Studies on the Restriction of Murine Leukemia Viruses by Mouse APOBEC3

APOBEC3 proteins function to restrict the replication of retroviruses. One mechanism of this restriction is deamination of cytidines to uridines in (−) strand DNA, resulting in hypermutation of guanosines to adenosines in viral (+) strands. However, Moloney murine leukemia virus (MoMLV) is partially resistant to restriction by mouse APOBEC3 (mA3) and virtually completely resistant to mA3-induced hypermutation. In contrast, the sequences of MLV genomes that are in mouse DNA suggest that they were susceptible to mA3-induced deamination when they infected the mouse germline. We tested the possibility that sensitivity to mA3 restriction and to deamination resides in the viral gag gene. We generated a chimeric MLV in which the gag gene was from an endogenous MLV in the mouse germline, while the remainder of the viral genome was from MoMLV. This chimera was fully infectious but its response to mA3 was indistinguishable from that of MoMLV. Thus, the Gag protein does not seem to control the sensitivity of MLVs to mA3. We also found that MLVs inactivated by mA3 do not synthesize viral DNA upon infection; thus mA3 restriction of MLV occurs before or at reverse transcription. In contrast, HIV-1 restricted by mA3 and MLVs restricted by human APOBEC3G do synthesize DNA; these DNAs exhibit APOBEC3-induced hypermutation

Digital ulcers predict a worse disease course in patients with systemic sclerosis

Objective: Systemic sclerosis (SSc) is a systemic autoimmune disease with high morbidity and significant mortality. There is a great need of predictors that would allow risk stratification of patients with SSc and ultimately initiation of treatment early enough to ensure optimal clinical results. In this study, we evaluated whether a history of digital ulcers (HDU) at presentation may be a predictor of vascular outcomes and of overall clinical worsening and death in patients with SSc. Methods: Patients from the EULAR Scleroderma Trials and Research (EUSTAR) database, satisfying at inclusion the 1980 American College of Rheumatology classification criteria for SSc, who had a follow-up of at least 3 years since baseline or who have died, were included in the analysis. HDU at presentation as a predictor of disease worsening or death was evaluated by Cox proportional hazards regression analysis. Results :3196 patients matched the inclusion criteria (male sex 13.2%, 33.4% diffuse subset). At presentation, 1092/3196 patients had an HDU (34.1%). In multivariable analysis adjusting for age, gender and all parameters considered potentially significant, HDU was predictive for the presence of active digital ulcers (DUs) at prospective visits (HR (95% CI)): 2.41(1.91 to 3.03), p<0.001, for an elevated systolic pulmonary arterial pressure on heart ultrasound (US-PAPs):1.36 (1.03 to 1.80), p=0.032, for any cardiovascular event (new DUs, elevated US-PAPs or LV failure):3.56 (2.26 to 5.62), p<0.001, and for death (1.53 (1.16 to 2.02), p=0.003). Conclusions :In patients with SSc, HDU at presentation predicts the occurrence of DUs at follow-up and is associated with cardiovascular worsening and decreased survival

Vasodilators and low-dose acetylsalicylic acid are associated with a lower incidence of distinct primary myocardial disease manifestations in systemic sclerosis: results of the DeSScipher inception cohort study

OBJECTIVES: To investigate the influence of vasodilator drugs on the occurrence of features depending on myocardial ischaemia/fibrosis (ventricular arrhythmias, Q waves, cardiac blocks, pacemaker implantation, left ventricular ejection fraction (LVEF) <55%, and/or congestive heart failure and sudden cardiac death) in systemic sclerosis (SSc). METHODS: 601 patients with SSc were enrolled from 1 December 2012 to 30 November 2015 and had a second visit 0.5-4 years apart. 153 received no vasodilators; 448 received vasodilator therapy (ie, calcium channel blockers and/or ACE inhibitors or angiotensin II receptor blockers or combinations of them), 89 of them being also treated with either endothelin receptor antagonists or PDE5 inhibitors or prostanoids. Associations between the occurrence of myocardial disease manifestations and any demographic, disease and therapeutic aspect were investigated by Cox regression analysis. A Cox frailty survival model with centre of enrolment as random effect was performed. RESULTS: During 914 follow-up patient-years, 12 ventricular arrhythmias, 5 Q waves, 40 cardiac blocks, 6 pacemaker implantations and 19 reduced LVEF and/or congestive heart failure (CHF) occurred. In multivariate Cox regression analysis, vasodilator therapy was associated with a lower incidence of ventricular arrhythmias (p=0.03); low-dose acetylsalicylic acid (ASA) with a lower incidence of cardiac blocks and/or Q waves and/or pacemaker implantation (p=0.02); active disease with a higher incidence of LVEF <55% and/or CHF and cardiac blocks and/or Q waves and/or pacemaker implantation (p=0.05). CONCLUSIONS: The present study might suggest a preventative effect on the occurrence of distinct myocardial manifestations by vasodilator therapy and low-dose ASA

Phenotypes Determined by Cluster Analysis and Their Survival in the Prospective European Scleroderma Trials and Research Cohort of Patients With Systemic Sclerosis

Objective: Systemic sclerosis (SSc) is a heterogeneous connective tissue disease that is typically subdivided into limited cutaneous SSc (lcSSc) and diffuse cutaneous SSc (dcSSc) depending on the extent of skin involvement. This subclassification may not capture the entire variability of clinical phenotypes. The European Scleroderma Trials and Research (EUSTAR) database includes data on a prospective cohort of SSc patients from 122 European referral centers. This study was undertaken to perform a cluster analysis of EUSTAR data to distinguish and characterize homogeneous phenotypes without any a priori assumptions, and to examine survival among the clusters obtained. / Methods: A total of 11,318 patients were registered in the EUSTAR database, and 6,927 were included in the study. Twenty‐four clinical and serologic variables were used for clustering. / Results: Clustering analyses provided a first delineation of 2 clusters showing moderate stability. In an exploratory attempt, we further characterized 6 homogeneous groups that differed with regard to their clinical features, autoantibody profile, and mortality. Some groups resembled usual dcSSc or lcSSc prototypes, but others exhibited unique features, such as a majority of lcSSc patients with a high rate of visceral damage and antitopoisomerase antibodies. Prognosis varied among groups and the presence of organ damage markedly impacted survival regardless of cutaneous involvement. / Conclusion: Our findings suggest that restricting subsets of SSc patients to only those based on cutaneous involvement may not capture the complete heterogeneity of the disease. Organ damage and antibody profile should be taken into consideration when individuating homogeneous groups of patients with a distinct prognosis

Digital ulcers predict a worse disease course in patients with systemic sclerosis

OBJECTIVE: Systemic sclerosis (SSc) is a systemic autoimmune disease with high morbidity and significant mortality. There is a great need of predictors that would allow risk stratification of patients with SSc and ultimately initiation of treatment early enough to ensure optimal clinical results. In this study, we evaluated whether a history of digital ulcers (HDU) at presentation may be a predictor of vascular outcomes and of overall clinical worsening and death in patients with SSc. METHODS: Patients from the EULAR Scleroderma Trials and Research (EUSTAR) database, satisfying at inclusion the 1980 American College of Rheumatology classification criteria for SSc, who had a follow-up of at least 3\u2005years since baseline or who have died, were included in the analysis. HDU at presentation as a predictor of disease worsening or death was evaluated by Cox proportional hazards regression analysis. RESULTS: 3196 patients matched the inclusion criteria (male sex 13.2%, 33.4% diffuse subset). At presentation, 1092/3196 patients had an HDU (34.1%). In multivariable analysis adjusting for age, gender and all parameters considered potentially significant, HDU was predictive for the presence of active digital ulcers (DUs) at prospective visits (HR (95% CI)): 2.41 (1.91 to 3.03), p<0.001, for an elevated systolic pulmonary arterial pressure on heart ultrasound (US-PAPs):1.36 (1.03 to 1.80), p=0.032, for any cardiovascular event (new DUs, elevated US-PAPs or LV failure): 3.56 (2.26 to 5.62), p<0.001, and for death (1.53 (1.16 to 2.02), p=0.003). CONCLUSIONS: In patients with SSc, HDU at presentation predicts the occurrence of DUs at follow-up and is associated with cardiovascular worsening and decreased survival

Structural imprints in vivo decode RNA regulatory mechanisms

Visualizing the physical basis for molecular behavior inside living cells is a grand challenge in biology. RNAs are central to biological regulation, and RNA’s ability to adopt specific structures intimately controls every step of the gene expression program(1). However, our understanding of physiological RNA structures is limited; current in vivo RNA structure profiles view only two of four nucleotides that make up RNA(2,3). Here we present a novel biochemical approach, In Vivo Click SHAPE (icSHAPE), that enables the first global view of RNA secondary structures of all four bases in living cells. icSHAPE of mouse embryonic stem cell transcriptome versus purified RNA folded in vitro shows that the structural dynamics of RNA in the cellular environment distinguishes different classes of RNAs and regulatory elements. Structural signatures at translational start sites and ribosome pause sites are conserved from in vitro, suggesting that these RNA elements are programmed by sequence. In contrast, focal structural rearrangements in vivo reveal precise interfaces of RNA with RNA binding proteins or RNA modification sites that are consistent with atomic-resolution structural data. Such dynamic structural footprints enable accurate prediction of RNA-protein interactions and N6-methyladenosine (m(6)A) modification genome-wide. These results open the door for structural genomics of RNA in living cells and reveal key physiological structures controlling gene expression