Synthesis of Terminal Ribose Analogues of Adenosine 5′-Diphosphate Ribose as Probes for the Transient Receptor Potential Cation Channel TRPM2

TRPM2 (transient receptor potential cation channel, subfamily M, member 2) is a nonselective cation channel involved in the response to oxidative stress and in inflammation. Its role in autoimmune and neurodegenerative diseases makes it an attractive pharmacological target. Binding of the nucleotide adenosine 5′-diphosphate ribose (ADPR) to the cytosolic NUDT9 homology (NUDT9H) domain activates the channel. A detailed understanding of how ADPR interacts with the TRPM2 ligand binding domain is lacking, hampering the rational design of modulators, but the terminal ribose of ADPR is known to be essential for activation. To study its role in more detail, we designed synthetic routes to novel analogues of ADPR and 2′-deoxy-ADPR that were modified only by removal of a single hydroxyl group from the terminal ribose. The ADPR analogues were obtained by coupling nucleoside phosphorimidazolides to deoxysugar phosphates. The corresponding C2″-based analogues proved to be unstable. The C1″- and C3″-ADPR analogues were evaluated electrophysiologically by patch-clamp in TRPM2-expressing HEK293 cells. In addition, a compound with all hydroxyl groups of the terminal ribose blocked as its 1″-β-O-methyl-2″,3″-O-isopropylidene derivative was evaluated. Removal of either C1″ or C3″ hydroxyl groups from ADPR resulted in loss of agonist activity. Both these modifications and blocking all three hydroxyl groups resulted in TRPM2 antagonists. Our results demonstrate the critical role of these hydroxyl groups in channel activation.</p

Second messenger analogues highlight unexpected substrate sensitivity of CD38: total synthesis of the hybrid “L-cyclic inosine 5'-diphosphate ribose”

The multifunctional, transmembrane glycoprotein human CD38 catalyses the synthesis of three key Ca2+-mobilising messengers, including cyclic adenosine 5′-diphosphate ribose (cADPR), and CD38 knockout studies have revealed the relevance of the related signalling pathways to disease. To generate inhibitors of CD38 by total synthesis, analogues based on the cyclic inosine 5′-diphosphate ribose (cIDPR) template were synthesised. In the first example of a sugar hybrid cIDPR analogue, “L-cIDPR”, the natural “northern” N1-linked D-ribose of cADPR was replaced by L-ribose. L-cIDPR is surprisingly still hydrolysed by CD38, whereas 8-Br-L-cIDPR is not cleaved, even at high enzyme concentrations. Thus, the inhibitory activity of L-cIDPR analogues appears to depend upon substitution of the base at C-8; 8-Br-L-cIDPR and 8-NH2-L-cIDPR inhibit CD38-mediated cADPR hydrolysis (IC50 7 μM and 21 µM respectively) with 8-Br-L-cIDPR over 20-fold more potent than 8-Br-cIDPR. In contrast, L-cIDPR displays a comparative 75-fold reduction in activity, but is only ca 2-fold less potent than cIDPR itself. Molecular modelling was used to explore the interaction of the CD38 catalytic residue Glu-226 with the “northern” ribose. We propose that Glu226 still acts as the catalytic residue even for an L-sugar substrate. 8-Br-L-cIDPR potentially binds non-productively in an upside-down fashion. Results highlight the key role of the “northern” ribose in the interaction of cADPR with CD38

Second messenger analogues highlight unexpected substrate sensitivity of CD38: total synthesis of the hybrid “L-cyclic inosine 5'-diphosphate ribose”

The multifunctional, transmembrane glycoprotein human CD38 catalyses the synthesis of three key Ca2+ - mobilising messengers, including cyclic adenosine 5-diphosphate ribose (cADPR), and CD38 knockout studies have revealed the relevance of the related signalling pathways to disease. To generate inhibitors of CD38 by total synthesis, analogues based on the cyclic inosine 5-diphosphate ribose (cIDPR) template were synthesised. In the first example of a sugar hybrid cIDPR analogue, “L-cIDPR”, the natural “northern” N1-linked D-ribose of cADPR was replaced by L-ribose. L-cIDPR is surprisingly still hydrolysed by CD38, whereas 8-Br-L-cIDPR is not cleaved, even at high enzyme concentrations. Thus, the inhibitory activity of L-cIDPR analogues appears to depend upon substitution of the base at C-8; 8- Br-L-cIDPR and 8-NH2-L-cIDPR inhibit CD38-mediated cADPR hydrolysis (IC50 7 M and 21 µM respectively) with 8-Br-L-cIDPR over 20-fold more potent than 8-Br-cIDPR. In contrast, L-cIDPR displays a comparative 75-fold reduction in activity, but is only ca 2-fold less potent than cIDPR itself. Molecular modelling was used to explore the interaction of the CD38 catalytic residue Glu-226 with the “northern” ribose. We propose that Glu226 still acts as the catalytic residue even for an L-sugar substrate. 8-Br-L-cIDPR potentially binds non-productively in an upside-down fashion. Results highlight the key role of the “northern” ribose in the interaction of cADPR with CD38

Direct activation of NADPH oxidase 2 by 2-deoxyribose-1-phosphate triggers nuclear factor kappa B-dependent angiogenesis.

AbstractAims: Deoxyribose-1-phosphate (dRP) is a proangiogenic paracrine stimulus released by cancer cells, platelets, and macrophages and acting on endothelial cells. The objective of this study was to clarify how dRP stimulates angiogenic responses in human endothelial cells.Results: Live cell imaging, electron paramagnetic resonance, pull-down of dRP-interacting proteins, followed by immunoblotting, gene silencing of different NADPH oxidases (NOXs), and their regulatory cosubunits by small interfering RNA (siRNA) transfection, and experiments with inhibitors of the sugar transporter glucose transporter 1 (GLUT1) were utilized to demonstrate that dRP acts intracellularly by directly activating the endothelial NOX2 complex, but not NOX4. Increased reactive oxygen species generation in response to NOX2 activity leads to redox-dependent activation of the transcription factor nuclear factor kappa B (NF-κB), which, in turn, induces vascular endothelial growth factor receptor 2 (VEGFR2) upregulation. Using endothelial tube formation assays, gene silencing by siRNA, and antibody-based receptor inhibition, we demonstrate that the activation of NF-κB and VEGFR2 is necessary for the angiogenic responses elicited by dRP. The upregulation of VEGFR2 and NOX2-dependent stimulation of angiogenesis by dRP were confirmed in excisional wound and Matrigel plug vascularization assays in vivo using NOX2−/− mice.Innovation: For the first time, we demonstrate that dRP acts intracellularly and stimulates superoxide anion generation by direct binding and activation of the NOX2 enzymatic complex.Conclusions: This study describes a novel molecular mechanism underlying the proangiogenic activity of dRP, which involves the sequential activation of NOX2 and NF-κB and upregulation of VEGFR2. Antioxid. Redox Signal. 28, 110–130

Genome-wide generation and systematic phenotyping of knockout mice reveals new roles for many genes.

Mutations in whole organisms are powerful ways of interrogating gene function in a realistic context. We describe a program, the Sanger Institute Mouse Genetics Project, that provides a step toward the aim of knocking out all genes and screening each line for a broad range of traits. We found that hitherto unpublished genes were as likely to reveal phenotypes as known genes, suggesting that novel genes represent a rich resource for investigating the molecular basis of disease. We found many unexpected phenotypes detected only because we screened for them, emphasizing the value of screening all mutants for a wide range of traits. Haploinsufficiency and pleiotropy were both surprisingly common. Forty-two percent of genes were essential for viability, and these were less likely to have a paralog and more likely to contribute to a protein complex than other genes. Phenotypic data and more than 900 mutants are openly available for further analysis. PAPERCLIP

Support for UNRWA's survival

The United Nations Relief and Works Agency for Palestine Refugees in the Near East (UNRWA) provides life-saving humanitarian aid for 5·4 million Palestine refugees now entering their eighth decade of statelessness and conflict. About a third of Palestine refugees still live in 58 recognised camps. UNRWA operates 702 schools and 144 health centres, some of which are affected by the ongoing humanitarian disasters in Syria and the Gaza Strip. It has dramatically reduced the prevalence of infectious diseases, mortality, and illiteracy. Its social services include rebuilding infrastructure and homes that have been destroyed by conflict and providing cash assistance and micro-finance loans for Palestinians whose rights are curtailed and who are denied the right of return to their homeland

Australasia

Observed changes and impacts Ongoing climate trends have exacerbated many extreme events (very high confidence). The Australian trends include further warming and sea level rise sea level rise (SLR), with more hot days and heatwaves, less snow, more rainfall in the north, less April–October rainfall in the southwest and southeast and more extreme fire weather days in the south and east. The New Zealand trends include further warming and sea level rise (SLR), more hot days and heatwaves, less snow, more rainfall in the south, less rainfall in the north and more extreme fire weather in the east. There have been fewer tropical cyclones and cold days in the region. Extreme events include Australia’s hottest and driest year in 2019 with a record-breaking number of days over 39°C, New Zealand’s hottest year in 2016, three widespread marine heatwaves during 2016–2020, Category 4 Cyclone Debbie in 2017, seven major hailstorms over eastern Australia and two over New Zealand from 2014–2020, three major floods in eastern Australia and three over New Zealand during 2019–2021 and major fires in southern and eastern Australia during 2019–2020

Repetitive Behavior in Rubinstein–Taybi Syndrome:Parallels with Autism Spectrum Phenomenology

Syndrome specific repetitive behavior profiles have been described previously. A detailed profile is absent for Rubinstein–Taybi syndrome (RTS). The Repetitive Behaviour Questionnaire and Social Communication Questionnaire were completed for children and adults with RTS (N = 87), Fragile-X (N = 196) and Down (N = 132) syndromes, and individuals reaching cut-off for autism spectrum disorder (N = 228). Total and matched group analyses were conducted. A phenotypic profile of repetitive behavior was found in RTS. The majority of behaviors in RTS were not associated with social-communication deficits or degree of disability. Repetitive behavior should be studied at a fine-grained level. A dissociation of the triad of impairments might be evident in RTS

The Polygenic and Monogenic Basis of Blood Traits and Diseases

Blood cells play essential roles in human health, underpinning physiological processes such as immunity, oxygen transport, and clotting, which when perturbed cause a significant global health burden. Here we integrate data from UK Biobank and a large-scale international collaborative effort, including data for 563,085 European ancestry participants, and discover 5,106 new genetic variants independently associated with 29 blood cell phenotypes covering a range of variation impacting hematopoiesis. We holistically characterize the genetic architecture of hematopoiesis, assess the relevance of the omnigenic model to blood cell phenotypes, delineate relevant hematopoietic cell states influenced by regulatory genetic variants and gene networks, identify novel splice-altering variants mediating the associations, and assess the polygenic prediction potential for blood traits and clinical disorders at the interface of complex and Mendelian genetics. These results show the power of large-scale blood cell trait GWAS to interrogate clinically meaningful variants across a wide allelic spectrum of human variation. Analysis of blood cell traits in the UK Biobank and other cohorts illuminates the full genetic architecture of hematopoietic phenotypes, with evidence supporting the omnigenic model for complex traits and linking polygenic burden with monogenic blood diseases