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Supplementary Figures file contains merged Figure S1 to S8 including description of cell lines, Human samples, PI3K inhibitor stability, SILAC setup and controls, STRING analysis, quantifications of WB and cellular assays, prediction of kinase motifs found regulated, effects of combination of treatment in vitro, correlation between cell survival and phosphopeptide intensity, effects of combination treatment in Human samples and in vivo.

Supplementary Table 1 contains the list of files deposited on PRIDE Proteome X Change under the number PXD008410.

This file contains the additional information for Material and method, including method for cell transduction, cell treatment and WB, RT-qPCR analysis, viablity assays, proliferation assays, DEVD cleavage assay, flow cytometry, soft agar assay, organoid and soft agar assay quantification and ImageJ plugin, scriot for analaysis of unpaired heavy/light phosphopeptides, method for SILAC labelling, SILAC sample preparation, data-dependent acquisition LC-MS/MS, phosphoproteomic data analysis, mice housing, details on statistics, as well as Legends of supplementary figures and tables, table with sequences of shRNAs, used antibodies, RT-qPCR primers.

Supplementary Table 3 contains the location of isoform selective phosphorylations in the protein.

Supplementary Table 2 contains the list of isoform selective peptides in each condition, with quantification and global pathway analysis.

Y avait-il des genres alternatifs chez les Anicinabek ? La question a émergé après qu’un centre de santé, désireux de lutter contre l’homophobie, eut voulu créer des outils culturellement adéquats, fondés sur la tradition anicinabe. En collaboration avec ce centre, les auteures ont effectué une recherche sur les sexes et les genres auprès de personnes reconnues comme porteuses de savoirs sur les temps anciens. S’aidant d’une revue de littérature sur les deux-esprits chez les Premières Nations en général, elles ont noté que rien en anicinabemowin ne traduit l’existence d’autres genres que homme/femme et d’autres pratiques qu’hétérosexuelles. La sexualité était un domaine caché et les gens étaient pudiques. L’importance de la discrétion, dans un contexte où la reproduction était capitale, a fait ressortir les valeurs qui sont fondamentales pour les Anicinabek, comme le respect et la non-ingérence. Ainsi, être un « bon homme » et une « bonne femme » semble plus important que les préférences amoureuses et sexuelles de chacun., Were there alternative genders among the Anicinabek? The question emerged after a health centre, wanting to fight homophobia, sought to develop culturally appropriate tools based on the Anicinabe tradition. In collaboration with this centre, we conducted a study on gender and sex with people who are recognized as bearers of traditional knowledge. With the help of a literature review on two-spiritedness among First Nations in general, we noted that nothing in Anicinabemowin shows the existence of genders other than male/female and practices other than heterosexual. Sexuality was a hidden domain and people were modest. The importance of discretion, in a context where reproduction was paramount, highlighted values that are fundamental to Anicinabek, such as respect and non-interference. Thus, being a “good man” and a “good woman” appears to be more important than one’s sexual or romantic preferences., ¿Existían géneros alternativos entre los Anicinabek? La cuestión surgió después de que un centro de salud, que deseaba luchar contra la homofobia, quisiera crear herramientas culturalmente apropiadas basadas en la tradición anicinabe. En colaboración con este centro, las autoras llevaron a cabo una investigación sobre género y sexo con personas reconocidas como portadoras de conocimientos ancestrales. A partir de una revisión de la bibliografía sobre la dualidad de espíritus entre las Primeras Naciones en general, observaron que nada en anicinabemowin refleja la existencia de géneros distintos al masculino/femenino y de prácticas distintas a la heterosexual. La sexualidad era un área oculta y la gente era púdica. La importancia de la discreción, en un contexto en el que la reproducción era primordial, puso de manifiesto valores fundamentales para los Anicinabek, como el respeto y la no injerencia. Así, ser un “buen hombre” y una “buena mujer” parece ser más importante que las propias preferencias amorosas y sexuales.

Statistical modeling of a screw press was established by using an experimental design based on the screw rotational speed, the pulp feed consistency, the pulp feed suspension freeness, the inlet pressure, and the counter-pressure at the discharge end. The statistical models showed that the screw press outputs for each pulp could be predicted. When including all data in a global model to predict the outputs of the press for any pulp, a global statistical model was found not to be efficient by using just the five fixed parameters. The solution to this problem was to use a multivariate analysis to include more parameters, mainly about the fiber characteristics (crowding factor, fiber length, fiber width, and fines content). By including these fiber properties, the differences between each pulp were more properly analyzed. The multivariate analysis predicted the press outsets very well in a global model by using eight parameters instead of five. The R2 values of the multivariate prediction model were all higher than 0.70 and had the goodness of prediction (Q2¬¬¬) higher than 0.60.

Proteasomes are major non-lysosomal proteolytic complexes localized in the cytoplasm and in the nucleus of eukaryotic cells. Strikingly, high levels of extracellular proteasome have also been evidenced in the plasma (p-proteasome) of patients with specific diseases. Here, we examined the process by which proteasomes are secreted, as well as their structural and functional features once in the extracellular space. We demonstrate that assembled 20S core particles are secreted by cells within microvesicles budding from the plasma membrane. Part of the extracellular proteasome pool is also free of membranes in the supernatant of cultured cells, and likely originates from microvesicles leakage. We further demonstrate that this free proteasome released by cells (cc-proteasome for cell culture proteasome) possesses latent proteolytic activity and can degrade various extracellular proteins. Both standard (no immune-subunits) and intermediate (containing some immune-subunits) forms of 20S are observed. Moreover, we show that galectin-3, which displays a highly disordered N-terminal region, is efficiently cleaved by purified cc-proteasome, without SDS activation, likely after its binding to PSMA3 (α7) subunit through its intrinsically disordered region. As a consequence, galectin-3 is unable to induce red blood cells agglutination when preincubated with cc-proteasome. These results highlight potential novel physio- and pathologic functions for the extracellular proteasome.

Mycobacterium abscessus(Mabs) drives life-shortening mortality in cystic fibrosis (CF) patients, primarily because of its resistance to chemotherapeutic agents. To date, our knowledge on the host and bacterial determinants driving Mabs pathology in CF patient lung remains rudimentary. Here, we used human airway organoids (AOs) microinjected with smooth (S) or rough (R-)Mabs to evaluate bacteria fitness, host responses to infection, and new treatment efficacy. We show that S Mabs formed biofilm, R Mabs formed cord serpentines and displayed a higher virulence. While Mabs infection triggers enhanced oxidative stress, pharmacological activation of antioxidant pathways resulted in better control of Mabs growth. Genetic and pharmacological inhibition of the CFTR is associated with better growth and higher virulence of S and R Mabs. Finally, pharmacological activation of antioxidant pathways inhibited Mabs growth and improved efficacy in combination with cefoxitin, a first line antibiotic. In conclusion, we have established AOs as a suitable human system to decipher mechanisms of CF-driven respiratory infection by Mabs and propose antioxidants as a potential host-directed strategy to improve Mabs infection control.

During spermatogenesis, spermatogonia undergo a series of mitotic and meiotic divisions on their path to spermatozoa. To achieve this, a succession of complex processes requiring high proteolytic activity are in part orchestrated by the proteasome. The spermatoproteasome (s20S) is a proteasome subtype specific to the developing gametes, in which the gamete-specific α4s subunit replaces the α4 isoform found in the constitutive proteasome (c20S). Although the s20S is conserved across species and was shown to be crucial for germ cell development, its mechanism, function and structure remain incompletely characterized. Here, we used advanced mass spectrometry (MS) methods to map the composition of proteasome complexes and their interactomes throughout spermatogenesis. We observed that the s20S becomes highly activated as germ cells enter meiosis, mainly through a particularly extensive 19S activation and, to a lesser extent, PA200 binding. Additionally, the proteasome population shifts from predominantly c20S (98%) to predominantly s20S (>82-92%) during differentiation, presumably due to the shift from α4 to α4s expression. We demonstrated that s20S, but not c20S, interacts with components of the meiotic synaptonemal complex, where it may localize via association with the PI31 adaptor protein. In vitro, s20S preferentially binds to 19S, and displays higher trypsin- and chymotrypsin-like activities, both with and without PA200 activation. Moreover, using MS methods to monitor protein dynamics, we identified significant differences in domain flexibility between α4 and α4s. We propose that these differences induced by α4s incorporation result in significant changes in the way the s20S interacts with its partners, and dictate its role in germ cell differentiation.

La recherche ethnologique et ethnolinguistique peut poser un large éventail de défis éthiques différents pour les chercheurs et les communautés dans lesquelles ils travaillent. Les ethnologues et les ethnolinguistes étudient les sociétés humaines sous différents angles pour comprendre leurs caractères culturels, leurs variabilités linguistiques, etc. Ces recherches peuvent faire progresser la compréhension que l’on a des groupes sociaux et contribuer à la production de connaissances, et peuvent même parfois être bénéfiques pour certaines communautés. Mais elle peut aussi créer des risques pour les participants et les communautés (par exemple perte de la vie privée, stigmatisation, persécution), et pour les chercheurs eux-mêmes (par exemple perte d’accès à une zone de terrain, perte de contrôle du processus et des résultats de la recherche). Ces risques et autres défis éthiques, ainsi que les moyens d’y faire face, méritent une attention particulière de la part des ethnologues et de l’ethnolinguistique., Ethnological and ethnolinguistic research can pose a wide range of different ethical challenges for researchers and the communities in which they work. Ethnologists and ethnolinguists study human societies from different angles to understand their cultural traits, linguistic variability, etc. Such research can advance our understanding of social groups and contribute to the production of knowledge and may even benefit certain communities. But it can also create risks for participants and communities (e.g., loss of privacy, stigmatization, persecution), and for researchers themselves (e.g., loss of access to a field area, loss of control over the research process and results). These risks and other ethical challenges, and the means to address them, deserve special attention from ethnologists and ethnolinguistics.

The drainage- and dewatering-controlling mechanisms in a screw press were detailed in this work. Three pulps (kraft pulp, bleached chemi-thermomechanical pulp, and thermomechanical pulp) were studied to compare a wide range of wood fibre types. The dewatering was controlled by the screw press parameters and the pulps’ properties. Filtration was found to be the controlling mechanism in the first part of the screw press for the three pulps, and it was less important when the fines content was greater. In the compression zone, the degree of compression was affected by the pulp flexibility and the fibres’ tendency to entangle. Filtrate flow rate monitoring along the screw press could be a good indicator of where the transition from filtration to consolidation occurs. The pressure along the screw press did not change much in the filtration zone, and it notably increased near the discharge end. When the drainage was very high, the pulp feed increased, causing the pulp axial velocity in the end part to be greater than the screw’s linear advance.

A Thune SP23 screw press dewatering parameters were studied. The dewatering efficiency was affected more by the rotational speed and the pulp properties. The counter-pressure affects dewatering near the discharge end, and it was observed to influence the outlet consistency and filtrate flow rate of Kraft, which has much longer fibres and fewer fines compared to TMP and BCTMP. The feed stock freeness and consistency are very important variables to consider in the screw press performance. The freeness reflects the degree of drainage, which is an important parameter to consider when optimising the screw press, while the feed consistency is a parameter of the fibre-fibre contact degree. The pulp properties, especially the fines content and fibre flexibility are also two very important parameters that affect the screw press performance. This study was to provide an insight of the screw press performance and to show the complex effect of the operational parameters on the dewatering characteristics. Using three different pulps, Kraft and TMP softwood fibres and a BCTMP hardwood fibres, we have shown that the fines content and fibre properties are two dominant properties that should be highly considered when operating a screw press.

20S proteasomes are very heterogeneous protein complexes involved in many cellular processes. In the present study, we combined an MRM-based assay with the production and purification of entire SILAC labelled proteasome to monitor absolute quantities of the different 20S proteasome subtypes in various human cells and tissues. This method applied to adipocyte-derived stem cells (ADSCs) amplified under various conditions highlights an increased expression of immunoproteasome when this type of cell is primed with IFNγ or amplified in a 20% O2 environment., Graphical Abstract Highlights Design of an MRM assay to determine the absolute quantity and stoichiometry of ubiquitous and tissue-specific human 20S proteasome subtypes. Use of purified isotopically labelled 20S proteasome as internal standard for accurate quantification. Variation in the expression of immunoproteasome in adipocyte-derived stem cells (ADSCs) grown under different O2 levels might be causal for change in cells differentiation capacity. The status of 20S proteasome during ADSCs expansion might constitute an additional relevant quality control parameter to contribute to predict, among other quality markers, their therapeutic capacity., The proteasome controls a multitude of cellular processes through protein degradation and has been identified as a therapeutic target in oncology. However, our understanding of its function and the development of specific modulators are hampered by the lack of a straightforward method to determine the overall proteasome status in biological samples. Here, we present a method to determine the absolute quantity and stoichiometry of ubiquitous and tissue-specific human 20S proteasome subtypes based on a robust, absolute SILAC-based multiplexed LC-Selected Reaction Monitoring (SRM) quantitative mass spectrometry assay with high precision, accuracy, and sensitivity. The method was initially optimized and validated by comparison with a reference ELISA assay and by analyzing the dynamics of catalytic subunits in HeLa cells following IFNγ-treatment and in range of human tissues. It was then successfully applied to reveal IFNγ- and O2-dependent variations of proteasome status during primary culture of Adipose-derived-mesenchymal Stromal/Stem Cells (ADSCs). The results show the critical importance of controlling the culture conditions during cell expansion for future therapeutic use in humans. We hypothesize that a shift from the standard proteasome to the immunoproteasome could serve as a predictor of immunosuppressive and differentiation capacities of ADSCs and, consequently, that quality control should include proteasomal quantification in addition to examining other essential cell parameters. The method presented also provides a new powerful tool to conduct more individualized protocols in cancer or inflammatory diseases where selective inhibition of the immunoproteasome has been shown to reduce side effects.

The PI3K pathway is highly active in human cancers. The four class I isoforms of PI3K are activated by distinct mechanisms leading to a common downstream signaling. Their downstream redundancy is thought to be responsible for treatment failures of PI3K inhibitors. We challenged this concept, by mapping the differential phosphoproteome evolution in response to PI3K inhibitors with different isoform selectivity patterns in pancreatic cancer, a disease currently without effective therapy. In this cancer, the PI3K signal was shown to control cell proliferation. We compared the effects of LY294002 that inhibit with equal potency all class I isoenzymes and downstream mTOR with the action of inhibitors with higher isoform-selectivity towards PI3Kα, PI3Kβ or PI3Kγ (namely A66, TGX-221 and AS-252424). A bioinformatics global pathway analysis of phosphoproteomics data allowed us to identify common and specific signals activated by PI3K inhibitors supported by the biological data. AS-252424 was the most effective treatment and induced apoptotic pathway activation as well as the highest changes in global phosphorylation-regulated cell signal. However, AS-252424 treatment induced re-activation of Akt, therefore decreasing the treatment outcome on cell survival. Reversely, AS-252424 and A66 combination treatment prevented p-Akt reactivation and led to synergistic action in cell lines and patient organoids. The combination of clinically approved α-selective BYL-719 with γ-selective IPI-549 was more efficient than single molecule treatment on xenograft growth. Mapping unique adaptive signaling responses to isoform-selective PI3K inhibition will help to design better combinative treatments that prevent the induction of selective compensatory signals.Graphical abstractSignificanceHalf of all human cancers show increased PI3K activity but the mutational pattern of the PI3K pathway is not sufficient to predict sensitivity to PI3K inhibitors. By identifying for the first time specific signaling induced by PI3K inhibitors with different isoform-selectivity patterns, we provide insight in how to handle heterogeneity of PI3K expression in tumor samples for the choice of available PI3K-targetting drugs. Our work provides a roadmap to target the PI3K pathway, balancing the use of isoform-selective PI3K inhibitors with personalized information extending beyond the specific mutational status.

We consider a quasilinear degenerate parabolic equation driven by the orthotropic $p-$Laplacian. We prove that local weak solutions are locally Lipschitz continuous in the spatial variable, uniformly in time., 29 pages

The proteasome is responsible for selective degradation of proteins. It exists in mammalian cells under four main subtypes, which differ by the combination of their catalytic subunits: the standard proteasome (β1–β2–β5), the immunoproteasome (β1i–β2i–β5i) and the two intermediate proteasomes (β1–β2–β5i and β1i–β2–β5i). The efficiency of the four proteasome subtypes to degrade ubiquitinated or oxidized proteins remains unclear. Using cells expressing exclusively one proteasome subtype, we observed that ubiquitinated p21 and c-­myc were degraded at similar rates, indicating that the four 26S proteasomes degrade ubiquitinated proteins equally well. Under oxidative stress, we observed a partial dissociation of 26S into 20S proteasomes, which can degrade non-ubiquitinated oxidized proteins. Oxidized calmodulin and hemoglobin were best degraded in vitro by the three β5i-containing 20S proteasomes, while their native forms were not degraded. Circular dichroism analyses indicated that ubiquitin-independent recognition of oxidized proteins by 20S proteasomes was triggered by the disruption of their structure. Accordingly, β5i-containing 20S proteasomes degraded unoxidized naturally disordered protein tau, while 26S proteasomes did not. Our results suggest that the three β5i-containing 20S proteasomes, namely the immunoproteasome and the two intermediate proteasomes, might help cells to eliminate proteins containing disordered domains, including those induced by oxidative stress.

Background & Aims Cancer-associated fibroblasts (CAFs) from pancreatic adenocarcinoma (PDA) present high protein synthesis rates. CAFs express the G-protein–coupled somatostatin receptor sst1. The sst1 agonist SOM230 blocks CAF protumoral features in vitro and in immunocompromised mice. We have explored here the therapeutic potential of SOM230, and underlying mechanisms, in immunocompetent models of murine PDA mimicking the heavy fibrotic and immunosuppressive stroma observed in patient tumors. Methods Large-scale mass spectrometry analyses were performed on media conditioned from 9 patient PDA-derived CAF primary cultures. Spontaneous transgenic and experimental (orthotopic co-graft of tumor cells plus CAFs) PDA-bearing mice were longitudinally ultrasound-monitored for tumor and metastatic progression. Histopathology and flow cytometry analyses were performed on primary tumors and metastases. Stromal signatures were functionally validated through bioinformatics using several published, and 1 original, PDA database. Results Proteomics on the CAF secretome showed that SOM230 controls stromal activities including inflammatory responses. Among the identified secreted proteins, we validated that colony-stimulating factor 1 (CSF-1) (a macrophage growth factor) was reduced by SOM230 in the tumor and plasma of PDA-harboring mice, alongside intratumor stromal normalization (reduced CAF and macrophage activities), and dramatic metastasis reduction. In transgenic mice, these SOM230 benefits alleviate the chemotherapy-induced (gemcitabine) immunosuppressive stroma reshaping. Mechanistically, SOM230 acts in vivo on CAFs through sst1 to disrupt prometastatic CAF production of CSF-1 and cross-talk with macrophages. We found that in patients, stromal CSF-1 was associated with aggressive PDA forms. Conclusions We propose SOM230 as an antimetastatic therapy in PDA for its capacity to remodel the fibrotic and immunosuppressive myeloid stroma. This pharmacotherapy should benefit PDA patients treated with chemotherapies., Graphical abstract

Proinsulin is a misfolding-prone protein, and its efficient breakdown is critical when β-cells are confronted with high-insulin biosynthetic demands, to prevent endoplasmic reticulum stress, a key trigger of secretory dysfunction and, if uncompensated, apoptosis. Proinsulin degradation is thought to be performed by the constitutively expressed standard proteasome, while the roles of other proteasomes are unknown. We recently demonstrated that deficiency of the proinsulin chaperone glucose-regulated protein 94 (GRP94) causes impaired proinsulin handling and defective insulin secretion associated with a compensated endoplasmic reticulum stress response. Taking advantage of this model of restricted folding capacity, we investigated the role of different proteasomes in proinsulin degradation, reasoning that insulin secretory dynamics require an inducible protein degradation system. We show that the expression of only one enzymatically active proteasome subunit, namely, the inducible β5i-subunit, was increased in GRP94 CRISPR/Cas9 knockout (KO) cells. Additionally, the level of β5i-containing intermediate proteasomes was significantly increased in these cells, as was β5i-related chymotrypsin-like activity. Moreover, proinsulin levels were restored in GRP94 KO upon β5i small interfering RNA-mediated knockdown. Finally, the fraction of β-cells expressing the β5i-subunit is increased in human islets from type 2 diabetes patients. We conclude that β5i is an inducible proteasome subunit dedicated to the degradation of mishandled proinsulin.

The interaction of insulin-degrading enzyme (IDE) with the main intracellular proteasome assemblies (i.e, 30S, 26S and 20S) was analyzed by enzymatic activity, mass spectrometry and native gel electrophoresis. IDE was mainly detected in association with assemblies with at least one free 20S end and biochemical investigations suggest that IDE competes with the 19S in vitro. IDE directly binds the 20S and affects its proteolytic activities in a bimodal fashion, very similar in human and yeast 20S, inhibiting at (IDE) 30 nM and activating at (IDE) 30 nM. Only an activating effect is observed in a yeast mutant locked in the "open" conformation (i.e., the alpha-3 Delta N 20S), envisaging a possible role of IDE as modulator of the 20S "open"-"closed" allosteric equilibrium. Protein-protein docking in silico proposes that the interaction between IDE and the 20S could involve the C-term helix of the 20S alpha-3 subunit which regulates the gate opening of the 20S.

Given a compact manifold $N^n subset mathbb{R}^u$ and real numbers $s ge 1$ and $1 le p < infty$, we prove that the class $C^infty(overline{Q}^m; N^n)$ of smooth maps on the cube with values into $N^n$ is strongly dense in the fractional Sobolev space $W^{s, p}(Q^m; N^n)$ when $N^n$ is $lfloor sp floor$ simply connected. For $sp$ integer, we prove weak sequential density of $C^infty(overline{Q}^m; N^n)$ when $N^n$ is $sp - 1$ simply connected. The proofs are based on the existence of a retraction of $mathbb{^ u$ onto $N^n$ except for a small subset of $N^n$ and on a pointwise estimate of fractional derivatives of composition of maps in $W^{s, p} cap W^{1, sp}$.

International audience; Protein-protein interactions play a major role in the molecular machinery of life, and various techniques such as AP-MS are dedicated to their identification. However, those techniques return lists of proteins devoid of organizational structure, not detailing which proteins interact with which others. Proposing a hierarchical view of the interactions between the members of the flat list becomes highly tedious for large data sets when done by hand. To help hierarchize this data, we introduce a new bioinformatics protocol that integrates information of the multimeric protein 3D structures available in the Protein Data Bank using remote homology detection, as well as information related to Short Linear Motifs and interaction data from the BioGRID. We illustrate on two unrelated use-cases of different complexity how our approach can be useful to decipher the network of interactions hidden in the list of input proteins, and how it provides added value compared to state-of-the-art resources such as Interactome3D or STRING. Particularly, we show the added value of using homology detection to distinguish between orthologs and paralogs, and to distinguish between core obligate and more facultative interactions. We also demonstrate the potential of considering interactions occurring through Short Linear Motifs.

International audience; A central and still open question regarding the pathogenesis of autoimmune diseases, such as type 1 diabetes, concerns the processes that underlie the generation of MHC-presented autoantigenic epitopes that become targets of autoimmune attack. Proteasomal degradation is a key step in processing of proteins for MHC class I presentation. Different types of proteasomes can be expressed in cells dictating the repertoire of peptides presented by the MHC class I complex. Of particular interest for type 1 diabetes is the proteasomal configuration of pancreatic β cells, as this might facilitate autoantigen presentation by β cells and thereby their T-cell mediated destruction. Here we investigated whether so-called inducible subunits of the proteasome are constitutively expressed in β cells, regulated by inflammatory signals and participate in the formation of active intermediate or immuno-proteasomes. We show that inducible proteasomal subunits are constitutively expressed in human and rodent islets and an insulin-secreting cell-line. Moreover, the β5i subunit is incorporated into active intermediate proteasomes that are bound to 19S or 11S regulatory particles. Finally, inducible subunit expression along with increase in total proteasome activities are further upregulated by low concentrations of IL-1β stimulating proinsulin biosynthesis. These findings suggest that the β cell proteasomal repertoire is more diverse than assumed previously and may be highly responsive to a local inflammatory islet environment.

A central and still open question regarding the pathogenesis of autoimmune diseases, such as type 1 diabetes, concerns the processes that underlie the generation of MHC-presented autoantigenic epitopes that become targets of autoimmune attack. Proteasomal degradation is a key step in processing of proteins for MHC class I presentation. Different types of proteasomes can be expressed in cells dictating the repertoire of peptides presented by the MHC class I complex. Of particular interest for type 1 diabetes is the proteasomal configuration of pancreatic β cells, as this might facilitate autoantigen presentation by β cells and thereby their T-cell mediated destruction. Here we investigated whether so-called inducible subunits of the proteasome are constitutively expressed in β cells, regulated by inflammatory signals and participate in the formation of active intermediate or immuno-proteasomes.We show that inducible proteasomal subunits are constitutively expressed in human and rodent islets and an insulin-secreting cell-line. Moreover, the β5i subunit is incorporated into active intermediate proteasomes that are bound to 19S or 11S regulatory particles. Finally, inducible subunit expression along with increase in total proteasome activities are further upregulated by non-toxic concentrations of IL-1β stimulating proinsulin biosynthesis. These findings suggest that the β cell proteasomal repertoire is more diverse than assumed previously and may be highly responsive to a local inflammatory islet environment.

International audience; The rise of intact protein analysis by mass spectrometry (MS) was accompanied by an increasing need for flexible tools allowing data visualization and analysis. These include inspection of the deconvoluted molecular weights of the proteoforms eluted alongside liquid chromatography (LC) through their representation in three-dimensional (3D) liquid chromatography coupled to mass spectrometry (LC-MS) maps (plots of deconvoluted molecular weights, retention times, and intensity of the MS signal). With this aim, we developed a free and open-source web application named VisioProt-MS (https://masstools.ipbs.fr/mstools/visioprot-ms/). VisioProt-MS is highly compatible with many algorithms and software developed by the community to integrate and deconvolute top-down and intact protein MS data. Its dynamic and userfriendly features greatly facilitate analysis through several graphical representations dedicated to MS and tandem mass spectrometry (MS/MS) analysis of proteoforms in complex samples. Here, we will illustrate the importance of LC-MS map visualization to optimize top-down acquisition/ search parameters and analyze intact protein MS data. We will go through the main features of VisioProt-MS using the human proteasomal 20S core particle as a user-case.