Integrative omics reveals subtle molecular perturbations following ischemic conditioning in a porcine kidney transplant model

BACKGROUND: Remote Ischemic Conditioning (RIC) has been proposed as a therapeutic intervention to circumvent the ischemia/reperfusion injury (IRI) that is inherent to organ transplantation. Using a porcine kidney transplant model, we aimed to decipher the subclinical molecular effects of a RIC regime, compared to non-RIC controls. METHODS: Kidney pairs (n = 8 + 8) were extracted from brain dead donor pigs and transplanted in juvenile recipient pigs following a period of cold ischemia. One of the two kidney recipients in each pair was subjected to RIC prior to kidney graft reperfusion, while the other served as non-RIC control. We designed an integrative Omics strategy combining transcriptomics, proteomics, and phosphoproteomics to deduce molecular signatures in kidney tissue that could be attributed to RIC. RESULTS: In kidney grafts taken out 10 h after transplantation we detected minimal molecular perturbations following RIC compared to non-RIC at the transcriptome level, which was mirrored at the proteome level. In particular, we noted that RIC resulted in suppression of tissue inflammatory profiles. Furthermore, an accumulation of muscle extracellular matrix assembly proteins in kidney tissues was detected at the protein level, which may be in response to muscle tissue damage and/or fibrosis. However, the majority of these protein changes did not reach significance (p < 0.05). CONCLUSIONS: Our data identifies subtle molecular phenotypes in porcine kidneys following RIC, and this knowledge could potentially aid optimization of remote ischemic conditioning protocols in renal transplantation. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12014-022-09343-3

USP16 is an ISG15 cross-reactive deubiquitinase targeting a subset of metabolic pathway-related proteins

The ubiquitin-like modifier ISG15 can modulate host and viral proteins to restrict viral and microbial infections, and act as a cytokine. Its expression and conjugation are strongly up-regulated by type I interferons. Here we identify the deubiquitinating enzyme USP16 as an ISG15 cross-reactive protease. Ubiquitin-specific protease 16 (USP16) was found to react with an ISG15 activity-based probe in pull-down experiments using chronic myeloid leukaemia-derived human cells (HAP1). Supporting this finding, recombinant USP16 cleaved pro-ISG15 and ISG15 iso-peptide linked model substrates in vitro, as well as ISGylated substrates present in cell lysates. Moreover, the interferon-induced stimulation of ISGylation in human HAP1 cells was increased by knockdown or knockout of USP16. Depletion of USP16 did not affect interferon signaling, and interferon treatment did not affect USP16 expression or enzymatic activity either. A USP16-dependent ISG15 interactome was established by anti-ISG15 immunoprecipitation mass spectrometry (IP-MS), which indicated that the deISGylating function of USP16 may regulate metabolic pathways involving GOT1, ALDOA, SOD1 and MDH1, all of which were further confirmed to be deISGylated by USP16 in HEK293T cells. Together, our results indicate that USP16 may contribute to regulating the ISGylation status of a subset of proteins related to metabolism during type I interferon responses

Constraining the Evolution of Zz Ceti

We report our analysis of the stability of pulsation periods in the DAV star (pulsating hydrogen atmosphere white dwarf) ZZ Ceti, also called R548. On the basis of observations that span 31 years, we conclude that the period 213.13 s observed in ZZ Ceti drifts at a rate dP/dt ≤ (5:5 ± 1:9) x 10-15 s s-1, after correcting for proper motion. Our results are consistent with previous Ṗ values for this mode and an improvement over them because of the larger time base. The characteristic stability timescale implied for the pulsation period is ⎸P / Ṗ ⎸=⎹≥ 1:2 Gyr, comparable to the theoretical cooling timescale for the star. Our current stability limit for the period 213.13 s is only slightly less than the present measurement for another DAV, G117-B15A, for the period 215.2 s, establishing this mode in ZZ Ceti as the second most stable optical clock known, comparable to atomic clocks and more stable than most pulsars. Constraining the cooling rate of ZZ Ceti aids theoretical evolutionary models and white dwarf cosmochronology. The drift rate of this clock is small enough that we can set interesting limits on reflex motion due to planetary companions

A blood atlas of COVID-19 defines hallmarks of disease severity and specificity.

Treatment of severe COVID-19 is currently limited by clinical heterogeneity and incomplete description of specific immune biomarkers. We present here a comprehensive multi-omic blood atlas for patients with varying COVID-19 severity in an integrated comparison with influenza and sepsis patients versus healthy volunteers. We identify immune signatures and correlates of host response. Hallmarks of disease severity involved cells, their inflammatory mediators and networks, including progenitor cells and specific myeloid and lymphocyte subsets, features of the immune repertoire, acute phase response, metabolism, and coagulation. Persisting immune activation involving AP-1/p38MAPK was a specific feature of COVID-19. The plasma proteome enabled sub-phenotyping into patient clusters, predictive of severity and outcome. Systems-based integrative analyses including tensor and matrix decomposition of all modalities revealed feature groupings linked with severity and specificity compared to influenza and sepsis. Our approach and blood atlas will support future drug development, clinical trial design, and personalized medicine approaches for COVID-19

MEMHDX: An interactive tool to expedite the statistical validation and visualization of large HDX-MS datasets

International audienceMotivation: With the continued improvement of requisite mass spectrometers and UHPLC systems, Hydrogen/Deuterium eXchange Mass Spectrometry (HDX-MS) workflows are rapidly evolving towards the investigation of more challenging biological systems, including large protein complexes and membrane proteins. The analysis of such extensive systems results in very large HDX-MS datasets for which specific analysis tools are required to speed up data validation and interpretation. Results: We introduce a web application and a new R-package named " MEMHDX " to help users analyze, validate and visualize large HDX-MS datasets. MEMHDX is composed of two elements. A statistical tool aids in the validation of the results by applying a mixed-effects model for each peptide, in each experimental condition, and at each time point, taking into account the time dependency of the HDX reaction and number of independent replicates. Two adjusted p-values are generated per peptide, one for the " Change in dynamics " and one for the " Magnitude of ∆D " , and are used to classify the data by means of a " Logit " representation. A user-friendly interface developed with Shiny by RStudio facilitates the use of the package. This interactive tool allows the user to easily and rapidly validate, visualize and compare the relative deuterium incorporation on the amino acid sequence and 3D structure, providing both spatial and temporal information. Availability: MEMHDX is freely available as a web tool at the project home page http://memhdx

Quantitative proteomics of pancreatic cyst fluid for early diagnosis of cancer.

In our EARLY DIAgnosis of PAncreatic Cancer (EARLY DIAPAC) study, we aim to analyse molecular changes or “signatures” associated with early stages of pancreatic carcinogenesis by using the combination of: (i) Label-Free Quantitative Proteomics (LFQP), (ii) Whole Genome Sequencing (WGS) and (iii) High Performance Computer (HPC) VIPER supercomputing capabilities. Building on the large volume of proteomics and genomics work acquired within the field of pancreatic cancer to-date, we focus on exploring the challenging concept of integrating these state-of-the-art platform technologies to conduct molecular analysis of pancreatic cystic fluid (PCyF, as a liquid biopsy) on unprecedented, global, or ‘multi-omics’ (proteogenomics) scale. This innovative approach of characterising PCyF at whole genome and proteome resolution levels in a single diagnostic test would allow for an unbiased profiling of the molecular events associated with early stages of pancreatic carcinogenesis. We have assembled a Multidisciplinary EARLY DIAPAC Consortium across the UK (Hull-Cambridge-Glasgow-Oxford) to conduct multi-omics data analysis of PCyF samples. Proof-of-principle pilot data has been acquired in the first instance. We compared findings from this dataset to confirmed malignant cases and a range of dysplasias to determine the potential of utilising the combination of these state-of-the-art platform technologies for early detection/diagnosis and clinical classification in an area of significant clinical need. Our LFQP and WGS data showed that the differences in PCyF from benign, pre-malignant, and malignant lesions are detectable and quantifiable. Our findings demonstrate that integrative Omics approach has the potential to distinguish molecular “signatures” of PCyF associated with different disease states involving formation of cysts and to advance the field of early detection of pancreatic cancer. The most recent updates from quantitative proteomics part of the EARLY DIAPAC study will be presented at the forum

Calmodulin fishing with a structurally disordered bait triggers CyaA catalysis.

Once translocated into the cytosol of target cells, the catalytic domain (AC) of the adenylate cyclase toxin (CyaA), a major virulence factor of Bordetella pertussis, is potently activated by binding calmodulin (CaM) to produce supraphysiological levels of cAMP, inducing cell death. Using a combination of small-angle X-ray scattering (SAXS), hydrogen/deuterium exchange mass spectrometry (HDX-MS), and synchrotron radiation circular dichroism (SR-CD), we show that, in the absence of CaM, AC exhibits significant structural disorder, and a 75-residue-long stretch within AC undergoes a disorder-to-order transition upon CaM binding. Beyond this local folding, CaM binding induces long-range allosteric effects that stabilize the distant catalytic site, whilst preserving catalytic loop flexibility. We propose that the high enzymatic activity of AC is due to a tight balance between the CaM-induced decrease of structural flexibility around the catalytic site and the preservation of catalytic loop flexibility, allowing for fast substrate binding and product release. The CaM-induced dampening of AC conformational disorder is likely relevant to other CaM-activated enzymes

Disorder-To-Order Transitions and Post-Translational Acylation Control the Folding and Activity of the Bordetella Pertussis CyaA Toxin

Place: Cambridge Publisher: Cell Press WOS:000513023201234Communication à la conférence : 64th Annual Meeting of the Biophysical-Society [San Diego, CA]Date: FEB 15-19, 2020International audienc