Circular RNAs (circRNAs) are broadly expressed in eukaryotic cells, but their molecular mechanism in human disease remains obscure. Here we show that circular antisense non-coding RNA in the INK4 locus (circANRIL), which is transcribed at a locus of atherosclerotic cardiovascular disease on chromosome 9p21, confers atheroprotection by controlling ribosomal RNA (rRNA) maturation and modulating pathways of atherogenesis. CircANRIL binds to pescadillo homologue 1 (PES1), an essential 60S-preribosomal assembly factor, thereby impairing exonuclease-mediated pre-rRNA processing and ribosome biogenesis in vascular smooth muscle cells and macrophages. As a consequence, circANRIL induces nucleolar stress and p53 activation, resulting in the induction of apoptosis and inhibition of proliferation, which are key cell functions in atherosclerosis. Collectively, these findings identify circANRIL as a prototype of a circRNA regulating ribosome biogenesis and conferring atheroprotection, thereby showing that circularization of long non-coding RNAs may alter RNA function and protect from human disease.

Objective— ADAM17 (a disintegrin and metalloproteinase 17) is a sheddase releasing different types of membrane-bound proteins, including adhesion molecules, cytokines, and their receptors as well as inflammatory mediators. Because these substrates modulate important mechanisms of atherosclerosis, we hypothesized that ADAM17 might be involved in the pathogenesis of this frequent disease. Approach and Results— Because Adam17 -knockout mice are not viable, we studied the effect of Adam17 deficiency on atherosclerosis in Adam17 hypomorphic mice ( Adam17 ex/ex ), which have low residual Adam17 expression. To induce atherosclerosis, mice were crossed onto the low-density lipoprotein receptor ( Ldlr )-deficient background. We found that Adam17 ex/ex .Ldlr −/− mice developed ≈1.5-fold larger atherosclerotic lesions, which contained more macrophages and vascular smooth muscle cells than wild-type littermate controls ( Adam17 wt/wt .Ldlr −/− ). Reduced Adam17 -mediated shedding led to significantly increased protein levels of membrane-resident TNFα (tumor necrosis factor) and TNFR2 (tumor necrosis factor receptor 2), resulting in a constitutive activation of TNFR2 signaling. At the same time, Adam17 deficiency promoted proatherosclerotic cellular functions, such as increased proliferation and reduced apoptosis in cultured macrophages and vascular smooth muscle cells and increased adhesion of macrophages to vascular endothelial cells. Because siRNA (small interfering RNA)-mediated knockdown of Tnfr2 rescued from aberrant proliferation and from misregulation of apoptosis in Adam17 -depleted cells, our data indicate that TNFR2 is an important effector of ADAM17 in our mouse model. Conclusions— Our results provide evidence for an atheroprotective role of ADAM17, which might be mediated by cleaving membrane-bound TNFα and TNFR2, thereby preventing overactivation of endogenous TNFR2 signaling in cells of the vasculature.

A current challenge in the era of genome-wide studies is to determine the responsible genes and mechanisms underlying newly identified loci. Screening of the plasma proteome by high-throughput mass spectrometry (MALDI-TOF MS) is considered a promising approach for identification of metabolic and disease processes. Therefore, plasma proteome screening might be particularly useful for identifying responsible genes when combined with analysis of variation in the genome. Here, we describe a proteomic quantitative trait locus (pQTL) study of plasma proteome screens in an F 2 intercross of 455 mice mapped with 177 genetic markers across the genome. A total of 69 of 176 peptides revealed significant LOD scores ($5.35) demonstrating strong genetic regulation of distinct components of the plasma proteome. Analyses were confirmed by mechanistic studies and MALDI-TOF/TOF, liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses of the two strongest pQTLs: A pQTL for mass-to-charge ratio (m/z) 3494 (LOD 24.9, D11Mit151) was identified as the N-terminal 35 amino acids of hemoglobin subunit A (Hba) and caused by genetic variation in Hba. Another pQTL for m/z 8713 (LOD 36.4; D1Mit111) was caused by variation in apolipoprotein A2 (Apoa2) and cosegregated with HDL cholesterol. Taken together, we show that genome-wide plasma proteome profiling in combination with genome-wide genetic screening aids in the identification of causal genetic variants affecting abundance of plasma proteins.T HE central dogma of molecular biology states that sequence information is carried from DNA to RNA to protein (Crick 1970) and it is commonly accepted that differences in protein function or abundance are in most cases responsible for phenotypic differences and susceptibility to disease. To date, mapping complex traits with global gene expression-designated expression quantitative trait loci (eQTL) mapping-has been remarkably successful in identifying the underlying molecular mechanisms (reviewed by Cookson et al. 2009). That approach has been greatly facilitated by advances in expression array technology. However, mRNA levels explain only 40% of the variability in protein levels (Schwanhausser et al. 2011). Therefore, using the proteome as a surrogate, which might be more closely related to the phenotype level, may provide novel insights into genetic regulation of certain phenotypes (Supporting Information, Figure S1).The global analysis of proteins has been exceedingly difficult in the past (Gstaiger and Aebersold 2009) and only few studies have previously attempted a combined analysis of proteome-wide and genome-wide data: An approach in yeast BY4716xRM11-1a segregants (n = 98) successfully identified genetically determined proteins using label-free liquid chromatography-tandem mass spectrometry (LC-MS/ MS) (Foss et al. 2007). In mice, Klose et al. (2002) have investigated brain proteome expression in 64 N2 progeny of a C57BL/6 · SPR backcross using two-dimensional gel electrophoresis. A recent study performed comparative analy...