Common Genetic Variants Contribute to Risk of Transposition of the Great Arteries.

RATIONALE: Dextro-transposition of the great arteries (D-TGA) is a severe congenital heart defect which affects approximately 1 in 4,000 live births. While there are several reports of D-TGA patients with rare variants in individual genes, the majority of D-TGA cases remain genetically elusive. Familial recurrence patterns and the observation that most cases with D-TGA are sporadic suggest a polygenic inheritance for the disorder, yet this remains unexplored. OBJECTIVE: We sought to study the role of common single nucleotide polymorphisms (SNPs) in risk for D-TGA. METHODS AND RESULTS: We conducted a genome-wide association study in an international set of 1,237 patients with D-TGA and identified a genome-wide significant susceptibility locus on chromosome 3p14.3, which was subsequently replicated in an independent case-control set (rs56219800, meta-analysis P=8.6x10 CONCLUSIONS: This work provides support for a polygenic architecture in D-TGA and identifies a susceptibility locus on chromosome 3p14.3 nea

A functional screen identifies specific microRNAs capable of inhibiting human melanoma cell viability

Malignant melanoma is an aggressive form of skin cancer with poor prognosis. Despite improvements in awareness and prevention of this disease, its incidence is rapidly increasing. MicroRNAs (miRNAs) are a class of small RNA molecules that regulate cellular processes by repressing messenger RNAs (mRNAs) with partially complementary target sites. Several miRNAs have already been shown to attenuate cancer phenotypes, by limiting proliferation, invasiveness, tumor angiogenesis, and stemness. Here, we employed a genome-scale lentiviral human miRNA expression library to systematically survey which miRNAs are able to decrease A375 melanoma cell viability. We highlight the strongest inhibitors of melanoma cell proliferation, including the miR-15/16, miR-141/200a and miR-96/182 families of miRNAs and miR-203. Ectopic expression of these miRNAs resulted in long-term inhibition of melanoma cell expansion, both in vitro and in vivo. We show specifically miR-16, miR-497, miR-96 and miR-182 are efficient effectors when introduced as synthetic miRNAs in several melanoma cell lines. Our study provides a comprehensive interrogation of miRNAs that interfere with melanoma cell proliferation and viability, and offers a selection of miRNAs that are especially promising candidates for application in melanoma therapy

An image-based miRNA screen identifies miRNA-135s as regulators of CNS axon growth and regeneration by targeting krüppel-like factor 4

During embryonic development, axons extend over long distances to establish functional connections. In contrast, axon regeneration in the adult mammalian CNS is limited in part by a reduced intrinsic capacity for axon growth. Therefore, insight into the intrinsic control of axon growth may provide new avenues for enhancing CNS regeneration. Here, we performed one of the first miRNome-wide functional miRNA screens to identify miRNAs with robust effects on axon growth. High-content screening identified miR-135a and miR-135b as potent stimulators of axon growth and cortical neuron migration in vitro and in vivo in male and female mice. Intriguingly, both of these developmental effects of miR-135s relied in part on silencing of Krüppel-like factor 4 (KLF4), a well known intrinsic inhibitor of axon growth and regeneration. These results prompted us to test the effect of miR-135s on axon regeneration after injury. Our results show that intravitreal application of miR-135s facilitates retinal ganglion cell (RGC) axon regeneration after optic nerve injury in adult mice in part by repressing KLF4. In contrast, depletion of miR-135s further reduced RGC axon regeneration. Together, these data identify a novel neuronal role for miR-135s and the miR-135–KLF4 pathway and highlight the potential of miRNAs as tools for enhancing CNS axon regeneration

Endogenous miRNA expression and expression after lentiviral transduction in A375 cells.

<p>qPCR data are relative to U6 small RNA. Endogenous: empty vector-transduced, ectopic: miRNA-transduced.</p

Transcriptome analysis after miR-203 transfection.

<p>(A) A375 cells were transfected with either miR-203 or scrambled control and the transcriptome was quantified by RNA-Seq. All differentially expressed genes are plotted in the left graph, while only the differentially expressed genes containing miR-203 target sites are plotted in the right graph. Genes with miR-203 target sites are much more likely to be downregulated after miR-203 overexpression, and downregulated genes are highly enriched for genes with miR-203 target sites (p<0.0001). (B) One of the differentially expressed genes after miR-203 transfection is BIRC5. Repression was examined at both the mRNA and the protein level by qPCR (left) and Western blot (right) respectively. The BIRC5 transcript and its protein product survivin are both reduced after miR-203 transfection, but also after siBRAF transfection.</p

A genome-wide screen for miRNAs that inhibit A375 melanoma cell growth.

<p>(A) Inhibition of melanoma growth was measured by means of cell viability and cell count. For each sample a B-score was calculated and B-scores from both assays are plotted against each other. There is a strong correlation between both assays. The B-scores for one miRNA, miR-518b, fell outside the range of the graph: they were −6 for cell count and −12 for cell viability. (B) A comparison with a normal distribution shows that the cell viability screen is sensitive for identifying growth-inhibitory miRNAs. A concomitant estimate of the false discovery rate is shown in grey fill (secondary axis). (C) 55 potential inhibitory miRNAs were tested in a confirmation screen against 11 empty vector samples and a population of 28 miRNAs with small or no effects in the primary screen. Box plots show values between 25^th and 75^th percentile in boxes, and the outermost values as whiskers. 20 of 55 inhibitory miRNAs scored better than any of the control miRNAs (below dashed line). *p = 6.8*10⁻⁵, **p = 1.6*10⁻⁶, ***p = 5.4*10⁻¹⁰. (D) Individual hits selected for follow-up, and their relative effect on cell viability. A virus containing a short-hairpin construct targeting BRAF was used as a positive control. Error bars represent standard deviation of three samples.</p