Preventing Civic Space Restrictions: an Exploratory Study of Successful Resistance Against NGO Laws

In many countries around the world, civil society organizations are facing increasing restrictions that constrain their autonomy, capacity and/or freedom of action. While the general phenomenon of shrinking civic space and the adoption of legal restrictions in particular have become more widespread since the early 2000s, there are also cases in which governmental attempts to adopt restrictive NGO laws have been frustrated, aborted or, at least, significantly mitigated as a consequence of domestic and/or international resistance. This PRIF Report takes a look at four such cases (Azerbaijan, Kenya, Kyrgyzstan and Zambia) and identifies conditions and dynamics that help understand successful resistance against legal civic space restrictions

Ex vivo drug response profiling for response and outcome prediction in hematologic malignancies: the prospective non-interventional SMARTrial

Ex vivo drug response profiling is a powerful tool to study genotype-drug response associations and is being explored as a tool set for precision medicine in cancer. Here we conducted a prospective non-interventional trial to investigate feasibility of ex vivo drug response profiling for treatment guidance in hematologic malignancies (SMARTrial, NCT03488641 ). The primary endpoint to provide drug response profiling reports within 7 d was met in 91% of all study participants (N = 80). Secondary endpoint analysis revealed that ex vivo resistance to chemotherapeutic drugs predicted chemotherapy treatment failure in vivo. We confirmed the predictive value of ex vivo response to chemotherapy in a validation cohort of 95 individuals with acute myeloid leukemia treated with daunorubicin and cytarabine. Ex vivo drug response profiles improved ELN-22 risk stratification in individuals with adverse risk. We conclude that ex vivo drug response profiling is clinically feasible and has the potential to predict chemotherapy response in individuals with hematologic malignancies beyond clinically established genetic markers

Comment on 'YcgC represents a new protein deacetylase family in prokaryotes'

Lysine acetylation is a post-translational modification that is conserved from bacteria to humans. It is catalysed by the activities of lysine acetyltransferases, which use acetyl-CoA as the acetyl-donor molecule, and lysine deacetylases, which remove the acetyl moiety. Recently, it was reported that YcgC represents a new prokaryotic deacetylase family with no apparent homologies to existing deacetylases (Tu et al., 2015). Here we report the results of experiments which demonstrate that YcgC is not a deacetylase

Insights into Lysine Deacetylation of Natively Folded Substrate Proteins by Sirtuins

Sirtuins are NAD(+)-dependent lysine deacylases, regulating a variety of cellular processes. The nuclear Sirt1, the cytosolic Sirt2, and the mitochondrial Sirt3 are robust deacetylases, whereas the other sirtuins have preferences for longer acyl chains. Most previous studies investigated sirtuin-catalyzed deacylation on peptide substrates only. We used the genetic code expansion concept to produce natively folded, site-specific, and lysine-acetylated Sirt1-3 substrate proteins, namely Ras-related nuclear, p53, PEPCK1, superoxide dismutase, cyclophilin D, and Hsp10, and analyzed the deacetylation reaction. Some acetylated proteins such as Ras-related nuclear, p53, and Hsp10 were robustly deacetylated by Sirt1-3. However, other reported sirtuin substrate proteins such as cyclophilin D, superoxide dismutase, and PEPCK1 were not deacetylated. Using a structural and functional approach, we describe the ability of Sirt1-3 to deacetylate two adjacent acetylated lysine residues. The dynamics of this process have implications for the lifetime of acetyl modifications on di-lysine acetylation sites and thus constitute a new mechanism for the regulation of proteins by acetylation. Our studies support that, besides the primary sequence context, the protein structure is a major determinant of sirtuin substrate specificity

Small GTP-binding protein Ran is regulated by posttranslational lysine acetylation

Ran is a small GTP-binding protein of the Ras superfamily regulating fundamental cellular processes: nucleo-cytoplasmic transport, nuclear envelope formation and mitotic spindle assembly. An intracellular Ran.GTP/Ran.GDP gradient created by the distinct subcellular localization of its regulators RCC1 and RanGAP mediates many of its cellular effects. Recent proteomic screens identified five Ran lysine acetylation sites in human and eleven sites inmouse/rat tissues. Some of these sites are located in functionally highly important regions such as switch I and switch II. Here, we show that lysine acetylation interferes with essential aspects of Ran function: nucleotide exchange and hydrolysis, subcellular Ran localization, GTP hydrolysis, and the interaction with import and export receptors. Deacetylation activity of certain sirtuins was detected for two Ran acetylation sites in vitro. Moreover, Ran was acetylated by CBP/p300 and Tip60 in vitro and on transferase overexpression in vivo. Overall, this study addresses many important challenges of the acetylome field, which will be discussed

Structural and Biochemical Basis for the Inhibitory Effect of Liprin-alpha 3 on Mouse Diaphanous 1 (mDia1) Function

Diaphanous-related formins are eukaryotic actin nucleation factors regulated by an autoinhibitory interaction between the N-terminal RhoGTPase-binding domain (mDia(N)) and the C-terminal Diaphanous-autoregulatory domain (DAD). Although the activation of formins by Rho proteins is well characterized, its inactivation is only marginally understood. Recently, liprin-alpha 3 was shown to interact with mDia1. Overexpression of liprin-alpha 3 resulted in a reduction of the cellular actin filament content. The molecular mechanisms of how liprin-alpha 3 exerts this effect and counteracts mDia1 activation by RhoA are unknown. Here, we functionally and structurally define a minimal liprin-alpha 3 core region, sufficient to recapitulate the liprin-alpha 3 determined mDia1-respective cellular functions. We show that liprin-alpha 3 alters the interaction kinetics and thermodynamics of mDia(N) with RhoA center dot GTP and DAD. RhoA displaces liprin-alpha 3 allosterically, whereas DAD competes with liprin-alpha 3 for a highly overlapping binding site on mDia(N). Liprin-alpha 3 regulates actin polymerization by lowering the regulatory potency of RhoA and DAD on mDia(N). We present a model of a mechanistically unexplored and new aspect of mDia(N) regulation by liprin-alpha 3

Additional file 1: of Cognitive Bias Modification for paranoia (CBM-pa): study protocol for a randomised controlled trial

Examples of CBM-pa and control conditions. Figure S1a. Instructions for the task are included within the programme and displayed before the participant begins. There is no time limit on the instructions. Figure S1b. A CBM-pa intervention passage. The passage is initially ambiguous, but the final word solution requires the participant to interpret in a non-paranoid way. Participants are initially given 20 s to read the passage, and 20 s to complete the word solution. Figure S1c. The participant is required to enter the first missing letter of the word, and is given positive feedback if they do so. Figure S1d. An incorrect response prompts the participant to try again, and more letters are given to help. Participants are given 23 s to respond. Figure S1e. The solution is then shown for 5 s regardless of whether a participant types the letter correctly or not. Figure S1f. A comprehension question is asked to encourage the participant to engage with the meaning of the passage. Participants are given 20 s to answer. Figure S1g. The figure above is shown when a participant responds in a non-paranoid way, for a maximum of 20 s. Figure S1h The figure above is shown when a participant responds in a paranoid way, for a maximum of 20 s. Figure S1i. A text-reading control passage. The text reading control programme is presented in an identical fashion to the CBM-pa. (DOCX 158 kb

Additional file 2: of Cognitive Bias Modification for paranoia (CBM-pa): study protocol for a randomised controlled trial

SPIRIT 2013 Checklist: recommended items to address in a clinical trial protocol and related documents. (DOC 120 kb