 mRNA 3′ untranslated region (3′-UTR). Previous reports have demonstrated that miRNAs (e.g., let-7 and miR-103/107) inhibited DICER1 expression posttranscriptionally. However, hAgo2 still suppressed DICER1 expression in the cells depleted of these miRNAs. Moreover, the reporter activities of the DICER1 mRNA 3′-UTR without these miRNA binding sites were still suppressed by hAgo2. Therefore, in addition to an miRNA-dependent pathway, hAgo2 can also modulate DICER1 expression through an miRNA-independent mechanism. Downregulation of DICER1 expression was further proven to be dependent on both hAgo2 and AUF1 proteins. Interactions of hAgo2 and AUF1 proteins were demonstrated by the coimmunoprecipitation assay. As expected, hAgo2 could not suppress the DICER1 mRNA 3′-UTR reporter with a mutation in the potential AUF1-binding site. Thus, downregulation of DICER1 expression through the 3′-UTR requires both hAgo2 and AUF1.

Brain Dicer1 is down-regulated in a mouse model of Alzheimer’s disease via Aβ42-induced repression of nuclear factor erythroid 2-related factor 2

Background Oxidative stress critically underlies the neurodegenerative pathogenesis of Alzheimer's disease (AD). Depletion of Dicer1, an endoribonuclease central to microRNA maturation, also leads to neurodegeneration. We therefore hypothesized that altered Dicer1 expression may play a role in AD. Results Using immunoblotting and quantitative real-time PCR, we found that Dicer1 protein and mRNA levels were reduced in the hippocampi of animals of the AD mouse model APPswe/PSEN1dE9 compared with littermate controls. SiRNA-meditated Dicer1 knockdown induced oxidative stress, reduced mitochondrial intermembrane potential, and increased apoptosis in cultured neurons. Aβ42 exposure decreased Dicer1 and also down-regulated the oxidative stress–induced transcriptional regulator nuclear factor erythroid 2-related factor 2 (Nrf2). Conversely, Nrf2 overexpression increased Dicer1 mRNA and protein levels and reverted the Aβ42-induced Dicer1 reduction. To further investigate Dicer1 regulation, we cloned Dicer1 promoter variants harboring the Nrf2-binding site, the antioxidant response elements (ARE), into a luciferase reporter and found that simultaneous transfection of Nrf2-expressing plasmid increased luciferase expression from these promoter constructs. ChIP assays indicated that Nrf2 directly interacted with the ARE motifs in the Dicer1 promoter. Furthermore, Dicer1 overexpression in cultured neurons reverted Aβ42-induced neurite deficits. Of note, injection of Dicer1-expressing adenovirus into the hippocampus of the AD mice significantly improved spatial learning. Conclusions These findings indicate that Dicer1 expression is reduced in the AD brain and that chronic Aβ exposure decreases Dicer1 levels in neurons via Nrf2–ARE signaling. Our results uncover a significant role for Dicer1 in AD and highlight that Dicer1 expression responds to oxidative stress in the brain.

Impaired miRNA processing by DICER1 downregulation endows thyroid cancer with increased aggressiveness

The global downregulation of miRNAs (miRs) is emerging as a common hallmark of cancer. However, the mechanisms underlying this phenomenon are not well known. We identified that the oncogenic miR-146b-5p attenuates miRNA biosynthesis by targeting DICER1 and reducing its expression. DICER1 overexpression inhibited all the miR-146b-induced aggressive phenotypes in thyroid cells. Systemic injection of an antimiR-146b in mice with orthotopic thyroid tumors suppressed tumor growth and recovered DICER1 levels. Notably, DICER1 downregulation promoted proliferation, migration, invasion and epithelial-mesenchymal transition through miR downregulation. Our analysis of TCGA revealed a general decrease in DICER1 expression in thyroid cancer that was associated with a worse clinical outcome. Administration of the small molecule enoxacin to promote DICER1 complex activity reduced tumor aggressiveness bothin vitroandin vivo. Overall, our data establish DICER1 as a tumor suppressor and that the oncogenic miR-146b contributes to its downregulation. Moreover, this study highlights a potential therapeutic application of RNA-based therapies including miR-inhibitors and restoration of the biogenesis machinery, which may provide treatments in for thyroid and other cancers.